Lean thinking transformation of the unsedated upper gastrointestinal endoscopy pathway improves efficiency and is associated with high levels of patient satisfaction.

PubMed

Hydes, Theresa; Hansi, Navjyot; Trebble, Timothy M

2012-01-01

Upper gastrointestinal (UGI) endoscopy is a routine healthcare procedure with a defined patient pathway. The objective of this study was to redesign this pathway for unsedated patients using lean thinking transformation to focus on patient-derived value-adding steps, remove waste and create a more efficient process. This was to form the basis of a pathway template that was transferrable to other endoscopy units. A literature search of patient expectations for UGI endoscopy identified patient-derived value. A value stream map was created of the current pathway. The minimum and maximum time per step, bottlenecks and staff-staff interactions were recorded. This information was used for service transformation using lean thinking. A patient pathway template was created and implemented into a secondary unit. Questionnaire studies were performed to assess patient satisfaction. In the primary unit the patient pathway reduced from 19 to 11 steps with a reduction in the maximum lead time from 375 to 80 min following lean thinking transformation. The minimum value/lead time ratio increased from 24% to 49%. The patient pathway was redesigned as a 'cellular' system with minimised patient and staff travelling distances, waiting times, paperwork and handoffs. Nursing staff requirements reduced by 25%. Patient-prioritised aspects of care were emphasised with increased patient-endoscopist interaction time. The template was successfully introduced into a second unit with an overall positive patient satisfaction rating of 95%. Lean thinking transformation of the unsedated UGI endoscopy pathway results in reduced waiting times, reduced staffing requirements and improved patient flow and can form the basis of a pathway template which may be successfully transferred into alternative endoscopy environments with high levels of patient satisfaction.

A three-tiered approach for linking pharmacokinetic ...

EPA Pesticide Factsheets

The power of the adverse outcome pathway (AOP) framework arises from its utilization of pathway-based data to describe the initial interaction of a chemical with a molecular target (molecular initiating event; (MIE), followed by a progression through a series of key events that lead to an adverse outcome relevant for regulatory purposes. The AOP itself is not chemical specific, thus providing the biological context necessary for interpreting high throughput (HT) toxicity screening results. Application of the AOP framework and HT predictions in ecological and human health risk assessment, however, requires the consideration of chemical-specific properties that influence external exposure doses and target tissue doses. To address this requirement, a three-tiered approach was developed to provide a workflow for connecting biology-based AOPs to biochemical-based pharmacokinetic properties (absorption, distribution, metabolism, excretion; ADME), and then to chemical/human activity-based exposure pathways. This approach included: (1) The power of the adverse outcome pathway (AOP) framework arisesfrom its utilization of pathway-based data to describe the initial interaction of a chemical with a molecular target (molecular initiating event; (MIE), followed by a progression through a series of key events that lead to an adverse outcome relevant for regulatory purposes. The AOP itself is not chemical specific, thus providing the biological context necessary for interpreti

Analysis of protein interactions within the cytokinin-signaling pathway of Arabidopsis thaliana.

PubMed

Dortay, Hakan; Mehnert, Nijuscha; Bürkle, Lukas; Schmülling, Thomas; Heyl, Alexander

2006-10-01

The signal of the plant hormone cytokinin is perceived by membrane-located sensor histidine kinases and transduced by other members of the plant two-component system. In Arabidopsis thaliana, 28 two-component system proteins (phosphotransmitters and response regulators) act downstream of three receptors, transmitting the signal from the membrane to the nucleus and modulating the cellular response. Although the principal signaling mechanism has been elucidated, redundancy in the system has made it difficult to understand which of the many components interact to control the downstream biological processes. Here, we present a large-scale interaction study comprising most members of the Arabidopsis cytokinin signaling pathway. Using the yeast two-hybrid system, we detected 42 new interactions, of which more than 90% were confirmed by in vitro coaffinity purification. There are distinct patterns of interaction between protein families, but only a few interactions between proteins of the same family. An interaction map of this signaling pathway shows the Arabidopsis histidine phosphotransfer proteins as hubs, which interact with members from all other protein families, mostly in a redundant fashion. Domain-mapping experiments revealed the interaction domains of the proteins of this pathway. Analyses of Arabidopsis histidine phosphotransfer protein 5 mutant proteins showed that the presence of the canonical phospho-accepting histidine residue is not required for the interactions. Interaction of A-type response regulators with Arabidopsis histidine phosphotransfer proteins but not with B-type response regulators suggests that their known activity in feedback regulation may be realized by interfering at the level of Arabidopsis histidine phosphotransfer protein-mediated signaling. This study contributes to our understanding of the protein interactions of the cytokinin-signaling system and provides a framework for further functional studies in planta.

Analysis of corkscrew signaling in the Drosophila epidermal growth factor receptor pathway during myogenesis.

PubMed

Johnson Hamlet, M R; Perkins, L A

2001-11-01

The Drosophila nonreceptor protein tyrosine phosphatase, Corkscrew (Csw), functions positively in multiple receptor tyrosine kinase (RTK) pathways, including signaling by the epidermal growth factor receptor (EGFR). Detailed phenotypic analyses of csw mutations have revealed that Csw activity is required in many of the same developmental processes that require EGFR function. However, it is still unclear where in the signaling hierarchy Csw functions relative to other proteins whose activities are also required downstream of the receptor. To address this issue, genetic interaction experiments were performed to place csw gene activity relative to the EGFR, spitz (spi), rhomboid (rho), daughter of sevenless (DOS), kinase-suppressor of ras (ksr), ras1, D-raf, pointed (pnt), and moleskin. We followed the EGFR-dependent formation of VA2 muscle precursor cells as a sensitive assay for these genetic interaction studies. First, we established that Csw has a positive function during mesoderm development. Second, we found that tissue-specific expression of a gain-of-function csw construct rescues loss-of-function mutations in other positive signaling genes upstream of rolled (rl)/MAPK in the EGFR pathway. Third, we were able to infer levels of EGFR signaling in various mutant backgrounds during myogenesis. This work extends previous studies of Csw during Torso and Sevenless RTK signaling to include an in-depth analysis of the role of Csw in the EGFR signaling pathway.

Analysis of corkscrew signaling in the Drosophila epidermal growth factor receptor pathway during myogenesis.

PubMed Central

Johnson Hamlet, M R; Perkins, L A

2001-01-01

The Drosophila nonreceptor protein tyrosine phosphatase, Corkscrew (Csw), functions positively in multiple receptor tyrosine kinase (RTK) pathways, including signaling by the epidermal growth factor receptor (EGFR). Detailed phenotypic analyses of csw mutations have revealed that Csw activity is required in many of the same developmental processes that require EGFR function. However, it is still unclear where in the signaling hierarchy Csw functions relative to other proteins whose activities are also required downstream of the receptor. To address this issue, genetic interaction experiments were performed to place csw gene activity relative to the EGFR, spitz (spi), rhomboid (rho), daughter of sevenless (DOS), kinase-suppressor of ras (ksr), ras1, D-raf, pointed (pnt), and moleskin. We followed the EGFR-dependent formation of VA2 muscle precursor cells as a sensitive assay for these genetic interaction studies. First, we established that Csw has a positive function during mesoderm development. Second, we found that tissue-specific expression of a gain-of-function csw construct rescues loss-of-function mutations in other positive signaling genes upstream of rolled (rl)/MAPK in the EGFR pathway. Third, we were able to infer levels of EGFR signaling in various mutant backgrounds during myogenesis. This work extends previous studies of Csw during Torso and Sevenless RTK signaling to include an in-depth analysis of the role of Csw in the EGFR signaling pathway. PMID:11729154

A taxonomy of visualization tasks for the analysis of biological pathway data.

PubMed

Murray, Paul; McGee, Fintan; Forbes, Angus G

2017-02-15

Understanding complicated networks of interactions and chemical components is essential to solving contemporary problems in modern biology, especially in domains such as cancer and systems research. In these domains, biological pathway data is used to represent chains of interactions that occur within a given biological process. Visual representations can help researchers understand, interact with, and reason about these complex pathways in a number of ways. At the same time, these datasets offer unique challenges for visualization, due to their complexity and heterogeneity. Here, we present taxonomy of tasks that are regularly performed by researchers who work with biological pathway data. The generation of these tasks was done in conjunction with interviews with several domain experts in biology. These tasks require further classification than is provided by existing taxonomies. We also examine existing visualization techniques that support each task, and we discuss gaps in the existing visualization space revealed by our taxonomy. Our taxonomy is designed to support the development and design of future biological pathway visualization applications. We conclude by suggesting future research directions based on our taxonomy and motivated by the comments received by our domain experts.

A RET-ER81-NRG1 Signaling Pathway Drives the Development of Pacinian Corpuscles.

PubMed

Fleming, Michael S; Li, Jian J; Ramos, Daniel; Li, Tong; Talmage, David A; Abe, Shin-Ichi; Arber, Silvia; Luo, Wenqin

2016-10-05

Axon-Schwann cell interactions are crucial for the development, function, and repair of the peripheral nervous system, but mechanisms underlying communication between axons and nonmyelinating Schwann cells are unclear. Here, we show that ER81 is functionally required in a subset of mouse RET + mechanosensory neurons for formation of Pacinian corpuscles, which are composed of a single myelinated axon and multiple layers of nonmyelinating Schwann cells, and Ret is required for the maintenance of Er81 expression. Interestingly, Er81 mutants have normal myelination but exhibit deficient interactions between axons and corpuscle-forming nonmyelinating Schwann cells. Finally, ablating Neuregulin-1 (Nrg1) in mechanosensory neurons results in no Pacinian corpuscles, and an Nrg1 isoform not required for communication with myelinating Schwann cells is specifically decreased in Er81-null somatosensory neurons. Collectively, our results suggest that a RET-ER81-NRG1 signaling pathway promotes axon communication with nonmyelinating Schwann cells, and that neurons use distinct mechanisms to interact with different types of Schwann cells. Communication between neurons and Schwann cells is critical for development, normal function, and regeneration of the peripheral nervous system. Despite many studies about axonal communication with myelinating Schwann cells, mostly via a specific isoform of Neuregulin1, the molecular nature of axonal communication with nonmyelinating Schwann cells is poorly understood. Here, we described a RET-ER81-Neuregulin1 signaling pathway in neurons innervating Pacinian corpuscle somatosensory end organs, which is essential for communication between the innervating axon and the end organ nonmyelinating Schwann cells. We also showed that this signaling pathway uses isoforms of Neuregulin1 that are not involved in myelination, providing evidence that neurons use different isoforms of Neuregulin1 to interact with different types of Schwann cells. Copyright © 2016 the authors 0270-6474/16/3610337-19$15.00/0.

Fault tolerance in protein interaction networks: stable bipartite subgraphs and redundant pathways.

PubMed

Brady, Arthur; Maxwell, Kyle; Daniels, Noah; Cowen, Lenore J

2009-01-01

As increasing amounts of high-throughput data for the yeast interactome become available, more system-wide properties are uncovered. One interesting question concerns the fault tolerance of protein interaction networks: whether there exist alternative pathways that can perform some required function if a gene essential to the main mechanism is defective, absent or suppressed. A signature pattern for redundant pathways is the BPM (between-pathway model) motif, introduced by Kelley and Ideker. Past methods proposed to search the yeast interactome for BPM motifs have had several important limitations. First, they have been driven heuristically by local greedy searches, which can lead to the inclusion of extra genes that may not belong in the motif; second, they have been validated solely by functional coherence of the putative pathways using GO enrichment, making it difficult to evaluate putative BPMs in the absence of already known biological annotation. We introduce stable bipartite subgraphs, and show they form a clean and efficient way of generating meaningful BPMs which naturally discard extra genes included by local greedy methods. We show by GO enrichment measures that our BPM set outperforms previous work, covering more known complexes and functional pathways. Perhaps most importantly, since our BPMs are initially generated by examining the genetic-interaction network only, the location of edges in the protein-protein physical interaction network can then be used to statistically validate each candidate BPM, even with sparse GO annotation (or none at all). We uncover some interesting biological examples of previously unknown putative redundant pathways in such areas as vesicle-mediated transport and DNA repair.

Fault Tolerance in Protein Interaction Networks: Stable Bipartite Subgraphs and Redundant Pathways

PubMed Central

Brady, Arthur; Maxwell, Kyle; Daniels, Noah; Cowen, Lenore J.

2009-01-01

As increasing amounts of high-throughput data for the yeast interactome become available, more system-wide properties are uncovered. One interesting question concerns the fault tolerance of protein interaction networks: whether there exist alternative pathways that can perform some required function if a gene essential to the main mechanism is defective, absent or suppressed. A signature pattern for redundant pathways is the BPM (between-pathway model) motif, introduced by Kelley and Ideker. Past methods proposed to search the yeast interactome for BPM motifs have had several important limitations. First, they have been driven heuristically by local greedy searches, which can lead to the inclusion of extra genes that may not belong in the motif; second, they have been validated solely by functional coherence of the putative pathways using GO enrichment, making it difficult to evaluate putative BPMs in the absence of already known biological annotation. We introduce stable bipartite subgraphs, and show they form a clean and efficient way of generating meaningful BPMs which naturally discard extra genes included by local greedy methods. We show by GO enrichment measures that our BPM set outperforms previous work, covering more known complexes and functional pathways. Perhaps most importantly, since our BPMs are initially generated by examining the genetic-interaction network only, the location of edges in the protein-protein physical interaction network can then be used to statistically validate each candidate BPM, even with sparse GO annotation (or none at all). We uncover some interesting biological examples of previously unknown putative redundant pathways in such areas as vesicle-mediated transport and DNA repair. PMID:19399174

Signal transduction at fertilization: the Ca2+ release pathway in echinoderms and other invertebrate deuterostomes.

PubMed

Townley, Ian K; Roux, Michelle M; Foltz, Kathy R

2006-04-01

Gamete interaction and fusion triggers a number of events that lead to egg activation and development of a new organism. A key event at fertilization is the rise in intracellular calcium. In deuterostomes, this calcium is released from the egg's endoplasmic reticulum and is necessary for proper activation. This article reviews recent data regarding how gamete interaction triggers the initial calcium release, focusing on the echinoderms (invertebrate deuterostomes) as model systems. In eggs of these animals, Src-type kinases and phospholipase C-gamma are required components of the initial calcium trigger pathway in eggs.

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

PubMed Central

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

2011-01-01

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

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

PubMed

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

2014-11-01

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. Copyright © 2014 Elsevier Inc. All rights reserved.

Ligand requirements for involvement of PKCε in synergistic analgesic interactions between spinal μ and δ opioid receptors.

PubMed

Schuster, D J; Metcalf, M D; Kitto, K F; Messing, R O; Fairbanks, C A; Wilcox, G L

2015-01-01

We recently found that PKCε was required for spinal analgesic synergy between two GPCRs, δ opioid receptors and α2 A adrenoceptors, co-located in the same cellular subpopulation. We sought to determine if co-delivery of μ and δ opioid receptor agonists would similarly result in synergy requiring PKCε. Combinations of μ and δ opioid receptor agonists were co-administered intrathecally by direct lumbar puncture to PKCε-wild-type (PKCε-WT) and -knockout (PKCε-KO) mice. Antinociception was assessed using the hot-water tail-flick assay. Drug interactions were evaluated by isobolographic analysis. All agonists produced comparable antinociception in both PKCε-WT and PKCε-KO mice. Of 19 agonist combinations that produced analgesic synergy, only 3 required PKCε for a synergistic interaction. In these three combinations, one of the agonists was morphine, although not all combinations involving morphine required PKCε. Morphine + deltorphin II and morphine + deltorphin I required PKCε for synergy, whereas a similar combination, morphine + deltorphin, did not. Additionally, morphine + oxymorphindole required PKCε for synergy, whereas a similar combination, morphine + oxycodindole, did not. We discovered biased agonism for a specific signalling pathway at the level of spinally co-delivered opioid agonists. As the bias is only revealed by an appropriate ligand combination and cannot be accounted for by a single drug, it is likely that the receptors these agonists act on are interacting with each other. Our results support the existence of μ and δ opioid receptor heteromers at the spinal level in vivo. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2. © 2014 The British Pharmacological Society.

Apoptosis-linked Gene-2 (ALG-2)/Sec31 Interactions Regulate Endoplasmic Reticulum (ER)-to-Golgi Transport

PubMed Central

Helm, Jared R.; Bentley, Marvin; Thorsen, Kevin D.; Wang, Ting; Foltz, Lauren; Oorschot, Viola; Klumperman, Judith; Hay, Jesse C.

2014-01-01

Luminal calcium released from secretory organelles has been suggested to play a regulatory role in vesicle transport at several steps in the secretory pathway; however, its functional roles and effector pathways have not been elucidated. Here we demonstrate for the first time that specific luminal calcium depletion leads to a significant decrease in endoplasmic reticulum (ER)-to-Golgi transport rates in intact cells. Ultrastructural analysis revealed that luminal calcium depletion is accompanied by increased accumulation of intermediate compartment proteins in COPII buds and clusters of unfused COPII vesicles at ER exit sites. Furthermore, we present several lines of evidence suggesting that luminal calcium affected transport at least in part through calcium-dependent interactions between apoptosis-linked gene-2 (ALG-2) and the Sec31A proline-rich region: 1) targeted disruption of ALG-2/Sec31A interactions caused severe defects in ER-to-Golgi transport in intact cells; 2) effects of luminal calcium and ALG-2/Sec31A interactions on transport mutually required each other; and 3) Sec31A function in transport required luminal calcium. Morphological phenotypes of disrupted ALG-2/Sec31A interactions were characterized. We found that ALG-2/Sec31A interactions were not required for the localization of Sec31A to ER exit sites per se but appeared to acutely regulate the stability and trafficking of the cargo receptor p24 and the distribution of the vesicle tether protein p115. These results represent the first outline of a mechanism that connects luminal calcium to specific protein interactions regulating vesicle trafficking machinery. PMID:25006245

Chemotropism and Cell Fusion in Neurospora crassa Relies on the Formation of Distinct Protein Complexes by HAM-5 and a Novel Protein HAM-14

PubMed Central

Jonkers, Wilfried; Fischer, Monika S.; Do, Hung P.; Starr, Trevor L.; Glass, N. Louise

2016-01-01

In filamentous fungi, communication is essential for the formation of an interconnected, multinucleate, syncytial network, which is constructed via hyphal fusion or fusion of germinated asexual spores (germlings). Anastomosis in filamentous fungi is comparable to other somatic cell fusion events resulting in syncytia, including myoblast fusion during muscle differentiation, macrophage fusion, and fusion of trophoblasts during placental development. In Neurospora crassa, fusion of genetically identical germlings is a highly dynamic and regulated process that requires components of a MAP kinase signal transduction pathway. The kinase pathway components (NRC-1, MEK-2 and MAK-2) and the scaffold protein HAM-5 are recruited to hyphae and germling tips undergoing chemotropic interactions. The MAK-2/HAM-5 protein complex shows dynamic oscillation to hyphae/germling tips during chemotropic interactions, and which is out-of-phase to the dynamic localization of SOFT, which is a scaffold protein for components of the cell wall integrity MAP kinase pathway. In this study, we functionally characterize HAM-5 by generating ham-5 truncation constructs and show that the N-terminal half of HAM-5 was essential for function. This region is required for MAK-2 and MEK-2 interaction and for correct cellular localization of HAM-5 to “fusion puncta.” The localization of HAM-5 to puncta was not perturbed in 21 different fusion mutants, nor did these puncta colocalize with components of the secretory pathway. We also identified HAM-14 as a novel member of the HAM-5/MAK-2 pathway by mining MAK-2 phosphoproteomics data. HAM-14 was essential for germling fusion, but not for hyphal fusion. Colocalization and coimmunoprecipitation data indicate that HAM-14 interacts with MAK-2 and MEK-2 and may be involved in recruiting MAK-2 (and MEK-2) to complexes containing HAM-5. PMID:27029735

Chemotropism and Cell Fusion in Neurospora crassa Relies on the Formation of Distinct Protein Complexes by HAM-5 and a Novel Protein HAM-14.

PubMed

Jonkers, Wilfried; Fischer, Monika S; Do, Hung P; Starr, Trevor L; Glass, N Louise

2016-05-01

In filamentous fungi, communication is essential for the formation of an interconnected, multinucleate, syncytial network, which is constructed via hyphal fusion or fusion of germinated asexual spores (germlings). Anastomosis in filamentous fungi is comparable to other somatic cell fusion events resulting in syncytia, including myoblast fusion during muscle differentiation, macrophage fusion, and fusion of trophoblasts during placental development. In Neurospora crassa, fusion of genetically identical germlings is a highly dynamic and regulated process that requires components of a MAP kinase signal transduction pathway. The kinase pathway components (NRC-1, MEK-2 and MAK-2) and the scaffold protein HAM-5 are recruited to hyphae and germling tips undergoing chemotropic interactions. The MAK-2/HAM-5 protein complex shows dynamic oscillation to hyphae/germling tips during chemotropic interactions, and which is out-of-phase to the dynamic localization of SOFT, which is a scaffold protein for components of the cell wall integrity MAP kinase pathway. In this study, we functionally characterize HAM-5 by generating ham-5 truncation constructs and show that the N-terminal half of HAM-5 was essential for function. This region is required for MAK-2 and MEK-2 interaction and for correct cellular localization of HAM-5 to "fusion puncta." The localization of HAM-5 to puncta was not perturbed in 21 different fusion mutants, nor did these puncta colocalize with components of the secretory pathway. We also identified HAM-14 as a novel member of the HAM-5/MAK-2 pathway by mining MAK-2 phosphoproteomics data. HAM-14 was essential for germling fusion, but not for hyphal fusion. Colocalization and coimmunoprecipitation data indicate that HAM-14 interacts with MAK-2 and MEK-2 and may be involved in recruiting MAK-2 (and MEK-2) to complexes containing HAM-5. Copyright © 2016 by the Genetics Society of America.

Sorting nexin 9 recruits clathrin heavy chain to the mitotic spindle for chromosome alignment and segregation.

PubMed

Ma, Maggie P C; Robinson, Phillip J; Chircop, Megan

2013-01-01

Sorting nexin 9 (SNX9) and clathrin heavy chain (CHC) each have roles in mitosis during metaphase. Since the two proteins directly interact for their other cellular function in endocytosis we investigated whether they also interact for metaphase and operate on the same pathway. We report that SNX9 and CHC functionally interact during metaphase in a specific molecular pathway that contributes to stabilization of mitotic spindle kinetochore (K)-fibres for chromosome alignment and segregation. This function is independent of their endocytic role. SNX9 residues in the clathrin-binding low complexity domain are required for CHC association and for targeting both CHC and transforming acidic coiled-coil protein 3 (TACC3) to the mitotic spindle. Mutation of these sites to serine increases the metaphase plate width, indicating inefficient chromosome congression. Therefore SNX9 and CHC function in the same molecular pathway for chromosome alignment and segregation, which is dependent on their direct association.

Sorting Nexin 9 Recruits Clathrin Heavy Chain to the Mitotic Spindle for Chromosome Alignment and Segregation

PubMed Central

Ma, Maggie P. C.; Robinson, Phillip J.; Chircop, Megan

2013-01-01

Sorting nexin 9 (SNX9) and clathrin heavy chain (CHC) each have roles in mitosis during metaphase. Since the two proteins directly interact for their other cellular function in endocytosis we investigated whether they also interact for metaphase and operate on the same pathway. We report that SNX9 and CHC functionally interact during metaphase in a specific molecular pathway that contributes to stabilization of mitotic spindle kinetochore (K)-fibres for chromosome alignment and segregation. This function is independent of their endocytic role. SNX9 residues in the clathrin-binding low complexity domain are required for CHC association and for targeting both CHC and transforming acidic coiled-coil protein 3 (TACC3) to the mitotic spindle. Mutation of these sites to serine increases the metaphase plate width, indicating inefficient chromosome congression. Therefore SNX9 and CHC function in the same molecular pathway for chromosome alignment and segregation, which is dependent on their direct association. PMID:23861900

The Pathway Tools software.

PubMed

Karp, Peter D; Paley, Suzanne; Romero, Pedro

2002-01-01

Bioinformatics requires reusable software tools for creating model-organism databases (MODs). The Pathway Tools is a reusable, production-quality software environment for creating a type of MOD called a Pathway/Genome Database (PGDB). A PGDB such as EcoCyc (see http://ecocyc.org) integrates our evolving understanding of the genes, proteins, metabolic network, and genetic network of an organism. This paper provides an overview of the four main components of the Pathway Tools: The PathoLogic component supports creation of new PGDBs from the annotated genome of an organism. The Pathway/Genome Navigator provides query, visualization, and Web-publishing services for PGDBs. The Pathway/Genome Editors support interactive updating of PGDBs. The Pathway Tools ontology defines the schema of PGDBs. The Pathway Tools makes use of the Ocelot object database system for data management services for PGDBs. The Pathway Tools has been used to build PGDBs for 13 organisms within SRI and by external users.

A novel method to identify hub pathways of rheumatoid arthritis based on differential pathway networks.

PubMed

Wei, Shi-Tong; Sun, Yong-Hua; Zong, Shi-Hua

2017-09-01

The aim of the current study was to identify hub pathways of rheumatoid arthritis (RA) using a novel method based on differential pathway network (DPN) analysis. The present study proposed a DPN where protein‑protein interaction (PPI) network was integrated with pathway‑pathway interactions. Pathway data was obtained from background PPI network and the Reactome pathway database. Subsequently, pathway interactions were extracted from the pathway data by building randomized gene‑gene interactions and a weight value was assigned to each pathway interaction using Spearman correlation coefficient (SCC) to identify differential pathway interactions. Differential pathway interactions were visualized using Cytoscape to construct a DPN. Topological analysis was conducted to identify hub pathways that possessed the top 5% degree distribution of DPN. Modules of DPN were mined according to ClusterONE. A total of 855 pathways were selected to build pathway interactions. By filtrating pathway interactions of weight values >0.7, a DPN with 312 nodes and 791 edges was obtained. Topological degree analysis revealed 15 hub pathways, such as heparan sulfate/heparin‑glycosaminoglycan (HS‑GAG) degradation, HS‑GAG metabolism and keratan sulfate degradation for RA based on DPN. Furthermore, hub pathways were also important in modules, which validated the significance of hub pathways. In conclusion, the proposed method is a computationally efficient way to identify hub pathways of RA, which identified 15 hub pathways that may be potential biomarkers and provide insight to future investigation and treatment of RA.

Exploring pathway interactions in insulin resistant mouse liver

PubMed Central

2011-01-01

Background Complex phenotypes such as insulin resistance involve different biological pathways that may interact and influence each other. Interpretation of related experimental data would be facilitated by identifying relevant pathway interactions in the context of the dataset. Results We developed an analysis approach to study interactions between pathways by integrating gene and protein interaction networks, biological pathway information and high-throughput data. This approach was applied to a transcriptomics dataset to investigate pathway interactions in insulin resistant mouse liver in response to a glucose challenge. We identified regulated pathway interactions at different time points following the glucose challenge and also studied the underlying protein interactions to find possible mechanisms and key proteins involved in pathway cross-talk. A large number of pathway interactions were found for the comparison between the two diet groups at t = 0. The initial response to the glucose challenge (t = 0.6) was typed by an acute stress response and pathway interactions showed large overlap between the two diet groups, while the pathway interaction networks for the late response were more dissimilar. Conclusions Studying pathway interactions provides a new perspective on the data that complements established pathway analysis methods such as enrichment analysis. This study provided new insights in how interactions between pathways may be affected by insulin resistance. In addition, the analysis approach described here can be generally applied to different types of high-throughput data and will therefore be useful for analysis of other complex datasets as well. PMID:21843341

Crosstalk between Fas and JNK determines lymphocyte apoptosis after ionizing radiation.

PubMed

Praveen, Koganti; Saxena, Nandita

2013-06-01

Radiation simultaneously activate Fas and JNK pathway in lymphocytes but their precise interaction is not clearly understood. Activation of Fas pathway is required for radiation induced apoptosis, however induction of JNK pathway may or may not contribute in apoptosis. Here we report that Fas, Fas associated death domain and total JNK are activated in a dose- and time-dependent radiation exposure. A biphasic pattern of phospho-JNK was found at lower doses (1 and 2 Gy), however at higher doses of radiation phospho-JNK was continuously activated. Interestingly, Fas ligand expression remained biphasic at all the doses of radiation. Our results suggest that the Fas pathway is the major player in radiation-induced apoptosis, with JNK playing a contributory role. We also observed that Fas ligand expression by radiation is dependent on JNK activation. We also propose that radiation activates JNK pathway, but sustained activation is required for maximal induction of apoptosis at later times. Our findings define a mechanism for crosstalk between JNK and Fas pathway in radiation-induced apoptosis, which may lead to the development of new therapeutic strategies.

Trapping of the Enoyl-Acyl Carrier Protein Reductase–Acyl Carrier Protein Interaction

PubMed Central

Tallorin, Lorillee; Finzel, Kara; Nguyen, Quynh G.; Beld, Joris; La Clair, James J.; Burkart, Michael D.

2016-01-01

An ideal target for metabolic engineering, fatty acid biosynthesis remains poorly understood on a molecular level. These carrier protein-dependent pathways require fundamental protein–protein interactions to guide reactivity and processivity, and their control has become one of the major hurdles in successfully adapting these biological machines. Our laboratory has developed methods to prepare acyl carrier proteins (ACPs) loaded with substrate mimetics and cross-linkers to visualize and trap interactions with partner enzymes, and we continue to expand the tools for studying these pathways. We now describe application of the slow-onset, tight-binding inhibitor triclosan to explore the interactions between the type II fatty acid ACP from Escherichia coli, AcpP, and its corresponding enoyl-ACP reductase, FabI. We show that the AcpP–triclosan complex demonstrates nM binding, inhibits in vitro activity, and can be used to isolate FabI in complex proteomes. PMID:26938266

Co-opting the Fanconi Anemia Genomic Stability Pathway Enables Herpesvirus DNA Synthesis and Productive Growth

PubMed Central

Karttunen, Heidi; Savas, Jeffrey N.; McKinney, Caleb; Chen, Yu-Hung; Yates, John R.; Hukkanen, Veijo; Huang, Tony T.; Mohr, Ian

2015-01-01

SUMMARY DNA damage associated with viral DNA synthesis can result in double strand breaks that threaten genome integrity and must be repaired. Here, we establish that the cellular Fanconi Anemia (FA) genomic stability pathway is exploited by HSV1 to promote viral DNA synthesis and enable its productive growth. Potent FA pathway activation in HSV1-infected cells resulted in monoubiquitination of FA effector proteins, FANCI and FANCD2 (FANCI-D2) and required the viral DNA polymerase. FANCD2 relocalized to viral replication compartments and FANCI-D2 interacted with a multi-subunit complex containing the virus-encoded single-stranded DNA-binding protein ICP8. Significantly, while HSV1 productive growth was impaired in monoubiquitination-defective FA patient cells, this restriction was partially surmounted by antagonizing the DNA-dependent protein kinase (DNA-PK), a critical enzyme required for non-homologous end-joining (NHEJ). This identifies the FA-pathway as a new cellular factor required for herpesvirus productive growth and suggests that FA-mediated suppression of NHEJ is a fundamental step in the viral lifecycle. PMID:24954902

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

Quorum Sensing Regulation of Competence and Bacteriocins in Streptococcus pneumoniae and mutans

PubMed Central

Shanker, Erin; Federle, Michael J.

2017-01-01

The human pathogens Streptococcus pneumoniae and Streptococcus mutans have both evolved complex quorum sensing (QS) systems that regulate the production of bacteriocins and the entry into the competent state, a requirement for natural transformation. Natural transformation provides bacteria with a mechanism to repair damaged genes or as a source of new advantageous traits. In S. pneumoniae, the competence pathway is controlled by the two-component signal transduction pathway ComCDE, which directly regulates SigX, the alternative sigma factor required for the initiation into competence. Over the past two decades, effectors of cellular killing (i.e., fratricides) have been recognized as important targets of the pneumococcal competence QS pathway. Recently, direct interactions between the ComCDE and the paralogous BlpRH pathway, regulating bacteriocin production, were identified, further strengthening the interconnections between these two QS systems. Interestingly, a similar theme is being revealed in S. mutans, the primary etiological agent of dental caries. This review compares the relationship between the bacteriocin and the competence QS pathways in both S. pneumoniae and S. mutans, and hopes to provide clues to regulatory pathways across the genus Streptococcus as a potential tool to efficiently investigate putative competence pathways in nontransformable streptococci. PMID:28067778

Modulation of the Akt Pathway Reveals a Novel Link with PERK/eIF2α, which Is Relevant during Hypoxia

PubMed Central

Sánchez, Manuel Alejandro; Urrutia, Carolina; Grande, Alicia; Risso, Guillermo; Srebrow, Anabella; Alfaro, Jennifer; Colman-Lerner, Alejandro

2013-01-01

The unfolded protein response (UPR) and the Akt signaling pathway share several regulatory functions and have the capacity to determine cell outcome under specific conditions. However, both pathways have largely been studied independently. Here, we asked whether the Akt pathway regulates the UPR. To this end, we used a series of chemical compounds that modulate PI3K/Akt pathway and monitored the activity of the three UPR branches: PERK, IRE1 and ATF6. The antiproliferative and antiviral drug Akt-IV strongly and persistently activated all three branches of the UPR. We present evidence that activation of PERK/eIF2α requires Akt and that PERK is a direct Akt target. Chemical activation of this novel Akt/PERK pathway by Akt-IV leads to cell death, which was largely dependent on the presence of PERK and IRE1. Finally, we show that hypoxia-induced activation of eIF2α requires Akt, providing a physiologically relevant condition for the interaction between Akt and the PERK branch of the UPR. These data suggest the UPR and the Akt pathway signal to one another as a means of controlling cell fate. PMID:23922774

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

Wnt9a secreted from the walls of hepatic sinusoids is essential for morphogenesis, proliferation, and glycogen accumulation of chick hepatic epithelium.

PubMed

Matsumoto, Ken; Miki, Rika; Nakayama, Mizuho; Tatsumi, Norifumi; Yokouchi, Yuji

2008-07-15

Hepatic epithelial morphogenesis, including hepatoblast migration and proliferation in the septum transversum, requires the interaction of hepatic epithelium with the embryonic sinusoidal wall. No factors that mediate this interaction have yet been identified. As the beta-catenin pathway is active in hepatoblast proliferation, then Wnt ligands might activate the canonical Wnt pathway during liver development. Here, we investigated the role of Wnts in mediating epithelial vessel interactions in the developing chick liver. We found that Wnt9a was specifically expressed in both endothelial and stellate cells of the embryonic sinusoidal wall. Induced overexpression of Wnt9a resulted in hepatomegaly with hyperplasia of the hepatocellular cords, and in hyperproliferation of hepatocytes. Knockdown of Wnt9a caused a reduction in liver size, with hypoplasia of hepatocellular cord branching, and hypoproliferation of hepatoblasts, and also inhibited glycogen accumulation at later developmental stages. Wnt9a promoted in vivo stabilization of beta-catenin through binding with Frizzled 4, 7, and 9, and activated TOPflash reporter expression in vitro via Frizzled 7 and 9. Our results demonstrate that Wnt9a from the embryonic sinusoidal wall is required for the proper morphogenesis of chick hepatocellular cords, proliferation of hepatoblasts/hepatocytes, and glycogen accumulation in hepatocytes. Wnt9a signaling appears to be mediated by an Fzd7/9-beta-catenin pathway.

Stimulation of the Salicylic Acid Pathway Aboveground Recruits Entomopathogenic Nematodes Belowground

PubMed Central

Filgueiras, Camila Cramer; Willett, Denis S.; Junior, Alcides Moino; Pareja, Martin; Borai, Fahiem El; Dickson, Donald W.; Stelinski, Lukasz L.; Duncan, Larry W.

2016-01-01

Plant defense pathways play a critical role in mediating tritrophic interactions between plants, herbivores, and natural enemies. While the impact of plant defense pathway stimulation on natural enemies has been extensively explored aboveground, belowground ramifications of plant defense pathway stimulation are equally important in regulating subterranean pests and still require more attention. Here we investigate the effect of aboveground stimulation of the salicylic acid pathway through foliar application of the elicitor methyl salicylate on belowground recruitment of the entomopathogenic nematode, Steinernema diaprepesi. Also, we implicate a specific root-derived volatile that attracts S. diaprepesi belowground following aboveground plant stimulation by an elicitor. In four-choice olfactometer assays, citrus plants treated with foliar applications of methyl salicylate recruited S. diaprepesi in the absence of weevil feeding as compared with negative controls. Additionally, analysis of root volatile profiles of citrus plants receiving foliar application of methyl salicylate revealed production of d-limonene, which was absent in negative controls. The entomopathogenic nematode S. diaprepesi was recruited to d-limonene in two-choice olfactometer trials. These results reinforce the critical role of plant defense pathways in mediating tritrophic interactions, suggest a broad role for plant defense pathway signaling belowground, and hint at sophisticated plant responses to pest complexes. PMID:27136916

Dmc1 Functions in a Saccharomyces Cerevisiae Meiotic Pathway That Is Largely Independent of the Rad51 Pathway

PubMed Central

Dresser, M. E.; Ewing, D. J.; Conrad, M. N.; Dominguez, A. M.; Barstead, R.; Jiang, H.; Kodadek, T.

1997-01-01

Meiotic recombination in the yeast Saccharomyces cerevisiae requires two similar recA-like proteins, Dmc1p and Rad51p. A screen for dominant meiotic mutants provided DMC1-G126D, a dominant allele mutated in the conserved ATP-binding site (specifically, the A-loop motif) that confers a null phenotype. A recessive null allele, dmc1-K69E, was isolated as an intragenic suppressor of DMC1-G126D. Dmc1-K69Ep, unlike Dmc1p, does not interact homotypically in a two-hybrid assay, although it does interact with other fusion proteins identified by two-hybrid screen with Dmc1p. Dmc1p, unlike Rad51p, does not interact in the two-hybrid assay with Rad52p or Rad54p. However, Dmc1p does interact with Tid1p, a Rad54p homologue, with Tid4p, a Rad16p homologue, and with other fusion proteins that do not interact with Rad51p, suggesting that Dmc1p and Rad51p function in separate, though possibly overlapping, recombinational repair complexes. Epistasis analysis suggests that DMC1 and RAD51 function in separate pathways responsible for meiotic recombination. Taken together, our results are consistent with a requirement for DMC1 for meiosis-specific entry of DNA double-strand break ends into chromatin. Interestingly, the pattern on CHEF gels of chromosome fragments that result from meiotic DNA double-strand break formation is different in DMC1 mutant strains from that seen in rad50S strains. PMID:9335591

Responsive eLearning exercises to enhance student interaction with metabolic pathways.

PubMed

Roesler, William J; Dreaver-Charles, Kristine

2018-05-01

Successful learning of biochemistry requires students to engage with the material. In the past this often involved students writing out pathways by hand, and more recently directing students to online resources such as videos, songs, and animated slide presentations. However, even these latter resources do not really provide students an opportunity to engage with the material in an active fashion. As part of an online introductory metabolism course that was developed at our university, we created a series of twelve online interactive activities using Adobe Captivate 9. These activities targeted glycolysis, gluconeogenesis, the pentose phosphate pathway, glycogen metabolism, the citric acid cycle, and fatty acid oxidation. The interactive exercises consisted of two types. One involved dragging objects such as names of enzymes or allosteric modifiers to their correct drop locations such as a particular point in a metabolic pathway, a specific enzyme, and so forth. A second type involved clicking on objects, locations within a pathway, and so forth, in response to a particular question. In both types of exercises, students received feedback on their decisions in order to enhance learning. The student feedback received on these activities was very positive, and indicated that they found them to increase their confidence in the material and that they had learned the key principles of each pathway. © 2018 by The International Union of Biochemistry and Molecular Biology, 46(3):223-229, 2018. © 2018 The International Union of Biochemistry and Molecular Biology.

RecQL4 is required for the association of Mcm10 and Ctf4 with replication origins in human cells

PubMed Central

Im, Jun-Sub; Park, Soon-Young; Cho, Won-Ho; Bae, Sung-Ho; Hurwitz, Jerard; Lee, Joon-Kyu

2015-01-01

Though RecQL4 was shown to be essential for the initiation of DNA replication in mammalian cells, its role in initiation is poorly understood. Here, we show that RecQL4 is required for the origin binding of Mcm10 and Ctf4, and their physical interactions and association with replication origins are controlled by the concerted action of both CDK and DDK activities. Although RecQL4-dependent binding of Mcm10 and Ctf4 to chromatin can occur in the absence of pre-replicative complex, their association with replication origins requires the presence of the pre-replicative complex and CDK and DDK activities. Their association with replication origins and physical interactions are also targets of the DNA damage checkpoint pathways which prevent initiation of DNA replication at replication origins. Taken together, the RecQL4-dependent association of Mcm10 and Ctf4 with replication origins appears to be the first important step controlled by S phase promoting kinases and checkpoint pathways for the initiation of DNA replication in human cells. PMID:25602958

Neuroglian activates Echinoid to antagonize the Drosophila EGF receptor signaling pathway.

PubMed

Islam, Rafique; Wei, Shu-Yi; Chiu, Wei-Hsin; Hortsch, Michael; Hsu, Jui-Chou

2003-05-01

echinoid (ed) encodes an cell-adhesion molecule (CAM) that contains immunoglobulin domains and regulates the EGFR signaling pathway during Drosophila eye development. Based on our previous genetic mosaic and epistatic analysis, we proposed that Ed, via homotypic interactions, activates a novel, as yet unknown pathway that antagonizes EGFR signaling. In this report, we demonstrate that Ed functions as a homophilic adhesion molecule and also engages in a heterophilic trans-interaction with Drosophila Neuroglian (Nrg), an L1-type CAM. Co-expression of ed and nrg in the eye exhibits a strong genetic synergy in inhibiting EGFR signaling. This synergistic effect requires the intracellular domain of Ed, but not that of Nrg. In addition, Ed and Nrg colocalize in the Drosophila eye and are efficiently co-immunoprecipitated. Together, our results suggest a model in which Nrg acts as a heterophilic ligand and activator of Ed, which in turn antagonizes EGFR signaling.

Replication protein A: directing traffic at the intersection of replication and repair.

PubMed

Oakley, Greg G; Patrick, Steve M

2010-06-01

Since the initial discovery of replication protein A (RPA) as a DNA replication factor, much progress has been made on elucidating critical roles for RPA in other DNA metabolic pathways. RPA has been shown to be required for DNA replication, DNA repair, DNA recombination, and the DNA damage response pathway with roles in checkpoint activation. This review summarizes the current understanding of RPA structure, phosphorylation and protein-protein interactions in mediating these DNA metabolic processes.

DISCO interacting protein 2 determines direction of axon projection under the regulation of c-Jun N-terminal kinase in the Drosophila mushroom body

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

Nitta, Yohei; Brain Research Institute, Niigata University; Sugie, Atsushi

Precisely controlled axon guidance for complex neuronal wiring is essential for appropriate neuronal function. c-Jun N-terminal kinase (JNK) was found to play a role in axon guidance recently as well as in cell proliferation, protection and apoptosis. In spite of many genetic and molecular studies on these biological processes regulated by JNK, how JNK regulates axon guidance accurately has not been fully explained thus far. To address this question, we use the Drosophila mushroom body (MB) as a model since the α/β axons project in two distinct directions. Here we show that DISCO interacting protein 2 (DIP2) is required formore » the accurate direction of axonal guidance. DIP2 expression is under the regulation of Basket (Bsk), the Drosophila homologue of JNK. We additionally found that the Bsk/DIP2 pathway is independent from the AP-1 transcriptional factor complex pathway, which is directly activated by Bsk. In conclusion, our findings revealed DIP2 as a novel effector downstream of Bsk modulating the direction of axon projection. - Highlights: • DIP2 is required for accurate direction of axon guidance in Drosophila mushroom body. • DIP2 is a downstream of JNK in the axon guidance of Drosophila mushroom body neuron. • JNK/DIP2 pathway is independent from JNK/AP-1 transcriptional factor complex pathway.« less

Intracellular Electric Field and pH Optimize Protein Localization and Movement

PubMed Central

Cunningham, Jessica; Estrella, Veronica; Lloyd, Mark; Gillies, Robert; Frieden, B. Roy; Gatenby, Robert

2012-01-01

Mammalian cell function requires timely and accurate transmission of information from the cell membrane (CM) to the nucleus (N). These pathways have been intensively investigated and many critical components and interactions have been identified. However, the physical forces that control movement of these proteins have received scant attention. Thus, transduction pathways are typically presented schematically with little regard to spatial constraints that might affect the underlying dynamics necessary for protein-protein interactions and molecular movement from the CM to the N. We propose messenger protein localization and movements are highly regulated and governed by Coulomb interactions between: 1. A recently discovered, radially directed E-field from the NM into the CM and 2. Net protein charge determined by its isoelectric point, phosphorylation state, and the cytosolic pH. These interactions, which are widely applied in elecrophoresis, provide a previously unknown mechanism for localization of messenger proteins within the cytoplasm as well as rapid shuttling between the CM and N. Here we show these dynamics optimize the speed, accuracy and efficiency of transduction pathways even allowing measurement of the location and timing of ligand binding at the CM –previously unknown components of intracellular information flow that are, nevertheless, likely necessary for detecting spatial gradients and temporal fluctuations in ligand concentrations within the environment. The model has been applied to the RAF-MEK-ERK pathway and scaffolding protein KSR1 using computer simulations and in-vitro experiments. The computer simulations predicted distinct distributions of phosphorylated and unphosphorylated components of this transduction pathway which were experimentally confirmed in normal breast epithelial cells (HMEC). PMID:22623963

Tiam–Rac signaling mediates trans-endocytosis of ephrin receptor EphB2 and is important for cell repulsion

PubMed Central

2016-01-01

Ephrin receptors interact with membrane-bound ephrin ligands to regulate contact-mediated attraction or repulsion between opposing cells, thereby influencing tissue morphogenesis. Cell repulsion requires bidirectional trans-endocytosis of clustered Eph–ephrin complexes at cell interfaces, but the mechanisms underlying this process are poorly understood. Here, we identified an actin-regulating pathway allowing ephrinB+ cells to trans-endocytose EphB receptors from opposing cells. Live imaging revealed Rac-dependent F-actin enrichment at sites of EphB2 internalization, but not during vesicle trafficking. Systematic depletion of Rho family GTPases and their regulatory proteins identified the Rac subfamily and the Rac-specific guanine nucleotide exchange factor Tiam2 as key components of EphB2 trans-endocytosis, a pathway previously implicated in Eph forward signaling, in which ephrins act as in trans ligands of Eph receptors. However, unlike in Eph signaling, this pathway is not required for uptake of soluble ligands in ephrinB+ cells. We also show that this pathway is required for EphB2-stimulated contact repulsion. These results support the existence of a conserved pathway for EphB trans-endocytosis that removes the physical tether between cells, thereby enabling cell repulsion. PMID:27597758

Necrotrophic pathogens use the salicylic acid signaling pathway to promote disease development in tomato.

PubMed

Rahman, Taha Abd El; Oirdi, Mohamed El; Gonzalez-Lamothe, Rocio; Bouarab, Kamal

2012-12-01

Plants use different immune pathways to combat pathogens. The activation of the jasmonic acid (JA)-signaling pathway is required for resistance against necrotrophic pathogens; however, to combat biotrophic pathogens, the plants activate mainly the salicylic acid (SA)-signaling pathway. SA can antagonize JA signaling and vice versa. NPR1 (noninducible pathogenesis-related 1) is considered a master regulator of SA signaling. NPR1 interacts with TGA transcription factors, ultimately leading to the activation of SA-dependent responses. SA has been shown to promote disease development caused by the necrotrophic pathogen Botrytis cinerea through NPR1, by suppressing the expression of two JA-dependent defense genes, proteinase inhibitors I and II. We show here that the transcription factor TGA1.a contributes to disease development caused by B. cinerea in tomato by suppressing the expression of proteinase inhibitors I and II. Finally, we present evidence that the SA-signaling pathway contributes to disease development caused by another necrotrophic pathogen, Alternaria solani, in tomato. Disease development promoted by SA through NPR1 requires the TGA1.a transcription factor. These data highlight how necrotrophs manipulate the SAsignaling pathway to promote their disease in tomato.

Investigating multiple dysregulated pathways in rheumatoid arthritis based on pathway interaction network.

PubMed

Song, Xian-Dong; Song, Xian-Xu; Liu, Gui-Bo; Ren, Chun-Hui; Sun, Yuan-Bo; Liu, Ke-Xin; Liu, Bo; Liang, Shuang; Zhu, Zhu

2018-03-01

The traditional methods of identifying biomarkers in rheumatoid arthritis (RA) have focussed on the differentially expressed pathways or individual pathways, which however, neglect the interactions between pathways. To better understand the pathogenesis of RA, we aimed to identify dysregulated pathway sets using a pathway interaction network (PIN), which considered interactions among pathways. Firstly, RA-related gene expression profile data, protein-protein interactions (PPI) data and pathway data were taken up from the corresponding databases. Secondly, principal component analysis method was used to calculate the pathway activity of each of the pathway, and then a seed pathway was identified using data gleaned from the pathway activity. A PIN was then constructed based on the gene expression profile, pathway data, and PPI information. Finally, the dysregulated pathways were extracted from the PIN based on the seed pathway using the method of support vector machines and an area under the curve (AUC) index. The PIN comprised of a total of 854 pathways and 1064 pathway interactions. The greatest change in the activity score between RA and control samples was observed in the pathway of epigenetic regulation of gene expression, which was extracted and regarded as the seed pathway. Starting with this seed pathway, one maximum pathway set containing 10 dysregulated pathways was extracted from the PIN, having an AUC of 0.8249, and the result indicated that this pathway set could distinguish RA from the controls. These 10 dysregulated pathways might be potential biomarkers for RA diagnosis and treatment in the future.

Metabolic pathways for the whole community.

PubMed

Hanson, Niels W; Konwar, Kishori M; Hawley, Alyse K; Altman, Tomer; Karp, Peter D; Hallam, Steven J

2014-07-22

A convergence of high-throughput sequencing and computational power is transforming biology into information science. Despite these technological advances, converting bits and bytes of sequence information into meaningful insights remains a challenging enterprise. Biological systems operate on multiple hierarchical levels from genomes to biomes. Holistic understanding of biological systems requires agile software tools that permit comparative analyses across multiple information levels (DNA, RNA, protein, and metabolites) to identify emergent properties, diagnose system states, or predict responses to environmental change. Here we adopt the MetaPathways annotation and analysis pipeline and Pathway Tools to construct environmental pathway/genome databases (ePGDBs) that describe microbial community metabolism using MetaCyc, a highly curated database of metabolic pathways and components covering all domains of life. We evaluate Pathway Tools' performance on three datasets with different complexity and coding potential, including simulated metagenomes, a symbiotic system, and the Hawaii Ocean Time-series. We define accuracy and sensitivity relationships between read length, coverage and pathway recovery and evaluate the impact of taxonomic pruning on ePGDB construction and interpretation. Resulting ePGDBs provide interactive metabolic maps, predict emergent metabolic pathways associated with biosynthesis and energy production and differentiate between genomic potential and phenotypic expression across defined environmental gradients. This multi-tiered analysis provides the user community with specific operating guidelines, performance metrics and prediction hazards for more reliable ePGDB construction and interpretation. Moreover, it demonstrates the power of Pathway Tools in predicting metabolic interactions in natural and engineered ecosystems.

Planar polarity pathway and Nance-Horan syndrome-like 1b have essential cell-autonomous functions in neuronal migration.

PubMed

Walsh, Gregory S; Grant, Paul K; Morgan, John A; Moens, Cecilia B

2011-07-01

Components of the planar cell polarity (PCP) pathway are required for the caudal tangential migration of facial branchiomotor (FBM) neurons, but how PCP signaling regulates this migration is not understood. In a forward genetic screen, we identified a new gene, nhsl1b, required for FBM neuron migration. nhsl1b encodes a WAVE-homology domain-containing protein related to human Nance-Horan syndrome (NHS) protein and Drosophila GUK-holder (Gukh), which have been shown to interact with components of the WAVE regulatory complex that controls cytoskeletal dynamics and with the polarity protein Scribble, respectively. Nhsl1b localizes to FBM neuron membrane protrusions and interacts physically and genetically with Scrib to control FBM neuron migration. Using chimeric analysis, we show that FBM neurons have two modes of migration: one involving interactions between the neurons and their planar-polarized environment, and an alternative, collective mode involving interactions between the neurons themselves. We demonstrate that the first mode of migration requires the cell-autonomous functions of Nhsl1b and the PCP components Scrib and Vangl2 in addition to the non-autonomous functions of Scrib and Vangl2, which serve to polarize the epithelial cells in the environment of the migrating neurons. These results define a role for Nhsl1b as a neuronal effector of PCP signaling and indicate that proper FBM neuron migration is directly controlled by PCP signaling between the epithelium and the migrating neurons.

Planar polarity pathway and Nance-Horan syndrome-like 1b have essential cell-autonomous functions in neuronal migration

PubMed Central

Walsh, Gregory S.; Grant, Paul K.; Morgan, John A.; Moens, Cecilia B.

2011-01-01

Components of the planar cell polarity (PCP) pathway are required for the caudal tangential migration of facial branchiomotor (FBM) neurons, but how PCP signaling regulates this migration is not understood. In a forward genetic screen, we identified a new gene, nhsl1b, required for FBM neuron migration. nhsl1b encodes a WAVE-homology domain-containing protein related to human Nance-Horan syndrome (NHS) protein and Drosophila GUK-holder (Gukh), which have been shown to interact with components of the WAVE regulatory complex that controls cytoskeletal dynamics and with the polarity protein Scribble, respectively. Nhsl1b localizes to FBM neuron membrane protrusions and interacts physically and genetically with Scrib to control FBM neuron migration. Using chimeric analysis, we show that FBM neurons have two modes of migration: one involving interactions between the neurons and their planar-polarized environment, and an alternative, collective mode involving interactions between the neurons themselves. We demonstrate that the first mode of migration requires the cell-autonomous functions of Nhsl1b and the PCP components Scrib and Vangl2 in addition to the non-autonomous functions of Scrib and Vangl2, which serve to polarize the epithelial cells in the environment of the migrating neurons. These results define a role for Nhsl1b as a neuronal effector of PCP signaling and indicate that proper FBM neuron migration is directly controlled by PCP signaling between the epithelium and the migrating neurons. PMID:21693519

Multiple interactions between maternally-activated signalling pathways control Xenopus nodal-related genes.

PubMed

Rex, Maria; Hilton, Emma; Old, Robert

2002-03-01

We have investigated the induction of the six Xenopus nodal-related genes, Xnr1-Xnr6, by maternal determinants. The beta-catenin pathway was modelled by stimulation using Xwnt8, activin-like signalling was modelled by activin, and VegT action was studied by overexpression in animal cap explants. Combinations of factors were examined, and previously unrecognised interactions were revealed in animal caps and whole embryos. For the induction of Xnr5 and Xnr6 in whole embryos, using a beta-catenin antisense morpholino oligonucleotide or a dominant negative XTcf3, we have demonstrated an absolute permissive requirement for the beta-catenin/Tcf pathway, in addition to the requirement for VegT action. In animal caps Xnr5 and Xnr6 are induced in response to VegT overexpression, and this induction is dependent upon the concomitant activation of the beta-catenin pathway that VegT initiates in animal caps. For the induction of Xnr3, VegT interacts negatively so as to inhibit the induction otherwise observed with wnt-signalling alone. The negative effect of VegT is not the result of a general inhibition of wnt-signalling, and does not result from an inhibition of wnt-induced siamois expression. A 294 bp proximal promoter fragment of the Xnr3 gene is sufficient to mediate the negative effect of VegT. Further experiments, employing cycloheximide to examine the dependence of Xnr gene expression upon proteins translated after the mid-blastula stage, demonstrated that Xnrs 4, 5 and 6 are 'primary' Xnr genes whose expression in the late blastula is solely dependent upon factors present before the mid-blastula stage.

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

Bacillus thuringiensis Crystal Protein Cry6Aa Triggers Caenorhabditis elegans Necrosis Pathway Mediated by Aspartic Protease (ASP-1)

PubMed Central

Zhang, Fengjuan; Peng, Donghai; Cheng, Chunsheng; Zhou, Wei; Ju, Shouyong; Wan, Danfeng; Yu, Ziquan; Shi, Jianwei; Deng, Yaoyao; Wang, Fenshan; Ye, Xiaobo; Hu, Zhenfei; Lin, Jian; Ruan, Lifang; Sun, Ming

2016-01-01

Cell death plays an important role in host-pathogen interactions. Crystal proteins (toxins) are essential components of Bacillus thuringiensis (Bt) biological pesticides because of their specific toxicity against insects and nematodes. However, the mode of action by which crystal toxins to induce cell death is not completely understood. Here we show that crystal toxin triggers cell death by necrosis signaling pathway using crystal toxin Cry6Aa-Caenorhabditis elegans toxin-host interaction system, which involves an increase in concentrations of cytoplasmic calcium, lysosomal lyses, uptake of propidium iodide, and burst of death fluorescence. We find that a deficiency in the necrosis pathway confers tolerance to Cry6Aa toxin. Intriguingly, the necrosis pathway is specifically triggered by Cry6Aa, not by Cry5Ba, whose amino acid sequence is different from that of Cry6Aa. Furthermore, Cry6Aa-induced necrosis pathway requires aspartic protease (ASP-1). In addition, ASP-1 protects Cry6Aa from over-degradation in C. elegans. This is the first demonstration that deficiency in necrosis pathway confers tolerance to Bt crystal protein, and that Cry6A triggers necrosis represents a newly added necrosis paradigm in the C. elegans. Understanding this model could lead to new strategies for nematode control. PMID:26795495

Hsp90 is required for the activity of a hepatitis B virus reverse transcriptase.

PubMed Central

Hu, J; Seeger, C

1996-01-01

The heat shock protein Hsp90 is known as an essential component of several signal transduction pathways and has now been identified as an essential host factor for hepatitis B virus replication. Hsp90 interacts with the viral reverse transcriptase to facilitate the formation of a ribonucleoprotein (RNP) complex between the polymerase and an RNA ligand. This RNP complex is required early in replication for viral assembly and initiation of DNA synthesis through a protein-priming mechanism. These results thus invoke a role for the Hsp90 pathway in the formation of an RNP. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:8577714

ANDSystem: an Associative Network Discovery System for automated literature mining in the field of biology

PubMed Central

2015-01-01

Background Sufficient knowledge of molecular and genetic interactions, which comprise the entire basis of the functioning of living systems, is one of the necessary requirements for successfully answering almost any research question in the field of biology and medicine. To date, more than 24 million scientific papers can be found in PubMed, with many of them containing descriptions of a wide range of biological processes. The analysis of such tremendous amounts of data requires the use of automated text-mining approaches. Although a handful of tools have recently been developed to meet this need, none of them provide error-free extraction of highly detailed information. Results The ANDSystem package was developed for the reconstruction and analysis of molecular genetic networks based on an automated text-mining technique. It provides a detailed description of the various types of interactions between genes, proteins, microRNA's, metabolites, cellular components, pathways and diseases, taking into account the specificity of cell lines and organisms. Although the accuracy of ANDSystem is comparable to other well known text-mining tools, such as Pathway Studio and STRING, it outperforms them in having the ability to identify an increased number of interaction types. Conclusion The use of ANDSystem, in combination with Pathway Studio and STRING, can improve the quality of the automated reconstruction of molecular and genetic networks. ANDSystem should provide a useful tool for researchers working in a number of different fields, including biology, biotechnology, pharmacology and medicine. PMID:25881313

T7 replisome directly overcomes DNA damage

NASA Astrophysics Data System (ADS)

Sun, Bo; Pandey, Manjula; Inman, James T.; Yang, Yi; Kashlev, Mikhail; Patel, Smita S.; Wang, Michelle D.

2015-12-01

Cells and viruses possess several known `restart' pathways to overcome lesions during DNA replication. However, these `bypass' pathways leave a gap in replicated DNA or require recruitment of accessory proteins, resulting in significant delays to fork movement or even cell division arrest. Using single-molecule and ensemble methods, we demonstrate that the bacteriophage T7 replisome is able to directly replicate through a leading-strand cyclobutane pyrimidine dimer (CPD) lesion. We show that when a replisome encounters the lesion, a substantial fraction of DNA polymerase (DNAP) and helicase stay together at the lesion, the replisome does not dissociate and the helicase does not move forward on its own. The DNAP is able to directly replicate through the lesion by working in conjunction with helicase through specific helicase-DNAP interactions. These observations suggest that the T7 replisome is fundamentally permissive of DNA lesions via pathways that do not require fork adjustment or replisome reassembly.

ATP can be dispensable for prespliceosome formation in yeast

PubMed Central

Perriman, Rhonda; Ares, Manuel

2000-01-01

The first ATP-dependent step in pre-mRNA splicing involves the stable binding of U2 snRNP to form the prespliceosome. We show that a prespliceosome-like complex forms in the absence of ATP in yeast extracts lacking the U2 suppressor protein CUS2. These complexes display the same pre-mRNA and U snRNA requirements as authentic prespliceosomes and can be chased through the splicing pathway, indicating that they are a functional intermediate in the spliceosome assembly pathway. ATP-independent prespliceosome-like complexes are also observed in extracts containing a mutant U2 snRNA. Loss of CUS2 does not bypass the role of PRP5, an RNA helicase family member required for ATP-dependent prespliceosome formation. Genetic interactions between CUS2 and a heat-sensitive prp5 allele parallel those observed between CUS2 and U2, and suggest that CUS2 mediates functional interactions between U2 RNA and PRP5. We propose that CUS2 enforces ATP dependence during formation of the prespliceosome by brokering an interaction between PRP5 and the U2 snRNP that depends on correct U2 RNA structure. PMID:10640279

Defining the Protein–Protein Interaction Network of the Human Hippo Pathway*

PubMed Central

Wang, Wenqi; Li, Xu; Huang, Jun; Feng, Lin; Dolinta, Keithlee G.; Chen, Junjie

2014-01-01

The Hippo pathway, which is conserved from Drosophila to mammals, has been recognized as a tumor suppressor signaling pathway governing cell proliferation and apoptosis, two key events involved in organ size control and tumorigenesis. Although several upstream regulators, the conserved kinase cascade and key downstream effectors including nuclear transcriptional factors have been defined, the global organization of this signaling pathway is not been fully understood. Thus, we conducted a proteomic analysis of human Hippo pathway, which revealed the involvement of an extensive protein–protein interaction network in this pathway. The mass spectrometry data were deposited to ProteomeXchange with identifier PXD000415. Our data suggest that 550 interactions within 343 unique protein components constitute the central protein–protein interaction landscape of human Hippo pathway. Our study provides a glimpse into the global organization of Hippo pathway, reveals previously unknown interactions within this pathway, and uncovers new potential components involved in the regulation of this pathway. Understanding these interactions will help us further dissect the Hippo signaling-pathway and extend our knowledge of organ size control. PMID:24126142

A novel requirement for C. elegans Alix/ALX-1 in RME-1 mediated membrane transport

PubMed Central

Shi, Anbing; Pant, Saumya; Balklava, Zita; Chen, Carlos Chih-Hsiung; Figueroa, Vanesa; Grant, Barth D.

2007-01-01

Summary Background Alix/Bro1p family proteins have recently been identified as important components of multivesicular endosomes (MVEs) involved in the sorting of endocytosed integral membrane proteins, interacting with components of the ESCRT complex, the unconventional phospholipid LBPA, and other known endocytosis regulators. During infection Alix can be co-opted by enveloped retroviruses, including HIV, providing an important function during virus budding from the plasma membrane. In addition Alix is associated with the actin cytoskeleton and may regulate cytoskeletal dynamics. Results Here we demonstrate a novel physical interaction between the only apparent Alix/Bro1p family protein in C. elegans, ALX-1, and a key regulator of receptor recycling from endosomes to the plasma membrane called RME-1. Analysis of alx-1 mutants indicates that ALX-1 is required for endocytic recycling of specific basolateral cargo in the C. elegans intestine, a pathway previously defined by analysis of rme-1 mutants. Expression of truncated human Alix in HeLa cells disrupts recycling of MHCI, a known Ehd1/RME-1 dependent transport step, suggesting phylogenetic conservation of this function. We show that the interaction of ALX-1 with RME-1 in C. elegans, mediated by RME-1/YPSL and ALX-1/NPF motifs, is required for this recycling process. In the C. elegans intestine ALX-1 localizes to both recycling endosomes and MVEs, but the ALX-1/RME-1 interaction appears dispensable for ALX-1 function in MVEs/late endosomes. Conclusions This work provides the first demonstration of a requirement for an Alix/Bro1p family member in the endocytic recycling pathway in association with the recycling regulator RME-1. PMID:17997305

ABA signaling in guard cells entails a dynamic protein-protein interaction relay from the PYL-RCAR family receptors to ion channels.

PubMed

Lee, Sung Chul; Lim, Chae Woo; Lan, Wenzhi; He, Kai; Luan, Sheng

2013-03-01

Plant hormone abscisic acid (ABA) serves as an integrator of environmental stresses such as drought to trigger stomatal closure by regulating specific ion channels in guard cells. We previously reported that SLAC1, an outward anion channel required for stomatal closure, was regulated via reversible protein phosphorylation events involving ABA signaling components, including protein phosphatase 2C members and a SnRK2-type kinase (OST1). In this study, we reconstituted the ABA signaling pathway as a protein-protein interaction relay from the PYL/RCAR-type receptors, to the PP2C-SnRK2 phosphatase-kinase pairs, to the ion channel SLAC1. The ABA receptors interacted with and inhibited PP2C phosphatase activity against the SnRK2-type kinase, releasing active SnRK2 kinase to phosphorylate, and activate the SLAC1 channel, leading to reduced guard cell turgor and stomatal closure. Both yeast two-hybrid and bimolecular fluorescence complementation assays were used to verify the interactions among the components in the pathway. These biochemical assays demonstrated activity modifications of phosphatases and kinases by their interaction partners. The SLAC1 channel activity was used as an endpoint readout for the strength of the signaling pathway, depending on the presence of different combinations of signaling components. Further study using transgenic plants overexpressing one of the ABA receptors demonstrated that changing the relative level of interacting partners would change ABA sensitivity.

Bcr-Abl induces abnormal cytoskeleton remodeling, beta1 integrin clustering and increased cell adhesion to fibronectin through the Abl interactor 1 pathway.

PubMed

Li, Yingzhu; Clough, Nancy; Sun, Xiaolin; Yu, Weidong; Abbott, Brian L; Hogan, Christopher J; Dai, Zonghan

2007-04-15

Hematopoietic cells isolated from patients with Bcr-Abl-positive leukemia exhibit multiple abnormalities of cytoskeletal and integrin function. These abnormalities are thought to play a role in the pathogenesis of leukemia; however, the molecular events leading to these abnormalities are not fully understood. We show here that the Abi1 pathway is required for Bcr-Abl to stimulate actin cytoskeleton remodeling, integrin clustering and cell adhesion. Expression of Bcr-Abl induces tyrosine phosphorylation of Abi1. This is accompanied by a subcellular translocation of Abi1/WAVE2 to a site adjacent to membrane, where an F-actin-enriched structure containing the adhesion molecules such as beta1-integrin, paxillin and vinculin is assembled. Bcr-Abl-induced membrane translocation of Abi1/WAVE2 requires direct interaction between Abi1 and Bcr-Abl, but is independent of the phosphoinositide 3-kinase pathway. Formation of the F-actin-rich complex correlates with an increased cell adhesion to fibronectin. More importantly, disruption of the interaction between Bcr-Abl and Abi1 by mutations either in Bcr-Abl or Abi1 not only abolished tyrosine phosphorylation of Abi1 and membrane translocation of Abi1/WAVE2, but also inhibited Bcr-Abl-stimulated actin cytoskeleton remodeling, integrin clustering and cell adhesion to fibronectin. Together, these data define Abi1/WAVE2 as a downstream pathway that contributes to Bcr-Abl-induced abnormalities of cytoskeletal and integrin function.

Benchmarking pathway interaction network for colorectal cancer to identify dysregulated pathways.

PubMed

Wang, Q; Shi, C-J; Lv, S-H

2017-03-30

Different pathways act synergistically to participate in many biological processes. Thus, the purpose of our study was to extract dysregulated pathways to investigate the pathogenesis of colorectal cancer (CRC) based on the functional dependency among pathways. Protein-protein interaction (PPI) information and pathway data were retrieved from STRING and Reactome databases, respectively. After genes were aligned to the pathways, each pathway activity was calculated using the principal component analysis (PCA) method, and the seed pathway was discovered. Subsequently, we constructed the pathway interaction network (PIN), where each node represented a biological pathway based on gene expression profile, PPI data, as well as pathways. Dysregulated pathways were then selected from the PIN according to classification performance and seed pathway. A PIN including 11,960 interactions was constructed to identify dysregulated pathways. Interestingly, the interaction of mRNA splicing and mRNA splicing-major pathway had the highest score of 719.8167. Maximum change of the activity score between CRC and normal samples appeared in the pathway of DNA replication, which was selected as the seed pathway. Starting with this seed pathway, a pathway set containing 30 dysregulated pathways was obtained with an area under the curve score of 0.8598. The pathway of mRNA splicing, mRNA splicing-major pathway, and RNA polymerase I had the maximum genes of 107. Moreover, we found that these 30 pathways had crosstalks with each other. The results suggest that these dysregulated pathways might be used as biomarkers to diagnose CRC.

Stapled peptides as a new technology to investigate protein-protein interactions in human platelets.

PubMed

Iegre, Jessica; Ahmed, Niaz S; Gaynord, Josephine S; Wu, Yuteng; Herlihy, Kara M; Tan, Yaw Sing; Lopes-Pires, Maria E; Jha, Rupam; Lau, Yu Heng; Sore, Hannah F; Verma, Chandra; O' Donovan, Daniel H; Pugh, Nicholas; Spring, David R

2018-05-28

Platelets are blood cells with numerous crucial pathophysiological roles in hemostasis, cardiovascular thrombotic events and cancer metastasis. Platelet activation requires the engagement of intracellular signalling pathways that involve protein-protein interactions (PPIs). A better understanding of these pathways is therefore crucial for the development of selective anti-platelet drugs. New strategies for studying PPIs in human platelets are required to overcome limitations associated with conventional platelet research methods. For example, small molecule inhibitors can lack selectivity and are often difficult to design and synthesise. Additionally, development of transgenic animal models is costly and time-consuming and conventional recombinant techniques are ineffective due to the lack of a nucleus in platelets. Herein, we describe the generation of a library of novel, functionalised stapled peptides and their first application in the investigation of platelet PPIs. Moreover, the use of platelet-permeable stapled Bim BH3 peptides confirms the part of Bim in phosphatidyl-serine (PS) exposure and reveals a role for the Bim protein in platelet activatory processes. Our work demonstrates that functionalised stapled peptides are a complementary alternative to conventional platelet research methods, and could make a significant contribution to the understanding of platelet signalling pathways and hence to the development of anti-platelet drugs.

VISIBIOweb: visualization and layout services for BioPAX pathway models

PubMed Central

Dilek, Alptug; Belviranli, Mehmet E.; Dogrusoz, Ugur

2010-01-01

With recent advancements in techniques for cellular data acquisition, information on cellular processes has been increasing at a dramatic rate. Visualization is critical to analyzing and interpreting complex information; representing cellular processes or pathways is no exception. VISIBIOweb is a free, open-source, web-based pathway visualization and layout service for pathway models in BioPAX format. With VISIBIOweb, one can obtain well-laid-out views of pathway models using the standard notation of the Systems Biology Graphical Notation (SBGN), and can embed such views within one's web pages as desired. Pathway views may be navigated using zoom and scroll tools; pathway object properties, including any external database references available in the data, may be inspected interactively. The automatic layout component of VISIBIOweb may also be accessed programmatically from other tools using Hypertext Transfer Protocol (HTTP). The web site is free and open to all users and there is no login requirement. It is available at: http://visibioweb.patika.org. PMID:20460470

A New Method, "Reverse Yeast Two-Hybrid Array" (RYTHA), Identifies Mutants that Dissociate the Physical Interaction Between Elg1 and Slx5.

PubMed

Lev, Ifat; Shemesh, Keren; Volpe, Marina; Sau, Soumitra; Levinton, Nelly; Molco, Maya; Singh, Shivani; Liefshitz, Batia; Ben Aroya, Shay; Kupiec, Martin

2017-07-01

The vast majority of processes within the cell are carried out by proteins working in conjunction. The Yeast Two-Hybrid (Y2H) methodology allows the detection of physical interactions between any two interacting proteins. Here, we describe a novel systematic genetic methodology, "Reverse Yeast Two-Hybrid Array" (RYTHA), that allows the identification of proteins required for modulating the physical interaction between two given proteins. Our assay starts with a yeast strain in which the physical interaction of interest can be detected by growth on media lacking histidine, in the context of the Y2H methodology. By combining the synthetic genetic array technology, we can systematically screen mutant libraries of the yeast Saccharomyces cerevisiae to identify trans -acting mutations that disrupt the physical interaction of interest. We apply this novel method in a screen for mutants that disrupt the interaction between the N-terminus of Elg1 and the Slx5 protein. Elg1 is part of an alternative replication factor C-like complex that unloads PCNA during DNA replication and repair. Slx5 forms, together with Slx8, a SUMO-targeted ubiquitin ligase (STUbL) believed to send proteins to degradation. Our results show that the interaction requires both the STUbL activity and the PCNA unloading by Elg1, and identify topoisomerase I DNA-protein cross-links as a major factor in separating the two activities. Thus, we demonstrate that RYTHA can be applied to gain insights about particular pathways in yeast, by uncovering the connection between the proteasomal ubiquitin-dependent degradation pathway, DNA replication, and repair machinery, which can be separated by the topoisomerase-mediated cross-links to DNA. Copyright © 2017 by the Genetics Society of America.

The Hedgehog Signal Transduction Network

PubMed Central

Robbins, David J.; Fei, Dennis Liang; Riobo, Natalia A.

2013-01-01

Hedgehog (Hh) proteins regulate the development of a wide range of metazoan embryonic and adult structures, and disruption of Hh signaling pathways results in various human diseases. Here, we provide a comprehensive review of the signaling pathways regulated by Hh, consolidating data from a diverse array of organisms in a variety of scientific disciplines. Similar to the elucidation of many other signaling pathways, our knowledge of Hh signaling developed in a sequential manner centered on its earliest discoveries. Thus, our knowledge of Hh signaling has for the most part focused on elucidating the mechanism by which Hh regulates the Gli family of transcription factors, the so-called “canonical” Hh signaling pathway. However, in the past few years, numerous studies have shown that Hh proteins can also signal through Gli-independent mechanisms collectively referred to as “noncanonical” signaling pathways. Noncanonical Hh signaling is itself subdivided into two distinct signaling modules: (i) those not requiring Smoothened (Smo) and (ii) those downstream of Smo that do not require Gli transcription factors. Thus, Hh signaling is now proposed to occur through a variety of distinct context-dependent signaling modules that have the ability to crosstalk with one another to form an interacting, dynamic Hh signaling network. PMID:23074268

Models for the mechanism for activating copper-zinc superoxide dismutase in the absence of the CCS Cu chaperone in Arabidopsis.

PubMed

Huang, Chien-Hsun; Kuo, Wen-Yu; Jinn, Tsung-Luo

2012-03-01

Copper-zinc superoxide dismutase (CuZnSOD; CSD) is an important antioxidant enzyme for oxidative stress protection. To date, two activation pathways have been identified in many species. One requiring the CCS, Cu chaperone for SOD, to insert Cu and activate CSD (referred to as CCS-dependent pathway), and the other works independently of CCS (referred to as CCS-independent pathway). In our previous study, we suggest an unidentified factor will work with glutathione (GSH) for CSD activation in the absence of the CCS. Here, two models of the CCS-independent mechanism are proposed. The role of the unidentified factor may work as a scaffold protein, which provides a platform for the CSD protein and Cu-GSH to interact, or as a Cu carrier, which itself can bind Cu and interact with CSD proteins. We also suggest that the CSD protein conformation at C-terminal is important in providing a docking site for unidentified factor to access.

Bmp signaling mediates endoderm pouch morphogenesis by regulating Fgf signaling in zebrafish.

PubMed

Lovely, C Ben; Swartz, Mary E; McCarthy, Neil; Norrie, Jacqueline L; Eberhart, Johann K

2016-06-01

The endodermal pouches are a series of reiterated structures that segment the pharyngeal arches and help pattern the vertebrate face. Multiple pathways regulate the complex process of endodermal development, including the Bone morphogenetic protein (Bmp) pathway. However, the role of Bmp signaling in pouch morphogenesis is poorly understood. Using genetic and chemical inhibitor approaches, we show that pouch morphogenesis requires Bmp signaling from 10-18 h post-fertilization, immediately following gastrulation. Blocking Bmp signaling during this window results in morphological defects to the pouches and craniofacial skeleton. Using genetic chimeras we show that Bmp signals directly to the endoderm for proper morphogenesis. Time-lapse imaging and analysis of reporter transgenics show that Bmp signaling is necessary for pouch outpocketing via the Fibroblast growth factor (Fgf) pathway. Double loss-of-function analyses demonstrate that Bmp and Fgf signaling interact synergistically in craniofacial development. Collectively, our analyses shed light on the tissue and signaling interactions that regulate development of the vertebrate face. © 2016. Published by The Company of Biologists Ltd.

Bmp signaling mediates endoderm pouch morphogenesis by regulating Fgf signaling in zebrafish

PubMed Central

Swartz, Mary E.; McCarthy, Neil; Norrie, Jacqueline L.; Eberhart, Johann K.

2016-01-01

The endodermal pouches are a series of reiterated structures that segment the pharyngeal arches and help pattern the vertebrate face. Multiple pathways regulate the complex process of endodermal development, including the Bone morphogenetic protein (Bmp) pathway. However, the role of Bmp signaling in pouch morphogenesis is poorly understood. Using genetic and chemical inhibitor approaches, we show that pouch morphogenesis requires Bmp signaling from 10-18 h post-fertilization, immediately following gastrulation. Blocking Bmp signaling during this window results in morphological defects to the pouches and craniofacial skeleton. Using genetic chimeras we show that Bmp signals directly to the endoderm for proper morphogenesis. Time-lapse imaging and analysis of reporter transgenics show that Bmp signaling is necessary for pouch outpocketing via the Fibroblast growth factor (Fgf) pathway. Double loss-of-function analyses demonstrate that Bmp and Fgf signaling interact synergistically in craniofacial development. Collectively, our analyses shed light on the tissue and signaling interactions that regulate development of the vertebrate face. PMID:27122171

Exploring the interactions of EGFR with phosphorylated Mig6 by molecular dynamics simulations and MM-PBSA calculations.

PubMed

Zhang, Yue; Zheng, Qing-Chuan

2018-06-14

Mig6, a negative regulator, directly binds to epidermal growth factor receptor (EGFR), including Mig6-segment1 and Mig6-segment2. Mig6 requires phosphorylation of Y394 on Mig6-segment2 in order to inhibit EGFR. Two phosphorylation pathways for Y394 have been previously reported and the first way may phosphorylate Y394 primed by Y395 phosphorylation. Besides, the binding mechanism of phosphorylated Mig6-segment2 with EGFR has not been elucidated clearly. Focused on EGFR complex with phosphorylated Mig6-segment2, molecular dynamics (MD) simulations were performed to explore the interactions of Mig6-segment2 with EGFR. Our results indicate a probable phosphorylation pathway on Y394 and some key residues of EGFR play important roles in binding to phosphorylated Mig6-segment2. In addition, a special L-shaped structure was found to be possibly associated with irreversible inhibition of EGFR by Mig6. Our work can give meaningful information to better understand the phosphorylation pathways for Y394 and the interactions of EGFR binding to phosphorylated Mig6-segment2. Copyright © 2018 Elsevier Ltd. All rights reserved.

JNK signalling is necessary for a Wnt- and stem cell-dependent regeneration programme

PubMed Central

Tejada-Romero, Belen; Carter, Jean-Michel; Mihaylova, Yuliana; Neumann, Bjoern; Aboobaker, A. Aziz

2015-01-01

Regeneration involves the integration of new and old tissues in the context of an adult life history. It is clear that the core conserved signalling pathways that orchestrate development also play central roles in regeneration, and further study of conserved signalling pathways is required. Here we have studied the role of the conserved JNK signalling cascade during planarian regeneration. Abrogation of JNK signalling by RNAi or pharmacological inhibition blocks posterior regeneration and animals fail to express posterior markers. While the early injury-induced expression of polarity markers is unaffected, the later stem cell-dependent phase of posterior Wnt expression is not established. This defect can be rescued by overactivation of the Hh or Wnt signalling pathway to promote posterior Wnt activity. Together, our data suggest that JNK signalling is required to establish stem cell-dependent Wnt expression after posterior injury. Given that Jun is known to be required in vertebrates for the expression of Wnt and Wnt target genes, we propose that this interaction may be conserved and is an instructive part of planarian posterior regeneration. PMID:26062938

The CASTOR proteins are arginine sensors for the mTORC1 pathway

PubMed Central

Chantranupong, Lynne; Scaria, Sonia M.; Saxton, Robert A.; Gygi, Melanie P.; Shen, Kuang; Wyant, Gregory A.; Wang, Tim; Harper, J. Wade; Gygi, Steven P.; Sabatini, David M.

2016-01-01

Amino acids signal to the mTOR complex I (mTORC1) growth pathway through the Rag GTPases. Multiple distinct complexes regulate the Rags, including GATOR1, a GTPase activating protein (GAP), and GATOR2, a positive regulator of unknown molecular function. Arginine stimulation of cells activates mTORC1, but how it is sensed is not well understood. Recently, SLC38A9 was identified as a putative lysosomal arginine sensor required for arginine to activate mTORC1 but how arginine deprivation represses mTORC1 is unknown. Here, we show that CASTOR1, a previously uncharacterized protein, interacts with GATOR2 and is required for arginine deprivation to inhibit mTORC1. CASTOR1 homodimerizes and can also heterodimerize with the related protein, CASTOR2. Arginine disrupts the CASTOR1-GATOR2 complex by binding to CASTOR1 with a dissociation constant of ~30 μM, and its arginine-binding capacity is required for arginine to activate mTORC1 in cells. Collectively, these results establish CASTOR1 as an arginine sensor for the mTORC1 pathway. PMID:26972053

The Helicase Aquarius/EMB-4 Is Required to Overcome Intronic Barriers to Allow Nuclear RNAi Pathways to Heritably Silence Transcription.

PubMed

Akay, Alper; Di Domenico, Tomas; Suen, Kin M; Nabih, Amena; Parada, Guillermo E; Larance, Mark; Medhi, Ragini; Berkyurek, Ahmet C; Zhang, Xinlian; Wedeles, Christopher J; Rudolph, Konrad L M; Engelhardt, Jan; Hemberg, Martin; Ma, Ping; Lamond, Angus I; Claycomb, Julie M; Miska, Eric A

2017-08-07

Small RNAs play a crucial role in genome defense against transposable elements and guide Argonaute proteins to nascent RNA transcripts to induce co-transcriptional gene silencing. However, the molecular basis of this process remains unknown. Here, we identify the conserved RNA helicase Aquarius/EMB-4 as a direct and essential link between small RNA pathways and the transcriptional machinery in Caenorhabditis elegans. Aquarius physically interacts with the germline Argonaute HRDE-1. Aquarius is required to initiate small-RNA-induced heritable gene silencing. HRDE-1 and Aquarius silence overlapping sets of genes and transposable elements. Surprisingly, removal of introns from a target gene abolishes the requirement for Aquarius, but not HRDE-1, for small RNA-dependent gene silencing. We conclude that Aquarius allows small RNA pathways to compete for access to nascent transcripts undergoing co-transcriptional splicing in order to detect and silence transposable elements. Thus, Aquarius and HRDE-1 act as gatekeepers coordinating gene expression and genome defense. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

AMP-activated protein kinase is involved in the activation of the Fanconi anemia/BRCA pathway in response to DNA interstrand crosslinks.

PubMed

Chun, Min Jeong; Kim, Sunshin; Hwang, Soo Kyung; Kim, Bong Sub; Kim, Hyoun Geun; Choi, Hae In; Kim, Jong Heon; Goh, Sung Ho; Lee, Chang-Hun

2016-08-16

Fanconi anemia complementation group (FANC) proteins constitute the Fanconi Anemia (FA)/BRCA pathway that is activated in response to DNA interstrand crosslinks (ICLs). We previously performed yeast two-hybrid screening to identify novel FANC-interacting proteins and discovered that the alpha subunit of AMP-activated protein kinase (AMPKα1) was a candidate binding partner of the FANCG protein, which is a component of the FA nuclear core complex. We confirmed the interaction between AMPKα and both FANCG using co-immunoprecipitation experiments. Additionally, we showed that AMPKα interacted with FANCA, another component of the FA nuclear core complex. AMPKα knockdown in U2OS cells decreased FANCD2 monoubiquitination and nuclear foci formation upon mitomycin C-induced ICLs. Furthermore, AMPKα knockdown enhanced cellular sensitivity to MMC. MMC treatment resulted in an increase in AMPKα phosphorylation/activation, indicating AMPK is involved in the cellular response to ICLs. FANCA was phosphorylated by AMPK at S347 and phosphorylation increased with MMC treatment. MMC-induced FANCD2 monoubiquitination and nuclear foci formation were compromised in a U2OS cell line that stably overexpressed the S347A mutant form of FANCA compared to wild-type FANCA-overexpressing cells, indicating a requirement for FANCA phosphorylation at S347 for proper activation of the FA/BRCA pathway. Our data suggest AMPK is involved in the activation of the FA/BRCA pathway.

Influence of Berry-Polyphenols on Receptor Signaling and Cell-Death Pathways: Implications for Breast Cancer Prevention

PubMed Central

Aiyer, Harini S; Warri, Anni M; Woode, Denzel R; Hilakivi-Clarke, Leena; Clarke, Robert

2012-01-01

Breast cancer is the most commonly diagnosed cancer among women worldwide. Many women have become more aware of the benefits of increasing fruit consumption, as part of a healthy lifestyle, for the prevention of cancer. The mechanisms by which fruits, including berries, prevent breast cancer can be partially explained by exploring their interactions with pathways known influence cell-proliferation and evasion of cell-death. Two receptor pathways- estrogen receptor (ER) and tyrosine kinase receptors, especially the epidermal growth factor receptor (EGFR) family- are drivers of cell-proliferation and play a significant role in the development of both primary and recurrent breast cancer. There is strong evidence to show that several phytochemicals present in berries such as cyanidin, delphinidin, quercetin, kaempferol, ellagic acid, resveratrol and pterostilbene, interact with and alter the effects of these pathways. Further, they also induce cell death (apoptosis and autophagy) via their influence on kinase signaling. In this review, we summarize in vitro data regarding the interaction of berry polyphenols with the specific receptors and the mechanisms by which they induce cell death. Further, we also present in vivo data of primary breast cancer prevention by individual compounds and whole berries. Finally, we present a possible role for berries and berry compounds in the prevention of breast cancer and our perspective on the areas that require further research. PMID:22300613

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

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.

The Toll-Dorsal Pathway Is Required for Resistance to Viral Oral Infection in Drosophila

PubMed Central

Ferreira, Álvaro Gil; Naylor, Huw; Esteves, Sara Santana; Pais, Inês Silva; Martins, Nelson Eduardo; Teixeira, Luis

2014-01-01

Pathogen entry route can have a strong impact on the result of microbial infections in different hosts, including insects. Drosophila melanogaster has been a successful model system to study the immune response to systemic viral infection. Here we investigate the role of the Toll pathway in resistance to oral viral infection in D. melanogaster. We show that several Toll pathway components, including Spätzle, Toll, Pelle and the NF-kB-like transcription factor Dorsal, are required to resist oral infection with Drosophila C virus. Furthermore, in the fat body Dorsal is translocated from the cytoplasm to the nucleus and a Toll pathway target gene reporter is upregulated in response to Drosophila C Virus infection. This pathway also mediates resistance to several other RNA viruses (Cricket paralysis virus, Flock House virus, and Nora virus). Compared with control, viral titres are highly increased in Toll pathway mutants. The role of the Toll pathway in resistance to viruses in D. melanogaster is restricted to oral infection since we do not observe a phenotype associated with systemic infection. We also show that Wolbachia and other Drosophila-associated microbiota do not interact with the Toll pathway-mediated resistance to oral infection. We therefore identify the Toll pathway as a new general inducible pathway that mediates strong resistance to viruses with a route-specific role. These results contribute to a better understanding of viral oral infection resistance in insects, which is particularly relevant in the context of transmission of arboviruses by insect vectors. PMID:25473839

Insulation of a synthetic hydrogen metabolism circuit in bacteria

PubMed Central

2010-01-01

Background The engineering of metabolism holds tremendous promise for the production of desirable metabolites, particularly alternative fuels and other highly reduced molecules. Engineering approaches must redirect the transfer of chemical reducing equivalents, preventing these electrons from being lost to general cellular metabolism. This is especially the case for high energy electrons stored in iron-sulfur clusters within proteins, which are readily transferred when two such clusters are brought in close proximity. Iron sulfur proteins therefore require mechanisms to ensure interaction between proper partners, analogous to many signal transduction proteins. While there has been progress in the isolation of engineered metabolic pathways in recent years, the design of insulated electron metabolism circuits in vivo has not been pursued. Results Here we show that a synthetic hydrogen-producing electron transfer circuit in Escherichia coli can be insulated from existing cellular metabolism via multiple approaches, in many cases improving the function of the pathway. Our circuit is composed of heterologously expressed [Fe-Fe]-hydrogenase, ferredoxin, and pyruvate-ferredoxin oxidoreductase (PFOR), allowing the production of hydrogen gas to be coupled to the breakdown of glucose. We show that this synthetic pathway can be insulated through the deletion of competing reactions, rational engineering of protein interaction surfaces, direct protein fusion of interacting partners, and co-localization of pathway components on heterologous protein scaffolds. Conclusions Through the construction and characterization of a synthetic metabolic circuit in vivo, we demonstrate a novel system that allows for predictable engineering of an insulated electron transfer pathway. The development of this system demonstrates working principles for the optimization of engineered pathways for alternative energy production, as well as for understanding how electron transfer between proteins is controlled. PMID:20184755

Profiling the HER3/PI3K Pathway in Breast Tumors Using Proximity-Directed Assays Identifies Correlations between Protein Complexes and Phosphoproteins

PubMed Central

Mukherjee, Ali; Badal, Youssouf; Nguyen, Xuan-Thao; Miller, Johanna; Chenna, Ahmed; Tahir, Hasan; Newton, Alicia; Parry, Gordon; Williams, Stephen

2011-01-01

Background The identification of patients for targeted antineoplastic therapies requires accurate measurement of therapeutic targets and associated signaling complexes. HER3 signaling through heterodimerization is an important growth-promoting mechanism in several tumor types and may be a principal resistance mechanism by which EGFR and HER2 expressing tumors elude targeted therapies. Current methods that can study these interactions are inadequate for formalin-fixed, paraffin-embedded (FFPE) tumor samples. Methodology and Principal Findings Herein, we describe a panel of proximity-directed assays capable of measuring protein-interactions and phosphorylation in FFPE samples in the HER3/PI3K/Akt pathway and examine the capability of these assays to inform on the functional state of the pathway. We used FFPE breast cancer cell line and tumor models for this study. In breast cancer cell lines we observe both ligand-dependent and independent activation of the pathway and strong correlations between measured activation of key analytes. When selected cell lines are treated with HER2 inhibitors, we not only observe the expected molecular effects based on mechanism of action knowledge, but also novel effects of HER2 inhibition on key targets in the HER receptor pathway. Significantly, in a xenograft model of delayed tumor fixation, HER3 phosphorylation is unstable, while alternate measures of pathway activation, such as formation of the HER3PI3K complex is preserved. Measurements in breast tumor samples showed correlations between HER3 phosphorylation and receptor interactions, obviating the need to use phosphorylation as a surrogate for HER3 activation. Significance This assay system is capable of quantitatively measuring therapeutically relevant responses and enables molecular profiling of receptor networks in both preclinical and tumor models. PMID:21297994

When Clock Time Governs Interaction: How Time Influences Health Professionals' Intersectoral Collaboration.

PubMed

Bendix Andersen, Anne; Beedholm, Kirsten; Kolbæk, Raymond; Frederiksen, Kirsten

2018-06-01

When setting up patient pathways that cross health care sectors, professionals in emergency units strive to fulfill system requirements by creating efficient patient pathways that comply with standards for length of stay. We conducted an ethnographic field study, focusing on health professionals' collaboration, of 10 elderly patients with chronic illnesses, following them from discharge to their home or other places where they received health care services. We found that clock time not only governed the professionals' ways of collaborating, but acceleration of patient pathways also became an overall goal in health care delivery. Professionals' efforts to save time came to represent a "monetary value," leading to speedier planning of patient pathways and consequent risks of disregarding important issues when treating and caring for elderly patients. We suggest that such issues are significant to the future planning and improvement of patient pathways that involve elderly citizens who are in need of intersectoral health care delivery.

User Interface Requirements for Web-Based Integrated Care Pathways: Evidence from the Evaluation of an Online Care Pathway Investigation Tool.

PubMed

Balatsoukas, Panos; Williams, Richard; Davies, Colin; Ainsworth, John; Buchan, Iain

2015-11-01

Integrated care pathways (ICPs) define a chronological sequence of steps, most commonly diagnostic or treatment, to be followed in providing care for patients. Care pathways help to ensure quality standards are met and to reduce variation in practice. Although research on the computerisation of ICP progresses, there is still little knowledge on what are the requirements for designing user-friendly and usable electronic care pathways, or how users (normally health care professionals) interact with interfaces that support design, analysis and visualisation of ICPs. The purpose of the study reported in this paper was to address this gap by evaluating the usability of a novel web-based tool called COCPIT (Collaborative Online Care Pathway Investigation Tool). COCPIT supports the design, analysis and visualisation of ICPs at the population level. In order to address the aim of this study, an evaluation methodology was designed based on heuristic evaluations and a mixed method usability test. The results showed that modular visualisation and direct manipulation of information related to the design and analysis of ICPs is useful for engaging and stimulating users. However, designers should pay attention to issues related to the visibility of the system status and the match between the system and the real world, especially in relation to the display of statistical information about care pathways and the editing of clinical information within a care pathway. The paper concludes with recommendations for interface design.

Immune complex-induced human monocyte procoagulant activity. I. a rapid unidirectional lymphocyte-instructed pathway.

PubMed

Schwartz, B S; Edgington, T S

1981-09-01

It has previously been described that soluble antigen:antibody complexes in antigen excess can induce an increase in the procoagulant activity of human peripheral blood mononuclear cells. It has been proposed that this response may explain the presence of fibrin in immune complex-mediated tissue lesions. In the present study we define cellular participants and their roles in the procoagulant response to soluble immune complexes. Monocytes were shown by cell fractionation and by a direct cytologic assay to be the cell of origin of the procoagulant activity; and virtually all monocytes were able to participate in the response. Monocytes, however, required the presence of lymphocytes to respond. The procoagulant response required cell cooperation, and this collaborative interaction between lymphocytes and monocytes appeared to be unidirectional. Lymphocytes once triggered by immune complexes induced monocytes to synthesize the procoagulant product. Intact viable lymphocytes were required to present instructions to monocytes; no soluble mediator could be found to subserve this function. Indeed, all that appeared necessary to induce monocytes to produce procoagulant activity was an encounter with lymphocytes that had previously been in contact with soluble immune complexes. The optimum cellular ratio for this interaction was four lymphocytes per monocyte, about half the ratio in peripheral blood. The procoagulant response was rapid, reaching a maximum within 6 h after exposure to antigen:antibody complexes. The procoagulant activity was consistent with tissue factor because Factors VII and X and prothrombin were required for clotting of fibrinogen. WE propose that this pathway differs from a number of others involving cells of the immune system. Elucidation of the pathway may clarify the role of this lymphocyte-instructed monocyte response in the Shwartzman phenomenon and other thrombohemorrhagic events associated with immune cell function and the formation of immune complexes.

Network analyses based on comprehensive molecular interaction maps reveal robust control structures in yeast stress response pathways

PubMed Central

Kawakami, Eiryo; Singh, Vivek K; Matsubara, Kazuko; Ishii, Takashi; Matsuoka, Yukiko; Hase, Takeshi; Kulkarni, Priya; Siddiqui, Kenaz; Kodilkar, Janhavi; Danve, Nitisha; Subramanian, Indhupriya; Katoh, Manami; Shimizu-Yoshida, Yuki; Ghosh, Samik; Jere, Abhay; Kitano, Hiroaki

2016-01-01

Cellular stress responses require exquisite coordination between intracellular signaling molecules to integrate multiple stimuli and actuate specific cellular behaviors. Deciphering the web of complex interactions underlying stress responses is a key challenge in understanding robust biological systems and has the potential to lead to the discovery of targeted therapeutics for diseases triggered by dysregulation of stress response pathways. We constructed large-scale molecular interaction maps of six major stress response pathways in Saccharomyces cerevisiae (baker’s or budding yeast). Biological findings from over 900 publications were converted into standardized graphical formats and integrated into a common framework. The maps are posted at http://www.yeast-maps.org/yeast-stress-response/ for browse and curation by the research community. On the basis of these maps, we undertook systematic analyses to unravel the underlying architecture of the networks. A series of network analyses revealed that yeast stress response pathways are organized in bow–tie structures, which have been proposed as universal sub-systems for robust biological regulation. Furthermore, we demonstrated a potential role for complexes in stabilizing the conserved core molecules of bow–tie structures. Specifically, complex-mediated reversible reactions, identified by network motif analyses, appeared to have an important role in buffering the concentration and activity of these core molecules. We propose complex-mediated reactions as a key mechanism mediating robust regulation of the yeast stress response. Thus, our comprehensive molecular interaction maps provide not only an integrated knowledge base, but also a platform for systematic network analyses to elucidate the underlying architecture in complex biological systems. PMID:28725465

Diet-gene interactions underlie metabolic individuality and influence brain development: Implications for clinical practice

PubMed Central

Zeisel, Steven H.

2014-01-01

One of the underlying mechanisms for metabolic individuality is genetic variation. Single nucleotide polymorphisms (SNPs) in genes of metabolic pathways can create metabolic inefficiencies that alter the dietary requirement for, and responses to nutrients. These SNPS can be detected using genetic profiling and the metabolic inefficiencies they cause can be detected using metabolomic profiling. Studies on the human dietary requirement for choline illustrate how useful these new approaches can be, as this requirement is influenced by SNPs in genes of choline and folate metabolism. In adults, these SNPs determine whether people develop fatty liver, liver damage and muscle damage when eating diets low in choline. Because choline is very important for fetal development, these SNPs may identify women who need to eat more choline during pregnancy. Some of the actions of choline are mediated by epigenetic mechanisms that permit “retuning” of metabolic pathways during early life. PMID:22614815

Interaction of Adverse Disease Related Pathways in Hypertrophic Cardiomyopathy.

PubMed

Rowin, Ethan J; Maron, Martin S; Chan, Raymond H; Hausvater, Anais; Wang, Wendy; Rastegar, Hassan; Maron, Barry J

2017-12-15

Hypertrophic cardiomyopathy (HC) has been characterized as a generally progressive genetic heart disease, creating an ominous perspective for patients and managing cardiologists. We explored the HC disease burden and interaction of adverse clinical pathways to clarify patient expectations over long time periods in the contemporary therapeutic era. We studied 1,000 consecutive HC patients (52 ± 17 years) at Tufts Medical Center, followed 9.3 ± 8 years from diagnosis, employing a novel disease pathway model: 46% experienced a benign course free of adverse pathways, but 42% of patients progressed along 1 major pathway, most commonly refractory heart failure to New York Heart Association class III or IV requiring surgical myectomy (or alcohol ablation) or heart transplant; repetitive or permanent atrial fibrillation; and least commonly arrhythmic sudden death events. Eleven percent experienced 2 of these therapeutic end points at different times in their clinical course, most frequently the combination of advanced heart failure and atrial fibrillation, whereas only 1% incurred all 3 pathways. Freedom of progression from 1 to 2 disease pathways, or from 2 to 3 was 80% and 93% at 5 years, respectively. Annual HC-related mortality did not differ according to the number of pathways: 1 (0.8%), 2 (0.8%), or 3 (2.4%) (p = 0.56), and 93% of patients were in New York Heart Association classes I or II at follow-up. In conclusion, it is uncommon for HC patients to experience multiple adverse (but treatable) disease pathways, underscoring the principle that HC is not a uniformly progressive disease. These observations provide a measure of clarity and/or reassurance to patients regarding the true long-term disease burden of HC. Copyright © 2017 Elsevier Inc. All rights reserved.

Design and Structure-Guided Development of Novel Inhibitors of the Xeroderma Pigmentosum Group A (XPA) Protein-DNA Interaction.

PubMed

Gavande, Navnath S; VanderVere-Carozza, Pamela; Mishra, Akaash K; Vernon, Tyler L; Pawelczak, Katherine S; Turchi, John J

2017-10-12

XPA is a unique and essential protein required for the nucleotide excision DNA repair pathway and represents a therapeutic target in oncology. Herein, we are the first to develop novel inhibitors of the XPA-DNA interaction through structure-guided drug design efforts. Ester derivatives of the compounds 1 (X80), 22, and 24 displayed excellent inhibitory activity (IC 50 of 0.82 ± 0.18 μM and 1.3 ± 0.22 μM, respectively) but poor solubility. We have synthesized novel amide derivatives that retain potency and have much improved solubility. Furthermore, compound 1 analogs exhibited good specificity for XPA over RPA (replication protein A), another DNA-binding protein that participates in the nucleotide excision repair (NER) pathway. Importantly, there were no significant interactions observed by the X80 class of compounds directly with DNA. Molecular docking studies revealed a mechanistic model for the interaction, and these studies could serve as the basis for continued analysis of structure-activity relationships and drug development efforts of this novel target.

Investigating dysregulated pathways in Staphylococcus aureus (SA) exposed macrophages based on pathway interaction network.

PubMed

Zhou, Wei; Zhang, Yan; Li, Yue-Hua; Wang, Shuang; Zhang, Jing-Jing; Zhang, Cui-Xia; Zhang, Zhi-Sheng

2017-02-01

This work aimed to identify dysregulated pathways for Staphylococcus aureus (SA) exposed macrophages based on pathway interaction network (PIN). The inference of dysregulated pathways was comprised of four steps: preparing gene expression data, protein-protein interaction (PPI) data and pathway data; constructing a PIN dependent on the data and Pearson correlation coefficient (PCC); selecting seed pathway from PIN by computing activity score for each pathway according to principal component analysis (PCA) method; and investigating dysregulated pathways in a minimum set of pathways (MSP) utilizing seed pathway and the area under the receiver operating characteristics curve (AUC) index implemented in support vector machines (SVM) model. A total of 20,545 genes, 449,833 interactions and 1189 pathways were obtained in the gene expression data, PPI data and pathway data, respectively. The PIN was consisted of 8388 interactions and 1189 nodes, and Respiratory electron transport, ATP synthesis by chemiosmotic coupling, and heat production by uncoupling proteins was identified as the seed pathway. Finally, 15 dysregulated pathways in MSP (AUC=0.999) were obtained for SA infected samples, such as Respiratory electron transport and DNA Replication. We have identified 15 dysregulated pathways for SA infected macrophages based on PIN. The findings might provide potential biomarkers for early detection and therapy of SA infection, and give insights to reveal the molecular mechanism underlying SA infections. However, how these dysregulated pathways worked together still needs to be studied. Copyright © 2016 Elsevier Ltd. All rights reserved.

The Ebola virus glycoprotein mediates entry via a non-classical dynamin-dependent macropinocytic pathway

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

Mulherkar, Nirupama; Raaben, Matthijs; Torre, Juan Carlos de la

2011-10-25

Ebola virus (EBOV) has been reported to enter cultured cell lines via a dynamin-2-independent macropinocytic pathway or clathrin-mediated endocytosis. The route(s) of productive EBOV internalization into physiologically relevant cell types remain unexplored, and viral-host requirements for this process are incompletely understood. Here, we use electron microscopy and complementary chemical and genetic approaches to demonstrate that the viral glycoprotein, GP, induces macropinocytic uptake of viral particles into cells. GP's highly-glycosylated mucin domain is dispensable for virus-induced macropinocytosis, arguing that interactions between other sequences in GP and the host cell surface are responsible. Unexpectedly, we also found a requirement for the largemore » GTPase dynamin-2, which is proposed to be dispensable for several types of macropinocytosis. Our results provide evidence that EBOV uses an atypical dynamin-dependent macropinocytosis-like entry pathway to enter Vero cells, adherent human peripheral blood-derived monocytes, and a mouse dendritic cell line.« less

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

Computational Reconstruction of NFκB Pathway Interaction Mechanisms during Prostate Cancer

PubMed Central

Börnigen, Daniela; Tyekucheva, Svitlana; Wang, Xiaodong; Rider, Jennifer R.; Lee, Gwo-Shu; Mucci, Lorelei A.; Sweeney, Christopher; Huttenhower, Curtis

2016-01-01

Molecular research in cancer is one of the largest areas of bioinformatic investigation, but it remains a challenge to understand biomolecular mechanisms in cancer-related pathways from high-throughput genomic data. This includes the Nuclear-factor-kappa-B (NFκB) pathway, which is central to the inflammatory response and cell proliferation in prostate cancer development and progression. Despite close scrutiny and a deep understanding of many of its members’ biomolecular activities, the current list of pathway members and a systems-level understanding of their interactions remains incomplete. Here, we provide the first steps toward computational reconstruction of interaction mechanisms of the NFκB pathway in prostate cancer. We identified novel roles for ATF3, CXCL2, DUSP5, JUNB, NEDD9, SELE, TRIB1, and ZFP36 in this pathway, in addition to new mechanistic interactions between these genes and 10 known NFκB pathway members. A newly predicted interaction between NEDD9 and ZFP36 in particular was validated by co-immunoprecipitation, as was NEDD9's potential biological role in prostate cancer cell growth regulation. We combined 651 gene expression datasets with 1.4M gene product interactions to predict the inclusion of 40 additional genes in the pathway. Molecular mechanisms of interaction among pathway members were inferred using recent advances in Bayesian data integration to simultaneously provide information specific to biological contexts and individual biomolecular activities, resulting in a total of 112 interactions in the fully reconstructed NFκB pathway: 13 (11%) previously known, 29 (26%) supported by existing literature, and 70 (63%) novel. This method is generalizable to other tissue types, cancers, and organisms, and this new information about the NFκB pathway will allow us to further understand prostate cancer and to develop more effective prevention and treatment strategies. PMID:27078000

Polyhydroxylated [60]fullerene binds specifically to functional recognition sites on a monomeric and a dimeric ubiquitin

NASA Astrophysics Data System (ADS)

Zanzoni, Serena; Ceccon, Alberto; Assfalg, Michael; Singh, Rajesh K.; Fushman, David; D'Onofrio, Mariapina

2015-04-01

The use of nanoparticles (NPs) in biomedical applications requires an in-depth understanding of the mechanisms by which NPs interact with biomolecules. NPs associating with proteins may interfere with protein-protein interactions and affect cellular communication pathways, however the impact of NPs on biomolecular recognition remains poorly characterized. In this respect, particularly relevant is the study of NP-induced functional perturbations of proteins implicated in the regulation of key biochemical pathways. Ubiquitin (Ub) is a prototypical protein post-translational modifier playing a central role in numerous essential biological processes. To contribute to the understanding of the interactions between this universally distributed biomacromolecule and NPs, we investigated the adsorption of polyhydroxylated [60]fullerene on monomeric Ub and on a minimal polyubiquitin chain in vitro at atomic resolution. Site-resolved chemical shift and intensity perturbations of Ub's NMR signals, together with 15N spin relaxation rate changes, exchange saturation transfer effects, and fluorescence quenching data were consistent with the reversible formation of soluble aggregates incorporating fullerenol clusters. The specific interaction epitopes were identified, coincident with functional recognition sites in a monomeric and lysine48-linked dimeric Ub. Fullerenol appeared to target the open state of the dynamic structure of a dimeric Ub according to a conformational selection mechanism. Importantly, the protein-NP association prevented the enzyme-catalyzed synthesis of polyubiquitin chains. Our findings provide an experiment-based insight into protein/fullerenol recognition, with implications in functional biomolecular communication, including regulatory protein turnover, and for the opportunity of therapeutic intervention in Ub-dependent cellular pathways.The use of nanoparticles (NPs) in biomedical applications requires an in-depth understanding of the mechanisms by which NPs interact with biomolecules. NPs associating with proteins may interfere with protein-protein interactions and affect cellular communication pathways, however the impact of NPs on biomolecular recognition remains poorly characterized. In this respect, particularly relevant is the study of NP-induced functional perturbations of proteins implicated in the regulation of key biochemical pathways. Ubiquitin (Ub) is a prototypical protein post-translational modifier playing a central role in numerous essential biological processes. To contribute to the understanding of the interactions between this universally distributed biomacromolecule and NPs, we investigated the adsorption of polyhydroxylated [60]fullerene on monomeric Ub and on a minimal polyubiquitin chain in vitro at atomic resolution. Site-resolved chemical shift and intensity perturbations of Ub's NMR signals, together with 15N spin relaxation rate changes, exchange saturation transfer effects, and fluorescence quenching data were consistent with the reversible formation of soluble aggregates incorporating fullerenol clusters. The specific interaction epitopes were identified, coincident with functional recognition sites in a monomeric and lysine48-linked dimeric Ub. Fullerenol appeared to target the open state of the dynamic structure of a dimeric Ub according to a conformational selection mechanism. Importantly, the protein-NP association prevented the enzyme-catalyzed synthesis of polyubiquitin chains. Our findings provide an experiment-based insight into protein/fullerenol recognition, with implications in functional biomolecular communication, including regulatory protein turnover, and for the opportunity of therapeutic intervention in Ub-dependent cellular pathways. Electronic supplementary information (ESI) available: Experimental details. Fig. S1. Characterization of fullerenol by dynamic light scattering. Fig. S2. Size-exclusion chromatography. Fig. S3. 15N R1 spin relaxation rates of Ub and Ub2 upon subsequent additions of fullerenol. See DOI: 10.1039/c5nr00539f

Rhizobium–legume symbiosis shares an exocytotic pathway required for arbuscule formation

PubMed Central

Ivanov, Sergey; Fedorova, Elena E.; Limpens, Erik; De Mita, Stephane; Genre, Andrea; Bonfante, Paola; Bisseling, Ton

2012-01-01

Endosymbiotic interactions are characterized by the formation of specialized membrane compartments, by the host in which the microbes are hosted, in an intracellular manner. Two well-studied examples, which are of major agricultural and ecological importance, are the widespread arbuscular mycorrhizal symbiosis and the Rhizobium–legume symbiosis. In both symbioses, the specialized host membrane that surrounds the microbes forms a symbiotic interface, which facilitates the exchange of, for example, nutrients in a controlled manner and, therefore, forms the heart of endosymbiosis. Despite their key importance, the molecular and cellular mechanisms underlying the formation of these membrane interfaces are largely unknown. Recent studies strongly suggest that the Rhizobium–legume symbiosis coopted a signaling pathway, including receptor, from the more ancient arbuscular mycorrhizal symbiosis to form a symbiotic interface. Here, we show that two highly homologous exocytotic vesicle-associated membrane proteins (VAMPs) are required for formation of the symbiotic membrane interface in both interactions. Silencing of these Medicago VAMP72 genes has a minor effect on nonsymbiotic plant development and nodule formation. However, it blocks symbiosome as well as arbuscule formation, whereas root colonization by the microbes is not affected. Identification of these VAMP72s as common symbiotic regulators in exocytotic vesicle trafficking suggests that the ancient exocytotic pathway forming the periarbuscular membrane compartment has also been coopted in the Rhizobium–legume symbiosis. PMID:22566631

Two novel WD40 domain–containing proteins, Ere1 and Ere2, function in the retromer-mediated endosomal recycling pathway

PubMed Central

Shi, Yufeng; Stefan, Christopher J.; Rue, Sarah M.; Teis, David; Emr, Scott D.

2011-01-01

Regulated secretion, nutrient uptake, and responses to extracellular signals depend on cell-surface proteins that are internalized and recycled back to the plasma membrane. However, the underlying mechanisms that govern membrane protein recycling to the cell surface are not fully known. Using a chemical-genetic screen in yeast, we show that the arginine transporter Can1 is recycled back to the cell surface via two independent pathways mediated by the sorting nexins Snx4/41/42 and the retromer complex, respectively. In addition, we identify two novel WD40-domain endosomal recycling proteins, Ere1 and Ere2, that function in the retromer pathway. Ere1 is required for Can1 recycling via the retromer-mediated pathway, but it is not required for the transport of other retromer cargoes, such as Vps10 and Ftr1. Biochemical studies reveal that Ere1 physically interacts with internalized Can1. Ere2 is present in a complex containing Ere1 on endosomes and functions as a regulator of Ere1. Taken together, our results suggest that Snx4/41/42 and the retromer comprise two independent pathways for the recycling of internalized cell-surface proteins. Moreover, a complex containing the two novel proteins Ere1 and Ere2 mediates cargo-specific recognition by the retromer pathway. PMID:21880895

Pathway-based discovery of genetic interactions in breast cancer

PubMed Central

Xu, Zack Z.; Boone, Charles; Lange, Carol A.

2017-01-01

Breast cancer is the second largest cause of cancer death among U.S. women and the leading cause of cancer death among women worldwide. Genome-wide association studies (GWAS) have identified several genetic variants associated with susceptibility to breast cancer, but these still explain less than half of the estimated genetic contribution to the disease. Combinations of variants (i.e. genetic interactions) may play an important role in breast cancer susceptibility. However, due to a lack of statistical power, the current tests for genetic interactions from GWAS data mainly leverage prior knowledge to focus on small sets of genes or SNPs that are known to have an association with breast cancer. Thus, many genetic interactions, particularly among novel variants, remain understudied. Reverse-genetic interaction screens in model organisms have shown that genetic interactions frequently cluster into highly structured motifs, where members of the same pathway share similar patterns of genetic interactions. Based on this key observation, we recently developed a method called BridGE to search for such structured motifs in genetic networks derived from GWAS studies and identify pathway-level genetic interactions in human populations. We applied BridGE to six independent breast cancer cohorts and identified significant pathway-level interactions in five cohorts. Joint analysis across all five cohorts revealed a high confidence consensus set of genetic interactions with support in multiple cohorts. The discovered interactions implicated the glutathione conjugation, vitamin D receptor, purine metabolism, mitotic prometaphase, and steroid hormone biosynthesis pathways as major modifiers of breast cancer risk. Notably, while many of the pathways identified by BridGE show clear relevance to breast cancer, variants in these pathways had not been previously discovered by traditional single variant association tests, or single pathway enrichment analysis that does not consider SNP-SNP interactions. PMID:28957314

MEDIATOR25 Acts as an Integrative Hub for the Regulation of Jasmonate-Responsive Gene Expression in Arabidopsis1[C][W

PubMed Central

Çevik, Volkan; Kidd, Brendan N.; Zhang, Peijun; Hill, Claire; Kiddle, Steve; Denby, Katherine J.; Holub, Eric B.; Cahill, David M.; Manners, John M.; Schenk, Peer M.; Beynon, Jim; Kazan, Kemal

2012-01-01

The PHYTOCHROME AND FLOWERING TIME1 gene encoding the MEDIATOR25 (MED25) subunit of the eukaryotic Mediator complex is a positive regulator of jasmonate (JA)-responsive gene expression in Arabidopsis (Arabidopsis thaliana). Based on the function of the Mediator complex as a bridge between DNA-bound transcriptional activators and the RNA polymerase II complex, MED25 has been hypothesized to function in association with transcriptional regulators of the JA pathway. However, it is currently not known mechanistically how MED25 functions to regulate JA-responsive gene expression. In this study, we show that MED25 physically interacts with several key transcriptional regulators of the JA signaling pathway, including the APETALA2 (AP2)/ETHYLENE RESPONSE FACTOR (ERF) transcription factors OCTADECANOID-RESPONSIVE ARABIDOPSIS AP2/ERF59 and ERF1 as well as the master regulator MYC2. Physical interaction detected between MED25 and four group IX AP2/ERF transcription factors was shown to require the activator interaction domain of MED25 as well as the recently discovered Conserved Motif IX-1/EDLL transcription activation motif of MED25-interacting AP2/ERFs. Using transcriptional activation experiments, we also show that OCTADECANOID-RESPONSIVE ARABIDOPSIS AP2/ERF59- and ERF1-dependent activation of PLANT DEFENSIN1.2 as well as MYC2-dependent activation of VEGETATIVE STORAGE PROTEIN1 requires a functional MED25. In addition, MED25 is required for MYC2-dependent repression of pathogen defense genes. These results suggest an important role for MED25 as an integrative hub within the Mediator complex during the regulation of JA-associated gene expression. PMID:22822211

The Crystal Structure of the Ubiquitin-like Domain of Ribosome Assembly Factor Ytm1 and Characterization of Its Interaction with the AAA-ATPase Midasin*

PubMed Central

Romes, Erin M.; Sobhany, Mack; Stanley, Robin E.

2016-01-01

The synthesis of eukaryotic ribosomes is a complex, energetically demanding process requiring the aid of numerous non-ribosomal factors, such as the PeBoW complex. The mammalian PeBoW complex, composed of Pes1, Bop1, and WDR12, is essential for the processing of the 32S preribosomal RNA. Previous work in Saccharomyces cerevisiae has shown that release of the homologous proteins in this complex (Nop7, Erb1, and Ytm1, respectively) from preribosomal particles requires Rea1 (midasin or MDN1 in humans), a large dynein-like protein. Midasin contains a C-terminal metal ion-dependent adhesion site (MIDAS) domain that interacts with the N-terminal ubiquitin-like (UBL) domain of Ytm1/WDR12 as well as the UBL domain of Rsa4/Nle1 in a later step in the ribosome maturation pathway. Here we present the crystal structure of the UBL domain of the WDR12 homologue from S. cerevisiae at 1.7 Å resolution and demonstrate that human midasin binds to WDR12 as well as Nle1 through their respective UBL domains. Midasin contains a well conserved extension region upstream of the MIDAS domain required for binding WDR12 and Nle1, and the interaction is dependent upon metal ion coordination because removal of the metal or mutation of residues that coordinate the metal ion diminishes the interaction. Mammalian WDR12 displays prominent nucleolar localization that is dependent upon active ribosomal RNA transcription. Based upon these results, we propose that release of the PeBoW complex and subsequent release of Nle1 by midasin is a well conserved step in the ribosome maturation pathway in both yeast and mammalian cells. PMID:26601951

Detection of characteristic sub pathway network for angiogenesis based on the comprehensive pathway network.

PubMed

Huang, Yezhou; Li, Shao

2010-01-18

Pathways in biological system often cooperate with each other to function. Changes of interactions among pathways tightly associate with alterations in the properties and functions of the cell and hence alterations in the phenotype. So, the pathway interactions and especially their changes over time corresponding to specific phenotype are critical to understanding cell functions and phenotypic plasticity. With prior-defined pathways and incorporated protein-protein interaction (PPI) data, we counted PPIs between corresponding gene sets of each pair of distinct pathways to construct a comprehensive pathway network. Then we proposed a novel concept, characteristic sub pathway network (CSPN), to realize the phenotype-specific pathway interactions. By adding gene expression data regarding a given phenotype, angiogenesis, active PPIs corresponding to stimulation of interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF-alpha) on human umbilical vein endothelial cells (HUVECs) respectively were derived. Two kinds of CSPN, namely the static or the dynamic CSPN, were detected by counting active PPIs. A comprehensive pathway network containing 37 signalling pathways as nodes and 263 pathway interactions were obtained. Two phenotype-specific CSPNs for angiogenesis, corresponding to stimulation of IL-1 and TNF-alpha on HUVEC respectively, were addressed. From phenotype-specific CSPNs, a static CSPN involving interactions among B cell receptor, T cell receptor, Toll-like receptor, MAPK, VEGF, and ErbB signalling pathways, and a dynamic CSPN involving interactions among TGF-beta, Wnt, p53 signalling pathways and cell cycle pathway, were detected for angiogenesis on HUVEC after stimulation of IL-1 and TNF-alpha respectively. We inferred that, in certain case, the static CSPN maintains related basic functions of the cells, whereas the dynamic CSPN manifests the cells' plastic responses to stimulus and therefore reflects the cells' phenotypic plasticity. The comprehensive pathway network helps us realize the cooperative behaviours among pathways. Moreover, two kinds of potential CSPNs found in this work, the static CSPN and the dynamic CSPN, are helpful to deeply understand the specific function of HUVEC and its phenotypic plasticity in regard to angiogenesis.

Detection of characteristic sub pathway network for angiogenesis based on the comprehensive pathway network

PubMed Central

2010-01-01

Background Pathways in biological system often cooperate with each other to function. Changes of interactions among pathways tightly associate with alterations in the properties and functions of the cell and hence alterations in the phenotype. So, the pathway interactions and especially their changes over time corresponding to specific phenotype are critical to understanding cell functions and phenotypic plasticity. Methods With prior-defined pathways and incorporated protein-protein interaction (PPI) data, we counted PPIs between corresponding gene sets of each pair of distinct pathways to construct a comprehensive pathway network. Then we proposed a novel concept, characteristic sub pathway network (CSPN), to realize the phenotype-specific pathway interactions. By adding gene expression data regarding a given phenotype, angiogenesis, active PPIs corresponding to stimulation of interleukin-1 (IL-1) and tumor necrosis factor α (TNF-α) on human umbilical vein endothelial cells (HUVECs) respectively were derived. Two kinds of CSPN, namely the static or the dynamic CSPN, were detected by counting active PPIs. Results A comprehensive pathway network containing 37 signalling pathways as nodes and 263 pathway interactions were obtained. Two phenotype-specific CSPNs for angiogenesis, corresponding to stimulation of IL-1 and TNF-α on HUVEC respectively, were addressed. From phenotype-specific CSPNs, a static CSPN involving interactions among B cell receptor, T cell receptor, Toll-like receptor, MAPK, VEGF, and ErbB signalling pathways, and a dynamic CSPN involving interactions among TGF-β, Wnt, p53 signalling pathways and cell cycle pathway, were detected for angiogenesis on HUVEC after stimulation of IL-1 and TNF-α respectively. We inferred that, in certain case, the static CSPN maintains related basic functions of the cells, whereas the dynamic CSPN manifests the cells' plastic responses to stimulus and therefore reflects the cells' phenotypic plasticity. Conclusion The comprehensive pathway network helps us realize the cooperative behaviours among pathways. Moreover, two kinds of potential CSPNs found in this work, the static CSPN and the dynamic CSPN, are helpful to deeply understand the specific function of HUVEC and its phenotypic plasticity in regard to angiogenesis. PMID:20122205

P4-ATPase Requirement for AP-1/Clathrin Function in Protein Transport from the trans-Golgi Network and Early Endosomes

PubMed Central

Liu, Ke; Surendhran, Kavitha; Nothwehr, Steven F.

2008-01-01

Drs2p is a resident type 4 P-type ATPase (P4-ATPase) and potential phospholipid translocase of the trans-Golgi network (TGN) where it has been implicated in clathrin function. However, precise protein transport pathways requiring Drs2p and how it contributes to clathrin-coated vesicle budding remain unclear. Here we show a functional codependence between Drs2p and the AP-1 clathrin adaptor in protein sorting at the TGN and early endosomes of Saccharomyces cerevisiae. Genetic criteria indicate that Drs2p and AP-1 operate in the same pathway and that AP-1 requires Drs2p for function. In addition, we show that loss of AP-1 markedly increases Drs2p trafficking to the plasma membrane, but does not perturb retrieval of Drs2p from the early endosome back to the TGN. Thus AP-1 is required at the TGN to sort Drs2p out of the exocytic pathway, presumably for delivery to the early endosome. Moreover, a conditional allele that inactivates Drs2p phospholipid translocase (flippase) activity disrupts its own transport in this AP-1 pathway. Drs2p physically interacts with AP-1; however, AP-1 and clathrin are both recruited normally to the TGN in drs2Δ cells. These results imply that Drs2p acts independently of coat recruitment to facilitate AP-1/clathrin-coated vesicle budding from the TGN. PMID:18508916

Apolipoprotein E Likely Contributes to a Maturation Step of Infectious Hepatitis C Virus Particles and Interacts with Viral Envelope Glycoproteins

PubMed Central

Lee, Ji-Young; Acosta, Eliana G.; Stoeck, Ina Karen; Long, Gang; Hiet, Marie-Sophie; Mueller, Birthe; Fackler, Oliver T.; Kallis, Stephanie

2014-01-01

ABSTRACT The assembly of infectious hepatitis C virus (HCV) particles is tightly linked to components of the very-low-density lipoprotein (VLDL) pathway. We and others have shown that apolipoprotein E (ApoE) plays a major role in production of infectious HCV particles. However, the mechanism by which ApoE contributes to virion assembly/release and how it gets associated with the HCV particle is poorly understood. We found that knockdown of ApoE reduces titers of infectious intra- and extracellular HCV but not of the related dengue virus. ApoE depletion also reduced amounts of extracellular HCV core protein without affecting intracellular core amounts. Moreover, we found that ApoE depletion affected neither formation of nucleocapsids nor their envelopment, suggesting that ApoE acts at a late step of assembly, such as particle maturation and infectivity. Importantly, we demonstrate that ApoE interacts with the HCV envelope glycoproteins, most notably E2. This interaction did not require any other viral proteins and depended on the transmembrane domain of E2 that also was required for recruitment of HCV envelope glycoproteins to detergent-resistant membrane fractions. These results suggest that ApoE plays an important role in HCV particle maturation, presumably by direct interaction with viral envelope glycoproteins. IMPORTANCE The HCV replication cycle is tightly linked to host cell lipid pathways and components. This is best illustrated by the dependency of HCV assembly on lipid droplets and the VLDL component ApoE. Although the role of ApoE for production of infectious HCV particles is well established, it is still poorly understood how ApoE contributes to virion formation and how it gets associated with HCV particles. Here, we provide experimental evidence that ApoE likely is required for an intracellular maturation step of HCV particles. Moreover, we demonstrate that ApoE associates with the viral envelope glycoproteins. This interaction appears to be dispensable for envelopment of virus particles but likely contributes to the quality control of secreted infectious virions. These results shed new light on the exploitation of host cell lipid pathways by HCV and the link of viral particle assembly to the VLDL component ApoE. PMID:25122793

Predicting the points of interaction of small molecules in the NF-κB pathway

PubMed Central

2011-01-01

Background The similarity property principle has been used extensively in drug discovery to identify small compounds that interact with specific drug targets. Here we show it can be applied to identify the interactions of small molecules within the NF-κB signalling pathway. Results Clusters that contain compounds with a predominant interaction within the pathway were created, which were then used to predict the interaction of compounds not included in the clustering analysis. Conclusions The technique successfully predicted the points of interactions of compounds that are known to interact with the NF-κB pathway. The method was also shown to be successful when compounds for which the interaction points were unknown were included in the clustering analysis. PMID:21342508

AMP-activated protein kinase is involved in the activation of the Fanconi anemia/BRCA pathway in response to DNA interstrand crosslinks

PubMed Central

Hwang, Soo Kyung; Kim, Bong Sub; Kim, Hyoun Geun; Choi, Hae In; Kim, Jong Heon; Goh, Sung Ho; Lee, Chang-Hun

2016-01-01

Fanconi anemia complementation group (FANC) proteins constitute the Fanconi Anemia (FA)/BRCA pathway that is activated in response to DNA interstrand crosslinks (ICLs). We previously performed yeast two-hybrid screening to identify novel FANC-interacting proteins and discovered that the alpha subunit of AMP-activated protein kinase (AMPKα1) was a candidate binding partner of the FANCG protein, which is a component of the FA nuclear core complex. We confirmed the interaction between AMPKα and both FANCG using co-immunoprecipitation experiments. Additionally, we showed that AMPKα interacted with FANCA, another component of the FA nuclear core complex. AMPKα knockdown in U2OS cells decreased FANCD2 monoubiquitination and nuclear foci formation upon mitomycin C-induced ICLs. Furthermore, AMPKα knockdown enhanced cellular sensitivity to MMC. MMC treatment resulted in an increase in AMPKα phosphorylation/activation, indicating AMPK is involved in the cellular response to ICLs. FANCA was phosphorylated by AMPK at S347 and phosphorylation increased with MMC treatment. MMC-induced FANCD2 monoubiquitination and nuclear foci formation were compromised in a U2OS cell line that stably overexpressed the S347A mutant form of FANCA compared to wild-type FANCA-overexpressing cells, indicating a requirement for FANCA phosphorylation at S347 for proper activation of the FA/BRCA pathway. Our data suggest AMPK is involved in the activation of the FA/BRCA pathway. PMID:27449087

Caenorhabditis elegans ABCRNAi transporters interact genetically with rde-2 and mut-7.

PubMed

Sundaram, Prema; Han, Wang; Cohen, Nancy; Echalier, Benjamin; Albin, John; Timmons, Lisa

2008-02-01

RNA interference (RNAi) mechanisms are conserved and consist of an interrelated network of activities that not only respond to exogenous dsRNA, but also perform endogenous functions required in the fine tuning of gene expression and in maintaining genome integrity. Not surprisingly, RNAi functions have widespread influences on cellular function and organismal development. Previously, we observed a reduced capacity to mount an RNAi response in nine Caenorhabditis elegans mutants that are defective in ABC transporter genes (ABC(RNAi) mutants). Here, we report an exhaustive study of mutants, collectively defective in 49 different ABC transporter genes, that allowed for the categorization of one additional transporter into the ABC(RNAi) gene class. Genetic complementation tests reveal functions for ABC(RNAi) transporters in the mut-7/rde-2 branch of the RNAi pathway. These second-site noncomplementation interactions suggest that ABC(RNAi) proteins and MUT-7/RDE-2 function together in parallel pathways and/or as multiprotein complexes. Like mut-7 and rde-2, some ABC(RNAi) mutants display transposon silencing defects. Finally, our analyses reveal a genetic interaction network of ABC(RNAi) gene function with respect to this part of the RNAi pathway. From our results, we speculate that the coordinated activities of ABC(RNAi) transporters, through their effects on endogenous RNAi-related mechanisms, ultimately affect chromosome function and integrity.

Caenorhabditis elegans ABCRNAi Transporters Interact Genetically With rde-2 and mut-7

PubMed Central

Sundaram, Prema; Han, Wang; Cohen, Nancy; Echalier, Benjamin; Albin, John; Timmons, Lisa

2008-01-01

RNA interference (RNAi) mechanisms are conserved and consist of an interrelated network of activities that not only respond to exogenous dsRNA, but also perform endogenous functions required in the fine tuning of gene expression and in maintaining genome integrity. Not surprisingly, RNAi functions have widespread influences on cellular function and organismal development. Previously, we observed a reduced capacity to mount an RNAi response in nine Caenorhabditis elegans mutants that are defective in ABC transporter genes (ABCRNAi mutants). Here, we report an exhaustive study of mutants, collectively defective in 49 different ABC transporter genes, that allowed for the categorization of one additional transporter into the ABCRNAi gene class. Genetic complementation tests reveal functions for ABCRNAi transporters in the mut-7/rde-2 branch of the RNAi pathway. These second-site noncomplementation interactions suggest that ABCRNAi proteins and MUT-7/RDE-2 function together in parallel pathways and/or as multiprotein complexes. Like mut-7 and rde-2, some ABCRNAi mutants display transposon silencing defects. Finally, our analyses reveal a genetic interaction network of ABCRNAi gene function with respect to this part of the RNAi pathway. From our results, we speculate that the coordinated activities of ABCRNAi transporters, through their effects on endogenous RNAi-related mechanisms, ultimately affect chromosome function and integrity. PMID:18245353

The insulator protein BEAF-32 is required for Hippo pathway activity in the terminal differentiation of neuronal subtypes.

PubMed

Jukam, David; Viets, Kayla; Anderson, Caitlin; Zhou, Cyrus; DeFord, Peter; Yan, Jenny; Cao, Jinshuai; Johnston, Robert J

2016-07-01

The Hippo pathway is crucial for not only normal growth and apoptosis but also cell fate specification during development. What controls Hippo pathway activity during cell fate specification is incompletely understood. In this article, we identify the insulator protein BEAF-32 as a regulator of Hippo pathway activity in Drosophila photoreceptor differentiation. Though morphologically uniform, the fly eye is composed of two subtypes of R8 photoreceptor neurons defined by expression of light-detecting Rhodopsin proteins. In one R8 subtype, active Hippo signaling induces Rhodopsin 6 (Rh6) and represses Rhodopsin 5 (Rh5), whereas in the other subtype, inactive Hippo signaling induces Rh5 and represses Rh6. The activity state of the Hippo pathway in R8 cells is determined by the expression of warts, a core pathway kinase, which interacts with the growth regulator melted in a double-negative feedback loop. We show that BEAF-32 is required for expression of warts and repression of melted Furthermore, BEAF-32 plays a second role downstream of Warts to induce Rh6 and prevent Rh5 fate. BEAF-32 is dispensable for Warts feedback, indicating that BEAF-32 differentially regulates warts and Rhodopsins. Loss of BEAF-32 does not noticeably impair the functions of the Hippo pathway in eye growth regulation. Our study identifies a context-specific regulator of Hippo pathway activity in post-mitotic neuronal fate, and reveals a developmentally specific role for a broadly expressed insulator protein. © 2016. Published by The Company of Biologists Ltd.

Identification of shared and unique susceptibility pathways among cancers of the lung, breast, and prostate from genome-wide association studies and tissue-specific protein interactions

PubMed Central

Qian, David C.; Byun, Jinyoung; Han, Younghun; Greene, Casey S.; Field, John K.; Hung, Rayjean J.; Brhane, Yonathan; Mclaughlin, John R.; Fehringer, Gordon; Landi, Maria Teresa; Rosenberger, Albert; Bickeböller, Heike; Malhotra, Jyoti; Risch, Angela; Heinrich, Joachim; Hunter, David J.; Henderson, Brian E.; Haiman, Christopher A.; Schumacher, Fredrick R.; Eeles, Rosalind A.; Easton, Douglas F.; Seminara, Daniela; Amos, Christopher I.

2015-01-01

Results from genome-wide association studies (GWAS) have indicated that strong single-gene effects are the exception, not the rule, for most diseases. We assessed the joint effects of germline genetic variations through a pathway-based approach that considers the tissue-specific contexts of GWAS findings. From GWAS meta-analyses of lung cancer (12 160 cases/16 838 controls), breast cancer (15 748 cases/18 084 controls) and prostate cancer (14 160 cases/12 724 controls) in individuals of European ancestry, we determined the tissue-specific interaction networks of proteins expressed from genes that are likely to be affected by disease-associated variants. Reactome pathways exhibiting enrichment of proteins from each network were compared across the cancers. Our results show that pathways associated with all three cancers tend to be broad cellular processes required for growth and survival. Significant examples include the nerve growth factor (P = 7.86 × 10−33), epidermal growth factor (P = 1.18 × 10−31) and fibroblast growth factor (P = 2.47 × 10−31) signaling pathways. However, within these shared pathways, the genes that influence risk largely differ by cancer. Pathways found to be unique for a single cancer focus on more specific cellular functions, such as interleukin signaling in lung cancer (P = 1.69 × 10−15), apoptosis initiation by Bad in breast cancer (P = 3.14 × 10−9) and cellular responses to hypoxia in prostate cancer (P = 2.14 × 10−9). We present the largest comparative cross-cancer pathway analysis of GWAS to date. Our approach can also be applied to the study of inherited mechanisms underlying risk across multiple diseases in general. PMID:26483192

Jupyter and Galaxy: Easing entry barriers into complex data analyses for biomedical researchers.

PubMed

Grüning, Björn A; Rasche, Eric; Rebolledo-Jaramillo, Boris; Eberhard, Carl; Houwaart, Torsten; Chilton, John; Coraor, Nate; Backofen, Rolf; Taylor, James; Nekrutenko, Anton

2017-05-01

What does it take to convert a heap of sequencing data into a publishable result? First, common tools are employed to reduce primary data (sequencing reads) to a form suitable for further analyses (i.e., the list of variable sites). The subsequent exploratory stage is much more ad hoc and requires the development of custom scripts and pipelines, making it problematic for biomedical researchers. Here, we describe a hybrid platform combining common analysis pathways with the ability to explore data interactively. It aims to fully encompass and simplify the "raw data-to-publication" pathway and make it reproducible.

Heterogeneous path ensembles for conformational transitions in semi–atomistic models of adenylate kinase

PubMed Central

Bhatt, Divesh; Zuckerman, Daniel M.

2010-01-01

We performed “weighted ensemble” path–sampling simulations of adenylate kinase, using several semi–atomistic protein models. The models have an all–atom backbone with various levels of residue interactions. The primary result is that full statistically rigorous path sampling required only a few weeks of single–processor computing time with these models, indicating the addition of further chemical detail should be readily feasible. Our semi–atomistic path ensembles are consistent with previous biophysical findings: the presence of two distinct pathways, identification of intermediates, and symmetry of forward and reverse pathways. PMID:21660120

General morphological and biological features of neoplasms: integration of molecular findings.

PubMed

Diaz-Cano, S J

2008-07-01

This review highlights the importance of morphology-molecular correlations for a proper implementation of new markers. It covers both general aspects of tumorigenesis (which are normally omitted in papers analysing molecular pathways) and the general mechanisms for the acquired capabilities of neoplasms. The mechanisms are also supported by appropriate diagrams for each acquired capability that include overlooked features such as mobilization of cellular resources and changes in chromatin, transcription and epigenetics; fully accepted oncogenes and tumour suppressor genes are highlighted, while the pathways are also presented as activating or inactivating with appropriate colour coding. Finally, the concepts and mechanisms presented enable us to understand the basic requirements for the appropriate implementation of molecular tests in clinical practice. In summary, the basic findings are presented to serve as a bridge to clinical applications. The current definition of neoplasm is descriptive and difficult to apply routinely. Biologically, neoplasms develop through acquisition of capabilities that involve tumour cell aspects and modified microenvironment interactions, resulting in unrestricted growth due to a stepwise accumulation of cooperative genetic alterations that affect key molecular pathways. The correlation of these molecular aspects with morphological changes is essential for better understanding of essential concepts as early neoplasms/precancerous lesions, progression/dedifferentiation, and intratumour heterogeneity. The acquired capabilities include self-maintained replication (cell cycle dysregulation), extended cell survival (cell cycle arrest, apoptosis dysregulation, and replicative lifespan), genetic instability (chromosomal and microsatellite), changes of chromatin, transcription and epigenetics, mobilization of cellular resources, and modified microenvironment interactions (tumour cells, stromal cells, extracellular, endothelium). The acquired capabilities defining neoplasms are the hallmarks of cancer, but they also comprise useful tools to improve diagnosis and prognosis, as well as potential therapeutic targets. The application of these concepts in oncological pathology leads to consideration of the molecular test requirements (Molecular Test Score System) for reliable implementation; these requirements should cover biological effects, molecular pathway, biological validation, and technical validation. Sensible application of molecular markers in tumour pathology always needs solid morphological support.

Interplay between DISC1 and GABA signaling regulates neurogenesis in mice and risk for schizophrenia.

PubMed

Kim, Ju Young; Liu, Cindy Y; Zhang, Fengyu; Duan, Xin; Wen, Zhexing; Song, Juan; Feighery, Emer; Lu, Bai; Rujescu, Dan; St Clair, David; Christian, Kimberly; Callicott, Joseph H; Weinberger, Daniel R; Song, Hongjun; Ming, Guo-li

2012-03-02

How extrinsic stimuli and intrinsic factors interact to regulate continuous neurogenesis in the postnatal mammalian brain is unknown. Here we show that regulation of dendritic development of newborn neurons by Disrupted-in-Schizophrenia 1 (DISC1) during adult hippocampal neurogenesis requires neurotransmitter GABA-induced, NKCC1-dependent depolarization through a convergence onto the AKT-mTOR pathway. In contrast, DISC1 fails to modulate early-postnatal hippocampal neurogenesis when conversion of GABA-induced depolarization to hyperpolarization is accelerated. Extending the period of GABA-induced depolarization or maternal deprivation stress restores DISC1-dependent dendritic regulation through mTOR pathway during early-postnatal hippocampal neurogenesis. Furthermore, DISC1 and NKCC1 interact epistatically to affect risk for schizophrenia in two independent case control studies. Our study uncovers an interplay between intrinsic DISC1 and extrinsic GABA signaling, two schizophrenia susceptibility pathways, in controlling neurogenesis and suggests critical roles of developmental tempo and experience in manifesting the impact of susceptibility genes on neuronal development and risk for mental disorders. Copyright © 2012 Elsevier Inc. All rights reserved.

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

SPV: a JavaScript Signaling Pathway Visualizer.

PubMed

Calderone, Alberto; Cesareni, Gianni

2018-03-24

The visualization of molecular interactions annotated in web resources is useful to offer to users such information in a clear intuitive layout. These interactions are frequently represented as binary interactions that are laid out in free space where, different entities, cellular compartments and interaction types are hardly distinguishable. SPV (Signaling Pathway Visualizer) is a free open source JavaScript library which offers a series of pre-defined elements, compartments and interaction types meant to facilitate the representation of signaling pathways consisting of causal interactions without neglecting simple protein-protein interaction networks. freely available under Apache version 2 license; Source code: https://github.com/Sinnefa/SPV_Signaling_Pathway_Visualizer_v1.0. Language: JavaScript; Web technology: Scalable Vector Graphics; Libraries: D3.js. sinnefa@gmail.com.

Wnt/β-catenin signaling integrates patterning and metabolism of the insect growth zone.

PubMed

Oberhofer, Georg; Grossmann, Daniela; Siemanowski, Janna L; Beissbarth, Tim; Bucher, Gregor

2014-12-01

Wnt/β-catenin and hedgehog (Hh) signaling are essential for transmitting signals across cell membranes in animal embryos. Early patterning of the principal insect model, Drosophila melanogaster, occurs in the syncytial blastoderm, where diffusion of transcription factors obviates the need for signaling pathways. However, in the cellularized growth zone of typical short germ insect embryos, signaling pathways are predicted to play a more fundamental role. Indeed, the Wnt/β-catenin pathway is required for posterior elongation in most arthropods, although which target genes are activated in this context remains elusive. Here, we use the short germ beetle Tribolium castaneum to investigate two Wnt and Hh signaling centers located in the head anlagen and in the growth zone of early embryos. We find that Wnt/β-catenin signaling acts upstream of Hh in the growth zone, whereas the opposite interaction occurs in the head. We determine the target gene sets of the Wnt/β-catenin and Hh pathways and find that the growth zone signaling center activates a much greater number of genes and that the Wnt and Hh target gene sets are essentially non-overlapping. The Wnt pathway activates key genes of all three germ layers, including pair-rule genes, and Tc-caudal and Tc-twist. Furthermore, the Wnt pathway is required for hindgut development and we identify Tc-senseless as a novel hindgut patterning gene required in the early growth zone. At the same time, Wnt acts on growth zone metabolism and cell division, thereby integrating growth with patterning. Posterior Hh signaling activates several genes potentially involved in a proteinase cascade of unknown function. © 2014. Published by The Company of Biologists Ltd.

Wnt/β-catenin signaling integrates patterning and metabolism of the insect growth zone

PubMed Central

Oberhofer, Georg; Grossmann, Daniela; Siemanowski, Janna L.; Beissbarth, Tim; Bucher, Gregor

2014-01-01

Wnt/β-catenin and hedgehog (Hh) signaling are essential for transmitting signals across cell membranes in animal embryos. Early patterning of the principal insect model, Drosophila melanogaster, occurs in the syncytial blastoderm, where diffusion of transcription factors obviates the need for signaling pathways. However, in the cellularized growth zone of typical short germ insect embryos, signaling pathways are predicted to play a more fundamental role. Indeed, the Wnt/β-catenin pathway is required for posterior elongation in most arthropods, although which target genes are activated in this context remains elusive. Here, we use the short germ beetle Tribolium castaneum to investigate two Wnt and Hh signaling centers located in the head anlagen and in the growth zone of early embryos. We find that Wnt/β-catenin signaling acts upstream of Hh in the growth zone, whereas the opposite interaction occurs in the head. We determine the target gene sets of the Wnt/β-catenin and Hh pathways and find that the growth zone signaling center activates a much greater number of genes and that the Wnt and Hh target gene sets are essentially non-overlapping. The Wnt pathway activates key genes of all three germ layers, including pair-rule genes, and Tc-caudal and Tc-twist. Furthermore, the Wnt pathway is required for hindgut development and we identify Tc-senseless as a novel hindgut patterning gene required in the early growth zone. At the same time, Wnt acts on growth zone metabolism and cell division, thereby integrating growth with patterning. Posterior Hh signaling activates several genes potentially involved in a proteinase cascade of unknown function. PMID:25395458

Viral exploitation of the MEK/ERK pathway - A tale of vaccinia virus and other viruses.

PubMed

Bonjardim, Cláudio A

2017-07-01

The VACV replication cycle is remarkable in the sense that it is performed entirely in the cytoplasmic compartment of vertebrate cells, due to its capability to encode enzymes required either for regulating the macromolecular precursor pool or the biosynthetic processes. Although remarkable, this gene repertoire is not sufficient to confer the status of a free-living microorganism to the virus, and, consequently, the virus relies heavily on the host to successfully generate its progeny. During the complex virus-host interaction, viruses must deal not only with the host pathways to accomplish their temporal demands but also with pathways that counteract viral infection, including the inflammatory, innate and acquired immune responses. This review focuses on VACV and other DNA or RNA viruses that stimulate the MEK (MAPK - Mitogen Activated Protein Kinase)/ERK- Extracellular signal-Regulated Kinase) pathway as part of their replication cycle. Copyright © 2016 Elsevier Inc. All rights reserved.

Molecular Genetics of the PI3K-AKT-mTOR Pathway in Genodermatoses: Diagnostic Implications and Treatment Opportunities.

PubMed

Vahidnezhad, Hassan; Youssefian, Leila; Uitto, Jouni

2016-01-01

A number of critical signaling pathways are required for homeostatic regulation of cell survival, differentiation, and proliferation during organogenesis. One of them is the PI3K-AKT-mTOR pathway consisting of a cascade of inhibitor/activator molecules. Recently, a number of heritable diseases with skin involvement, manifesting particularly with tissue overgrowth, have been shown to result from mutations in the genes in the PI3K-AKT-mTOR and interacting intracellular pathways. Many of these conditions represent an overlapping spectrum of phenotypic manifestations forming a basis for novel, unifying classifications. Identification of the mutant genes and specific mutations in these patients has implications for diagnostics and genetic counseling and provides a rational basis for the development of novel treatment modalities for this currently intractable group of disorders. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

RNA-binding protein GLD-1/quaking genetically interacts with the mir-35 and the let-7 miRNA pathways in Caenorhabditis elegans

PubMed Central

Akay, Alper; Craig, Ashley; Lehrbach, Nicolas; Larance, Mark; Pourkarimi, Ehsan; Wright, Jane E.; Lamond, Angus; Miska, Eric; Gartner, Anton

2013-01-01

Messenger RNA translation is regulated by RNA-binding proteins and small non-coding RNAs called microRNAs. Even though we know the majority of RNA-binding proteins and microRNAs that regulate messenger RNA expression, evidence of interactions between the two remain elusive. The role of the RNA-binding protein GLD-1 as a translational repressor is well studied during Caenorhabditis elegans germline development and maintenance. Possible functions of GLD-1 during somatic development and the mechanism of how GLD-1 acts as a translational repressor are not known. Its human homologue, quaking (QKI), is essential for embryonic development. Here, we report that the RNA-binding protein GLD-1 in C. elegans affects multiple microRNA pathways and interacts with proteins required for microRNA function. Using genome-wide RNAi screening, we found that nhl-2 and vig-1, two known modulators of miRNA function, genetically interact with GLD-1. gld-1 mutations enhance multiple phenotypes conferred by mir-35 and let-7 family mutants during somatic development. We used stable isotope labelling with amino acids in cell culture to globally analyse the changes in the proteome conferred by let-7 and gld-1 during animal development. We identified the histone mRNA-binding protein CDL-1 to be, in part, responsible for the phenotypes observed in let-7 and gld-1 mutants. The link between GLD-1 and miRNA-mediated gene regulation is further supported by its biochemical interaction with ALG-1, CGH-1 and PAB-1, proteins implicated in miRNA regulation. Overall, we have uncovered genetic and biochemical interactions between GLD-1 and miRNA pathways. PMID:24258276

Axonal guidance signaling pathway interacting with smoking in modifying the risk of pancreatic cancer: a gene- and pathway-based interaction analysis of GWAS data.

PubMed

Tang, Hongwei; Wei, Peng; Duell, Eric J; Risch, Harvey A; Olson, Sara H; Bueno-de-Mesquita, H Bas; Gallinger, Steven; Holly, Elizabeth A; Petersen, Gloria; Bracci, Paige M; McWilliams, Robert R; Jenab, Mazda; Riboli, Elio; Tjønneland, Anne; Boutron-Ruault, Marie Christine; Kaaks, Rudolph; Trichopoulos, Dimitrios; Panico, Salvatore; Sund, Malin; Peeters, Petra H M; Khaw, Kay-Tee; Amos, Christopher I; Li, Donghui

2014-05-01

Cigarette smoking is the best established modifiable risk factor for pancreatic cancer. Genetic factors that underlie smoking-related pancreatic cancer have previously not been examined at the genome-wide level. Taking advantage of the existing Genome-wide association study (GWAS) genotype and risk factor data from the Pancreatic Cancer Case Control Consortium, we conducted a discovery study in 2028 cases and 2109 controls to examine gene-smoking interactions at pathway/gene/single nucleotide polymorphism (SNP) level. Using the likelihood ratio test nested in logistic regression models and ingenuity pathway analysis (IPA), we examined 172 KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways, 3 manually curated gene sets, 3 nicotine dependency gene ontology pathways, 17 912 genes and 468 114 SNPs. None of the individual pathway/gene/SNP showed significant interaction with smoking after adjusting for multiple comparisons. Six KEGG pathways showed nominal interactions (P < 0.05) with smoking, and the top two are the pancreatic secretion and salivary secretion pathways (major contributing genes: RAB8A, PLCB and CTRB1). Nine genes, i.e. ZBED2, EXO1, PSG2, SLC36A1, CLSTN1, MTHFSD, FAT2, IL10RB and ATXN2 had P interaction < 0.0005. Five intergenic region SNPs and two SNPs of the EVC and KCNIP4 genes had P interaction < 0.00003. In IPA analysis of genes with nominal interactions with smoking, axonal guidance signaling $$\\left(P=2.12\\times 1{0}^{-7}\\right)$$ and α-adrenergic signaling $$\\left(P=2.52\\times 1{0}^{-5}\\right)$$ genes were significantly overrepresented canonical pathways. Genes contributing to the axon guidance signaling pathway included the SLIT/ROBO signaling genes that were frequently altered in pancreatic cancer. These observations need to be confirmed in additional data set. Once confirmed, it will open a new avenue to unveiling the etiology of smoking-associated pancreatic cancer.

Pathway Interaction Network Analysis Identifies Dysregulated Pathways in Human Monocytes Infected by Listeria monocytogenes.

PubMed

Fan, Wufeng; Zhou, Yuhan; Li, Hao

2017-01-01

In our study, we aimed to extract dysregulated pathways in human monocytes infected by Listeria monocytogenes (LM) based on pathway interaction network (PIN) which presented the functional dependency between pathways. After genes were aligned to the pathways, principal component analysis (PCA) was used to calculate the pathway activity for each pathway, followed by detecting seed pathway. A PIN was constructed based on gene expression profile, protein-protein interactions (PPIs), and cellular pathways. Identifying dysregulated pathways from the PIN was performed relying on seed pathway and classification accuracy. To evaluate whether the PIN method was feasible or not, we compared the introduced method with standard network centrality measures. The pathway of RNA polymerase II pretranscription events was selected as the seed pathway. Taking this seed pathway as start, one pathway set (9 dysregulated pathways) with AUC score of 1.00 was identified. Among the 5 hub pathways obtained using standard network centrality measures, 4 pathways were the common ones between the two methods. RNA polymerase II transcription and DNA replication owned a higher number of pathway genes and DEGs. These dysregulated pathways work together to influence the progression of LM infection, and they will be available as biomarkers to diagnose LM infection.

Qualitative evaluation of a local coronary heart disease treatment pathway: practical implications and theoretical framework

PubMed Central

2012-01-01

Background Coronary heart disease (CHD) is a common medical problem in general practice. Due to its chronic character, shared care of the patient between general practitioner (GP) and cardiologist (C) is required. In order to improve the cooperation between both medical specialists for patients with CHD, a local treatment pathway was developed. The objective of this study was first to evaluate GPs’ opinions regarding the pathway and its practical implications, and secondly to suggest a theoretical framework of the findings by feeding the identified key factors influencing the pathway implementation into a multi-dimensional model. Methods The evaluation of the pathway was conducted in a qualitative design on a sample of 12 pathway developers (8 GPs and 4 cardiologists) and 4 pathway users (GPs). Face-to face interviews, which were aligned with previously conducted studies of the department and assumptions of the theory of planned behaviour (TPB), were performed following a semi-structured interview guideline. These were audio-taped, transcribed verbatim, coded, and analyzed according to the standards of qualitative content analysis. Results We identified 10 frequently mentioned key factors having an impact on the implementation success of the CHD treatment pathway. We thereby differentiated between pathway related (pathway content, effort, individual flexibility, ownership), behaviour related (previous behaviour, support), interaction related (patient, shared care/colleagues), and system related factors (context, health care system). The overall evaluation of the CHD pathway was positive, but did not automatically lead to a change of clinical behaviour as some GPs felt to have already acted as the pathway recommends. Conclusions By providing an account of our experience creating and implementing an intersectoral care pathway for CHD, this study contributes to our knowledge of factors that may influence physicians’ decisions regarding the use of a local treatment pathway. An improved adaptation of the pathway in daily practice might be best achieved by a combined implementation strategy addressing internal and external factors. A simple, direct adaptation regards the design of the pathway material (e.g. layout, PC version), or the embedding of the pathway in another programme, like a Disease Management Programme (DMP). In addition to these practical implications, we propose a theoretical framework to understand the key factors’ influence on the pathway implementation, with the identified factors along the microlevel (pathway related factors), the mesolevel (interaction related factors), and system- related factors along the macrolevel. PMID:22584032

Qualitative evaluation of a local coronary heart disease treatment pathway: practical implications and theoretical framework.

PubMed

Kramer, Lena; Schlößler, Kathrin; Träger, Susanne; Donner-Banzhoff, Norbert

2012-05-14

Coronary heart disease (CHD) is a common medical problem in general practice. Due to its chronic character, shared care of the patient between general practitioner (GP) and cardiologist (C) is required. In order to improve the cooperation between both medical specialists for patients with CHD, a local treatment pathway was developed. The objective of this study was first to evaluate GPs' opinions regarding the pathway and its practical implications, and secondly to suggest a theoretical framework of the findings by feeding the identified key factors influencing the pathway implementation into a multi-dimensional model. The evaluation of the pathway was conducted in a qualitative design on a sample of 12 pathway developers (8 GPs and 4 cardiologists) and 4 pathway users (GPs). Face-to face interviews, which were aligned with previously conducted studies of the department and assumptions of the theory of planned behaviour (TPB), were performed following a semi-structured interview guideline. These were audio-taped, transcribed verbatim, coded, and analyzed according to the standards of qualitative content analysis. We identified 10 frequently mentioned key factors having an impact on the implementation success of the CHD treatment pathway. We thereby differentiated between pathway related (pathway content, effort, individual flexibility, ownership), behaviour related (previous behaviour, support), interaction related (patient, shared care/colleagues), and system related factors (context, health care system). The overall evaluation of the CHD pathway was positive, but did not automatically lead to a change of clinical behaviour as some GPs felt to have already acted as the pathway recommends. By providing an account of our experience creating and implementing an intersectoral care pathway for CHD, this study contributes to our knowledge of factors that may influence physicians' decisions regarding the use of a local treatment pathway. An improved adaptation of the pathway in daily practice might be best achieved by a combined implementation strategy addressing internal and external factors. A simple, direct adaptation regards the design of the pathway material (e.g. layout, PC version), or the embedding of the pathway in another programme, like a Disease Management Programme (DMP). In addition to these practical implications, we propose a theoretical framework to understand the key factors' influence on the pathway implementation, with the identified factors along the microlevel (pathway related factors), the mesolevel (interaction related factors), and system- related factors along the macrolevel.

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.

Consortium analysis of gene and gene-folate interactions in purine and pyrimidine metabolism pathways with ovarian carcinoma risk

PubMed Central

Kelemen, Linda E.; Terry, Kathryn L.; Goodman, Marc T.; Webb, Penelope M.; Bandera, Elisa V.; McGuire, Valerie; Rossing, Mary Anne; Wang, Qinggang; Dicks, Ed; Tyrer, Jonathan P.; Song, Honglin; Kupryjanczyk, Jolanta; Dansonka-Mieszkowska, Agnieszka; Plisiecka-Halasa, Joanna; Timorek, Agnieszka; Menon, Usha; Gentry-Maharaj, Aleksandra; Gayther, Simon A.; Ramus, Susan J.; Narod, Steven A.; Risch, Harvey A.; McLaughlin, John R.; Siddiqui, Nadeem; Glasspool, Rosalind; Paul, James; Carty, Karen; Gronwald, Jacek; Lubiński, Jan; Jakubowska, Anna; Cybulski, Cezary; Kiemeney, Lambertus A.; Massuger, Leon F. A. G.; van Altena, Anne M.; Aben, Katja K. H.; Olson, Sara H.; Orlow, Irene; Cramer, Daniel W.; Levine, Douglas A.; Bisogna, Maria; Giles, Graham G.; Southey, Melissa C.; Bruinsma, Fiona; Kjær, Susanne Krüger; Høgdall, Estrid; Jensen, Allan; Høgdall, Claus K.; Lundvall, Lene; Engelholm, Svend-Aage; Heitz, Florian; du Bois, Andreas; Harter, Philipp; Schwaab, Ira; Butzow, Ralf; Nevanlinna, Heli; Pelttari, Liisa M.; Leminen, Arto; Thompson, Pamela J.; Lurie, Galina; Wilkens, Lynne R.; Lambrechts, Diether; Van Nieuwenhuysen, Els; Lambrechts, Sandrina; Vergote, Ignace; Beesley, Jonathan; Fasching, Peter A.; Beckmann, Matthias W.; Hein, Alexander; Ekici, Arif B.; Doherty, Jennifer A.; Wu, Anna H.; Pearce, Celeste L.; Pike, Malcolm C.; Stram, Daniel; Chang-Claude, Jenny; Rudolph, Anja; Dörk, Thilo; Dürst, Matthias; Hillemanns, Peter; Runnebaum, Ingo B.; Bogdanova, Natalia; Antonenkova, Natalia; Odunsi, Kunle; Edwards, Robert P.; Kelley, Joseph L.; Modugno, Francesmary; Ness, Roberta B.; Karlan, Beth Y.; Walsh, Christine; Lester, Jenny; Orsulic, Sandra; Fridley, Brooke L.; Vierkant, Robert A.; Cunningham, Julie M.; Wu, Xifeng; Lu, Karen; Liang, Dong; Hildebrandt, Michelle A.T.; Weber, Rachel Palmieri; Iversen, Edwin S.; Tworoger, Shelley S.; Poole, Elizabeth M.; Salvesen, Helga B.; Krakstad, Camilla; Bjorge, Line; Tangen, Ingvild L.; Pejovic, Tanja; Bean, Yukie; Kellar, Melissa; Wentzensen, Nicolas; Brinton, Louise A.; Lissowska, Jolanta; Garcia-Closas, Montserrat; Campbell, Ian G.; Eccles, Diana; Whittemore, Alice S.; Sieh, Weiva; Rothstein, Joseph H.; Anton-Culver, Hoda; Ziogas, Argyrios; Phelan, Catherine M.; Moysich, Kirsten B.; Goode, Ellen L.; Schildkraut, Joellen M.; Berchuck, Andrew; Pharoah, Paul D.P.; Sellers, Thomas A.; Brooks-Wilson, Angela; Cook, Linda S.; Le, Nhu D.

2014-01-01

Scope We re-evaluated previously reported associations between variants in pathways of one-carbon (folate) transfer genes and ovarian carcinoma (OC) risk, and in related pathways of purine and pyrimidine metabolism, and assessed interactions with folate intake. Methods and Results Odds ratios (OR) for 446 genetic variants were estimated among 13,410 OC cases and 22,635 controls and among 2,281 cases and 3,444 controls with folate information. Following multiple testing correction, the most significant main effect associations were for DPYD variants rs11587873 (OR=0.92, P=6x10−5) and rs828054 (OR=1.06, P=1x10−4). Thirteen variants in the pyrimidine metabolism genes, DPYD, DPYS, PPAT and TYMS, also interacted significantly with folate in a multi-variant analysis (corrected P=9.9x10−6) but collectively explained only 0.2% of OC risk. Although no other associations were significant after multiple testing correction, variants in SHMT1 in one-carbon transfer, previously reported with OC, suggested lower risk at higher folate (Pinteraction=0.03-0.006). Conclusions Variation in pyrimidine metabolism genes, particularly DPYD, which was previously reported to be associated with OC, may influence risk; however, stratification by folate intake is unlikely to modify disease risk appreciably in these women. SHMT1 SNP-byfolate interactions are plausible but require further validation. Polymorphisms in selected genes in purine metabolism were not associated with OC. PMID:25066213

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

PubMed Central

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

2016-01-01

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

TGMI: an efficient algorithm for identifying pathway regulators through evaluation of triple-gene mutual interaction

PubMed Central

Gunasekara, Chathura; Zhang, Kui; Deng, Wenping; Brown, Laura

2018-01-01

Abstract Despite their important roles, the regulators for most metabolic pathways and biological processes remain elusive. Presently, the methods for identifying metabolic pathway and biological process regulators are intensively sought after. We developed a novel algorithm called triple-gene mutual interaction (TGMI) for identifying these regulators using high-throughput gene expression data. It first calculated the regulatory interactions among triple gene blocks (two pathway genes and one transcription factor (TF)), using conditional mutual information, and then identifies significantly interacted triple genes using a newly identified novel mutual interaction measure (MIM), which was substantiated to reflect strengths of regulatory interactions within each triple gene block. The TGMI calculated the MIM for each triple gene block and then examined its statistical significance using bootstrap. Finally, the frequencies of all TFs present in all significantly interacted triple gene blocks were calculated and ranked. We showed that the TFs with higher frequencies were usually genuine pathway regulators upon evaluating multiple pathways in plants, animals and yeast. Comparison of TGMI with several other algorithms demonstrated its higher accuracy. Therefore, TGMI will be a valuable tool that can help biologists to identify regulators of metabolic pathways and biological processes from the exploded high-throughput gene expression data in public repositories. PMID:29579312

Requirement of zebrafish pcdh10a and pcdh10b in melanocyte precursor migration.

PubMed

Williams, Jason S; Hsu, Jessica Y; Rossi, Christy Cortez; Artinger, Kristin Bruk

2018-03-29

Melanocytes derive from neural crest cells, which are a highly migratory population of cells that play an important role in pigmentation of the skin and epidermal appendages. In most vertebrates, melanocyte precursor cells migrate solely along the dorsolateral pathway to populate the skin. However, zebrafish melanocyte precursors also migrate along the ventromedial pathway, in route to the yolk, where they interact with other neural crest derivative populations. Here, we demonstrate the requirement for zebrafish paralogs pcdh10a and pcdh10b in zebrafish melanocyte precursor migration. pcdh10a and pcdh10b are expressed in a subset of melanocyte precursor and somatic cells respectively, and knockdown and TALEN mediated gene disruption of pcdh10a results in aberrant migration of melanocyte precursors resulting in fully melanized melanocytes that differentiate precociously in the ventromedial pathway. Live cell imaging analysis demonstrates that loss of pchd10a results in a reduction of directed cell migration of melanocyte precursors, caused by both increased adhesion and a loss of cell-cell contact with other migratory neural crest cells. Also, we determined that the paralog pcdh10b is upregulated and can compensate for the genetic loss of pcdh10a. Disruption of pcdh10b alone by CRISPR mutagenesis results in somite defects, while the loss of both paralogs results in enhanced migratory melanocyte precursor phenotype and embryonic lethality. These results reveal a novel role for pcdh10a and pcdh10b in zebrafish melanocyte precursor migration and suggest that pcdh10 paralogs potentially interact for proper transient migration along the ventromedial pathway. Copyright © 2018 Elsevier Inc. All rights reserved.

The chaperone-histone partnership: for the greater good of histone traffic and chromatin plasticity.

PubMed

Hondele, Maria; Ladurner, Andreas G

2011-12-01

Histones are highly positively charged proteins that wrap our genome. Their surface properties also make them prone to nonspecific interactions and aggregation. A class of proteins known as histone chaperones is dedicated to safeguard histones by aiding their proper incorporation into nucleosomes. Histone chaperones facilitate ordered nucleosome assembly and disassembly reactions through the formation of semi-stable histone-chaperone intermediates without requiring ATP, but merely providing a complementary protein surface for histones to dynamically interact with. Recurrent 'chaperoning' mechanisms involve the masking of the histone's positive charge and the direct blocking of crucial histone surface sites, including those required for H3-H4 tetramerization or the binding of nucleosomal DNA. This shielding prevents histones from engaging in premature or unwanted interactions with nucleic acids and other cellular components. In this review, we analyze recent structural studies on chaperone-histone interactions and discuss the implications of this vital partnership for nucleosome assembly and disassembly pathways. Copyright © 2011 Elsevier Ltd. All rights reserved.

Mediator-dependent Nuclear Receptor Functions

PubMed Central

Chen, Wei; Roeder, Robert

2011-01-01

As gene-specific transcription factors, nuclear hormone receptors are broadly involved in many important biological processes. Their function on target genes requires the stepwise assembly of different coactivator complexes that facilitate chromatin remodeling and subsequent preinitiation complex (PIC) formation and function. Mediator has proved to be a crucial, and general, nuclear receptor-interacting coactivator, with demonstrated functions in transcription steps ranging from chromatin remodeling to subsequent PIC formation and function. Here we discuss (i) our current understanding of pathways that nuclear receptors and other interacting cofactors employ to recruit Mediator to target gene enhancers and promoters, including conditional requirements for the strong NR-Mediator interactions mediated by the NR AF2 domain and the MED1 LXXLLL motifs and (ii) mechanisms by which Mediator acts to transmit signals from enhancer-bound nuclear receptors to the general transcription machinery at core promoters to effect PIC formation and function. PMID:21854863

Competition for vitamin B1 (thiamin) structures numerous ecological interactions.

PubMed

Kraft, Clifford E; Angert, Esther R

2017-06-01

Thiamin (vitamin B1) is a cofactor required for essential biochemical reactions in all living organisms, yet free thiamin is scarce in the environment. The diversity of biochemical pathways involved in the acquisition, degradation, and synthesis of thiamin indicates that organisms have evolved numerous ecological strategies for meeting this nutritional requirement. In this review we synthesize information from multiple disciplines to show how the complex biochemistry of thiamin influences ecological outcomes of interactions between organisms in environments ranging from the open ocean and the Australian outback to the gastrointestinal tract of animals. We highlight population and ecosystem responses to the availability or absence of thiamin. These include widespread mortality of fishes, birds, and mammals, as well as the thiamin-dependent regulation of ocean productivity. Overall, we portray thiamin biochemistry as the foundation for molecularly mediated ecological interactions that influence survival and abundance of a vast array of organisms.

Root Systems Biology: Integrative Modeling across Scales, from Gene Regulatory Networks to the Rhizosphere1

PubMed Central

Hill, Kristine; Porco, Silvana; Lobet, Guillaume; Zappala, Susan; Mooney, Sacha; Draye, Xavier; Bennett, Malcolm J.

2013-01-01

Genetic and genomic approaches in model organisms have advanced our understanding of root biology over the last decade. Recently, however, systems biology and modeling have emerged as important approaches, as our understanding of root regulatory pathways has become more complex and interpreting pathway outputs has become less intuitive. To relate root genotype to phenotype, we must move beyond the examination of interactions at the genetic network scale and employ multiscale modeling approaches to predict emergent properties at the tissue, organ, organism, and rhizosphere scales. Understanding the underlying biological mechanisms and the complex interplay between systems at these different scales requires an integrative approach. Here, we describe examples of such approaches and discuss the merits of developing models to span multiple scales, from network to population levels, and to address dynamic interactions between plants and their environment. PMID:24143806

Sestrin2 is a leucine sensor for the mTORC1 pathway

PubMed Central

Wolfson, Rachel L.; Chantranupong, Lynne; Saxton, Robert A.; Shen, Kuang; Scaria, Sonia M.; Cantor, Jason R.; Sabatini, David M.

2015-01-01

Leucine is a proteogenic amino acid that also regulates many aspects of mammalian physiology, in large part by activating the mTOR complex 1 (mTORC1) protein kinase, a master growth controller. Amino acids signal to mTORC1 through the Rag guanine triphosphatases (GTPases). Several factors regulate the Rags, including GATOR1, a GTPase activating protein (GAP); GATOR2, a positive regulator of unknown function; and Sestrin2, a GATOR2-interacting protein that inhibits mTORC1 signaling. We find that leucine, but not arginine, disrupts the Sestrin2-GATOR2 interaction by binding to Sestrin2 with a Kd of 20 µM, which is the leucine concentration that half-maximally activates mTORC1. The leucine-binding capacity of Sestrin2 is required for leucine to activate mTORC1 in cells. These results indicate that Sestrin2 is a leucine sensor for the mTORC1 pathway. PMID:26449471

Insulin receptors internalize by a rapid, saturable pathway requiring receptor autophosphorylation and an intact juxtamembrane region

PubMed Central

1991-01-01

The effect of receptor occupancy on insulin receptor endocytosis was examined in CHO cells expressing normal human insulin receptors (CHO/IR), autophosphorylation- and internalization-deficient receptors (CHO/IRA1018), and receptors which undergo autophosphorylation but lack a sequence required for internalization (CHO/IR delta 960). The rate of [125I]insulin internalization in CHO/IR cells at 37 degrees C was rapid at physiological concentrations, but decreased markedly in the presence of increasing unlabeled insulin (ED50 = 1-3 nM insulin, or 75,000 occupied receptors/cell). In contrast, [125I]insulin internalization by CHO/IRA1018 and CHO/IR delta 960 cells was slow and was not inhibited by unlabeled insulin. At saturating insulin concentrations, the rate of internalization by wild-type and mutant receptors was similar. Moreover, depletion of intracellular potassium, which has been shown to disrupt coated pit formation, inhibited the rapid internalization of [125I]insulin at physiological insulin concentrations by CHO/IR cells, but had little or no effect on [125I]insulin uptake by CHO/IR delta 960 and CHO/IRA1018 cells or wild-type cells at high insulin concentrations. These data suggest that the insulin-stimulated entry of the insulin receptor into a rapid, coated pit-mediated internalization pathway is saturable and requires receptor autophosphorylation and an intact juxtamembrane region. Furthermore, CHO cells also contain a constitutive nonsaturable pathway which does not require receptor autophosphorylation or an intact juxtamembrane region; this second pathway is unaffected by depletion of intracellular potassium, and therefore may be independent of coated pits. Our data suggest that the ligand-stimulated internalization of the insulin receptor may require specific saturable interactions between the receptor and components of the endocytic system. PMID:1757462

Minimal metabolic pathway structure is consistent with associated biomolecular interactions

PubMed Central

Bordbar, Aarash; Nagarajan, Harish; Lewis, Nathan E; Latif, Haythem; Ebrahim, Ali; Federowicz, Stephen; Schellenberger, Jan; Palsson, Bernhard O

2014-01-01

Pathways are a universal paradigm for functionally describing cellular processes. Even though advances in high-throughput data generation have transformed biology, the core of our biological understanding, and hence data interpretation, is still predicated on human-defined pathways. Here, we introduce an unbiased, pathway structure for genome-scale metabolic networks defined based on principles of parsimony that do not mimic canonical human-defined textbook pathways. Instead, these minimal pathways better describe multiple independent pathway-associated biomolecular interaction datasets suggesting a functional organization for metabolism based on parsimonious use of cellular components. We use the inherent predictive capability of these pathways to experimentally discover novel transcriptional regulatory interactions in Escherichia coli metabolism for three transcription factors, effectively doubling the known regulatory roles for Nac and MntR. This study suggests an underlying and fundamental principle in the evolutionary selection of pathway structures; namely, that pathways may be minimal, independent, and segregated. PMID:24987116

Dysregulation of Uterine Signaling Pathways in Progesterone Receptor-Cre Knockout of Dicer

PubMed Central

Andreu-Vieyra, Claudia V.; Kim, Tae Hoon; Jeong, Jae-Wook; Hodgson, Myles C.; Chen, Ruihong; Creighton, Chad J.; Lydon, John P.; Gunaratne, Preethi H.; DeMayo, Francesco J.; Matzuk, Martin M.

2012-01-01

Epithelial-stromal interactions in the uterus are required for normal uterine functions such as pregnancy, and multiple signaling pathways are essential for this process. Although Dicer and microRNA (miRNA) have been implicated in several reproductive processes, the specific roles of Dicer and miRNA in uterine development are not known. To address the roles of miRNA in the regulation of key uterine pathways, we generated a conditional knockout of Dicer in the postnatal uterine epithelium and stroma using progesterone receptor-Cre. These Dicer conditional knockout females are sterile with small uteri, which demonstrate significant defects, including absence of glandular epithelium and enhanced stromal apoptosis, beginning at approximately postnatal d 15, with coincident expression of Cre and deletion of Dicer. Specific miRNA (miR-181c, −200b, −101, let-7d) were down-regulated and corresponding predicted proapoptotic target genes (Bcl2l11, Aldh1a3) were up-regulated, reflecting the apoptotic phenomenon. Although these mice had normal serum hormone levels, critical uterine signaling pathways, including progesterone-responsive genes, Indian hedgehog signaling, and the Wnt/β-catenin canonical pathway, were dysregulated at the mRNA level. Importantly, uterine stromal cell proliferation in response to progesterone was absent, whereas uterine epithelial cell proliferation in response to estradiol was maintained in adult uteri. These data implicate Dicer and appropriate miRNA expression as essential players in the regulation of multiple uterine signaling pathways required for uterine development and appropriate function. PMID:22798293

The Tandem CARDs of NOD2: Intramolecular Interactions and Recognition of RIP2

PubMed Central

Fridh, Veronica; Rittinger, Katrin

2012-01-01

Caspase recruitment domains (CARDs) are homotypic protein interaction modules that link the stimulus-dependent assembly of large signaling platforms such as inflammasomes to the activation of downstream effectors that often include caspases and kinases and thereby play an important role in the regulation of inflammatory and apoptotic signaling pathways. NOD2 belongs to the NOD-like (NLR) family of intracellular pattern recognition receptors (PRR) and induces activation of the NF-κB pathway in response to the recognition of bacterial components. This process requires the specific recognition of the CARD of the protein kinase RIP2 by the tandem CARDs of NOD2. Here we demonstrate that the tandem CARDs of NOD2 are engaged in an intramolecular interaction that is important for the structural stability of this region. Using a combination of ITC and pull-down experiments we identify distinct surface areas that are involved in the intramolecular tandem CARD interaction and the interaction with the downstream effector RIP2. Our findings indicate that while CARDa of NOD2 might be the primary binding partner of RIP2 the two CARDs of NOD2 do not act independently of one another but may cooperate to from a binding surface that is distinct from that of single CARDs. PMID:22470564

Topological Transitions in Mitochondrial Membranes controlled by Apoptotic Proteins

NASA Astrophysics Data System (ADS)

Hwee Lai, Ghee; Sanders, Lori K.; Mishra, Abhijit; Schmidt, Nathan W.; Wong, Gerard C. L.; Ivashyna, Olena; Schlesinger, Paul H.

2010-03-01

The Bcl-2 family comprises pro-apoptotic proteins, capable of permeabilizing the mitochondrial membrane, and anti-apoptotic members interacting in an antagonistic fashion to regulate programmed cell death (apoptosis). They offer potential therapeutic targets to re-engage cellular suicide in tumor cells but the extensive network of implicated protein-protein interactions has impeded full understanding of the decision pathway. We show, using synchrotron x-ray diffraction, that pro-apoptotic proteins interact with mitochondrial-like model membranes to generate saddle-splay (negative Gaussian) curvature topologically required for pore formation, while anti-apoptotic proteins can deactivate curvature generation by molecules drastically different from Bcl-2 family members and offer evidence for membrane-curvature mediated interactions general enough to affect very disparate systems.

Studying emotion theories through connectivity analysis: Evidence from generalized psychophysiological interactions and graph theory.

PubMed

Huang, Yun-An; Jastorff, Jan; Van den Stock, Jan; Van de Vliet, Laura; Dupont, Patrick; Vandenbulcke, Mathieu

2018-05-15

Psychological construction models of emotion state that emotions are variable concepts constructed by fundamental psychological processes, whereas according to basic emotion theory, emotions cannot be divided into more fundamental units and each basic emotion is represented by a unique and innate neural circuitry. In a previous study, we found evidence for the psychological construction account by showing that several brain regions were commonly activated when perceiving different emotions (i.e. a general emotion network). Moreover, this set of brain regions included areas associated with core affect, conceptualization and executive control, as predicted by psychological construction models. Here we investigate directed functional brain connectivity in the same dataset to address two questions: 1) is there a common pathway within the general emotion network for the perception of different emotions and 2) if so, does this common pathway contain information to distinguish between different emotions? We used generalized psychophysiological interactions and information flow indices to examine the connectivity within the general emotion network. The results revealed a general emotion pathway that connects neural nodes involved in core affect, conceptualization, language and executive control. Perception of different emotions could not be accurately classified based on the connectivity patterns from the nodes of the general emotion pathway. Successful classification was achieved when connections outside the general emotion pathway were included. We propose that the general emotion pathway functions as a common pathway within the general emotion network and is involved in shared basic psychological processes across emotions. However, additional connections within the general emotion network are required to classify different emotions, consistent with a constructionist account. Copyright © 2018 Elsevier Inc. All rights reserved.

Casein Kinase 2 Reverses Tail-Independent Inactivation of Kinesin-1

NASA Astrophysics Data System (ADS)

Xu, Jing

2013-03-01

Kinesin-1 is a plus-end microtubule-based motor, and defects in kinesin-based transport are linked to diseases including neurodegeneration. Kinesin can auto-inhibit via a head-tail interaction, but is believed to be active otherwise. Here we report a tail-independent inactivation of kinesin, reversible by the disease-relevant signalling protein, casein kinase 2 (CK2). The majority of initially active kinesin (native or tail-less) loses its ability to interact with microtubules in vitro, and CK2 reverses this inactivation (approximately fourfold) without altering kinesin's single motor properties. This activation pathway does not require motor phosphorylation, and is independent of head-tail auto-inhibition. In cultured mammalian cells, reducing CK2 expression, but not its kinase activity, decreases the force required to stall lipid droplet transport, consistent with a decreased number of active kinesin motors. Our results (Nat. Commun., 3:754, 2012) provide the first direct evidence of a protein kinase upregulating kinesin-based transport, and suggest a novel pathway for regulating the activity of cargo-bound kinesin. Work supported by NIGMS grants GM64624 to SPG, GM74830-06A1 to LH, GM76516 to LB, NS048501 to SJK, and AHA grant 825278F to JX.

An allelic series reveals essential roles for FY in plant development in addition to flowering-time control.

PubMed

Henderson, Ian R; Liu, Fuquan; Drea, Sinead; Simpson, Gordon G; Dean, Caroline

2005-08-01

The autonomous pathway functions to promote flowering in Arabidopsis by limiting the accumulation of the floral repressor FLOWERING LOCUS C (FLC). Within this pathway FCA is a plant-specific, nuclear RNA-binding protein, which interacts with FY, a highly conserved eukaryotic polyadenylation factor. FCA and FY function to control polyadenylation site choice during processing of the FCA transcript. Null mutations in the yeast FY homologue Pfs2p are lethal. This raises the question as to whether these essential RNA processing functions are conserved in plants. Characterisation of an allelic series of fy mutations reveals that null alleles are embryo lethal. Furthermore, silencing of FY, but not FCA, is deleterious to growth in Nicotiana. The late-flowering fy alleles are hypomorphic and indicate a requirement for both intact FY WD repeats and the C-terminal domain in repression of FLC. The FY C-terminal domain binds FCA and in vitro assays demonstrate a requirement for both C-terminal FY-PPLPP repeats during this interaction. The expression domain of FY supports its roles in essential and flowering-time functions. Hence, FY may mediate both regulated and constitutive RNA 3'-end processing.

HEXIM1 and NEAT1 Long Non-coding RNA Form a Multi-subunit Complex that Regulates DNA-Mediated Innate Immune Response.

PubMed

Morchikh, Mehdi; Cribier, Alexandra; Raffel, Raoul; Amraoui, Sonia; Cau, Julien; Severac, Dany; Dubois, Emeric; Schwartz, Olivier; Bennasser, Yamina; Benkirane, Monsef

2017-08-03

The DNA-mediated innate immune response underpins anti-microbial defenses and certain autoimmune diseases. Here we used immunoprecipitation, mass spectrometry, and RNA sequencing to identify a ribonuclear complex built around HEXIM1 and the long non-coding RNA NEAT1 that we dubbed the HEXIM1-DNA-PK-paraspeckle components-ribonucleoprotein complex (HDP-RNP). The HDP-RNP contains DNA-PK subunits (DNAPKc, Ku70, and Ku80) and paraspeckle proteins (SFPQ, NONO, PSPC1, RBM14, and MATRIN3). We show that binding of HEXIM1 to NEAT1 is required for its assembly. We further demonstrate that the HDP-RNP is required for the innate immune response to foreign DNA, through the cGAS-STING-IRF3 pathway. The HDP-RNP interacts with cGAS and its partner PQBP1, and their interaction is remodeled by foreign DNA. Remodeling leads to the release of paraspeckle proteins, recruitment of STING, and activation of DNAPKc and IRF3. Our study establishes the HDP-RNP as a key nuclear regulator of DNA-mediated activation of innate immune response through the cGAS-STING pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Diet-gene interactions underlie metabolic individuality and influence brain development: implications for clinical practice derived from studies on choline metabolism.

PubMed

Zeisel, Steven H

2012-01-01

One of the underlying mechanisms for metabolic individuality is genetic variation. Single nucleotide polymorphisms (SNPs) in genes of metabolic pathways can create metabolic inefficiencies that alter the dietary requirement for, and responses to, nutrients. These SNPs can be detected using genetic profiling and the metabolic inefficiencies they cause can be detected using metabolomic profiling. Studies on the human dietary requirement for choline illustrate how useful these new approaches can be, as this requirement is influenced by SNPs in genes of choline and folate metabolism. In adults, these SNPs determine whether people develop fatty liver, liver damage and muscle damage when eating diets low in choline. Because choline is very important for fetal development, these SNPs may identify women who need to eat more choline during pregnancy. Some of the actions of choline are mediated by epigenetic mechanisms that permit 'retuning' of metabolic pathways during early life. Copyright © 2012 S. Karger AG, Basel.

Loss of CHK1 function impedes DNA damage-induced FANCD2 monoubiquitination but normalizes the abnormal G2 arrest in Fanconi anemia.

PubMed

Guervilly, Jean-Hugues; Macé-Aimé, Gaëtane; Rosselli, Filippo

2008-03-01

Fanconi anemia (FA) is a cancer-prone hereditary disease resulting from mutations in one of the 13 genes defining the FANC/BRCA pathway. This pathway is involved in the cellular resistance to DNA-cross-linking agents. How the FANC/BRCA pathway is activated and why its deficiency leads to the accumulation of FA cells with a 4N DNA content are still poorly answered questions. We investigated the involvement of ATR pathway members in these processes. We show here that RAD9 and RAD17 are required for DNA interstrand cross-link (ICL) resistance and for the optimal activation of FANCD2. Moreover, we demonstrate that CHK1 and its interacting partner CLASPIN that act downstream in the ATR pathway are required for both FANCD2 monoubiquitination and assembling in subnuclear foci in response to DNA damage. Paradoxically, in the absence of any genotoxic stress, CHK1 or CLASPIN depletion results in an increased basal level of FANCD2 monoubiquitination and focalization. We also demonstrate that the ICL-induced accumulation of FA cells in late S/G2 phase is dependent on ATR and CHK1. In agreement with this, CHK1 phosphorylation is enhanced in FA cells, and chemical inhibition of the ATR/CHK1 axis in FA lymphoblasts decreases their sensitivity to mitomycin C. In conclusion, this work describes a complex crosstalk between CHK1 and the FANC/BRCA pathway: CHK1 activates this pathway through FANCD2 monoubiquitination, whereas FA deficiency leads to a CHK1-dependent G2 accumulation, raising the possibility that the FANC/BRCA pathway downregulates CHK1 activation.

Additional targets of the Arabidopsis autonomous pathway members, FCA and FY.

PubMed

Marquardt, S; Boss, P K; Hadfield, J; Dean, C

2006-01-01

A central player in the Arabidopsis floral transition is the floral repressor FLC, the MADS-box transcriptional regulator that inhibits the activity of genes required to switch the meristem from vegetative to floral development. One of the many pathways that regulate FLC expression is the autonomous promotion pathway composed of FCA, FY, FLD, FPA, FVE, LD, and FLK. Rather than a hierarchical set of activities the autonomous promotion pathway comprises sub-pathways of genes with different biochemical functions that all share FLC as a target. One sub-pathway involves FCA and FY, which interact to regulate RNA processing of FLC. Several of the identified components (FY, FVE, and FLD) are homologous to yeast and mammalian proteins with rather generic roles in gene regulation. So why do mutations in these genes specifically show a late-flowering phenotype in Arabidopsis? One reason, found during the analysis of fy alleles, is that the mutant alleles identified in flowering screens can be hypomorphic, they still have partial function. A broader role for the autonomous promotion pathway is supported by a microarray analysis which has identified genes mis-regulated in fca mutants, and whose expression is also altered in fy mutants.

cPath: open source software for collecting, storing, and querying biological pathways.

PubMed

Cerami, Ethan G; Bader, Gary D; Gross, Benjamin E; Sander, Chris

2006-11-13

Biological pathways, including metabolic pathways, protein interaction networks, signal transduction pathways, and gene regulatory networks, are currently represented in over 220 diverse databases. These data are crucial for the study of specific biological processes, including human diseases. Standard exchange formats for pathway information, such as BioPAX, CellML, SBML and PSI-MI, enable convenient collection of this data for biological research, but mechanisms for common storage and communication are required. We have developed cPath, an open source database and web application for collecting, storing, and querying biological pathway data. cPath makes it easy to aggregate custom pathway data sets available in standard exchange formats from multiple databases, present pathway data to biologists via a customizable web interface, and export pathway data via a web service to third-party software, such as Cytoscape, for visualization and analysis. cPath is software only, and does not include new pathway information. Key features include: a built-in identifier mapping service for linking identical interactors and linking to external resources; built-in support for PSI-MI and BioPAX standard pathway exchange formats; a web service interface for searching and retrieving pathway data sets; and thorough documentation. The cPath software is freely available under the LGPL open source license for academic and commercial use. cPath is a robust, scalable, modular, professional-grade software platform for collecting, storing, and querying biological pathways. It can serve as the core data handling component in information systems for pathway visualization, analysis and modeling.

Methylation interactions in Arabidopsis hybrids require RNA-directed DNA methylation and are influenced by genetic variation

PubMed Central

Zhang, Qingzhu; Wang, Dong; Lang, Zhaobo; He, Li; Yang, Lan; Zeng, Liang; Li, Yanqiang; Zhao, Cheng; Huang, Huan; Zhang, Heng; Zhang, Huiming; Zhu, Jian-Kang

2016-01-01

DNA methylation is a conserved epigenetic mark in plants and many animals. How parental alleles interact in progeny to influence the epigenome is poorly understood. We analyzed the DNA methylomes of Arabidopsis Col and C24 ecotypes, and their hybrid progeny. Hybrids displayed nonadditive DNA methylation levels, termed methylation interactions, throughout the genome. Approximately 2,500 methylation interactions occurred at regions where parental DNA methylation levels are similar, whereas almost 1,000 were at differentially methylated regions in parents. Methylation interactions were characterized by an abundance of 24-nt small interfering RNAs. Furthermore, dysfunction of the RNA-directed DNA methylation pathway abolished methylation interactions but did not affect the increased biomass observed in hybrid progeny. Methylation interactions correlated with altered genetic variation within the genome, suggesting that they may play a role in genome evolution. PMID:27382183

The F-box-containing protein UFO and AGAMOUS participate in antagonistic pathways governing early petal development in Arabidopsis.

PubMed

Durfee, Tim; Roe, Judith L; Sessions, R Allen; Inouye, Carla; Serikawa, Kyle; Feldmann, Kenneth A; Weigel, Detlef; Zambryski, Patricia C

2003-07-08

The UNUSUAL FLORAL ORGANS (UFO) gene is required for multiple processes in the developing Arabidopsis flower, including the proper patterning and identity of both petals and stamens. The gene encodes an F-box-containing protein, UFO, which interacts physically and genetically with the Skp1 homolog, ASK1. In this report, we describe four ufo alleles characterized by the absence of petals, which uncover another role for UFO in promoting second whorl development. This UFO-dependent pathway is required regardless of the second whorl organ to be formed, arguing that it affects a basic process acting in parallel with those establishing organ identity. However, the pathway is dispensable in the absence of AGAMOUS (AG), a known inhibitor of petal development. In situ hybridization results argue that AG is not transcribed in the petal region, suggesting that it acts non-cell-autonomously to inhibit second whorl development in ufo mutants. These results are combined into a genetic model explaining early second whorl initiation/proliferation, in which UFO functions to inhibit an AG-dependent activity.

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

Proteases for Processing Proneuropeptides into Peptide Neurotransmitters and Hormones

PubMed Central

Hook, Vivian; Funkelstein, Lydiane; Lu, Douglas; Bark, Steven; Wegrzyn, Jill; Hwang, Shin-Rong

2009-01-01

Peptide neurotransmitters and peptide hormones, collectively known as neuropeptides, are required for cell-cell communication in neurotransmission and for regulation of endocrine functions. Neuropeptides are synthesized from protein precursors (termed proneuropeptides or prohormones) that require proteolytic processing primarily within secretory vesicles that store and secrete the mature neuropeptides to control target cellular and organ systems. This review describes interdisciplinary strategies that have elucidated two primary protease pathways for prohormone processing consisting of the cysteine protease pathway mediated by secretory vesicle cathepsin L and the well-known subtilisin-like proprotein convertase pathway that together support neuropeptide biosynthesis. Importantly, this review discusses important areas of current and future biomedical neuropeptide research with respect to biological regulation, inhibitors, structural features of proneuropeptide and protease interactions, and peptidomics combined with proteomics for systems biological approaches. Future studies that gain in-depth understanding of protease mechanisms for generating active neuropeptides will be instrumental for translational research to develop pharmacological strategies for regulation of neuropeptide functions. Pharmacological applications for neuropeptide research may provide valuable therapeutics in health and disease. PMID:18184105

Obscurin Targets Ankyrin-B and Protein Phosphatase 2A to the Cardiac M-line*

PubMed Central

Cunha, Shane R.; Mohler, Peter J.

2008-01-01

Ankyrin-B targets ion channels and transporters in excitable cells. Dysfunction in ankyrin-B-based pathways results in defects in cardiac physiology. Despite a wealth of knowledge regarding the role of ankyrin-B for cardiac function, little is known regarding the mechanisms underlying ankyrin-B regulation. Moreover, the pathways underlying ankyrin-B targeting in heart are unclear. We report that alternative splicing regulates ankyrin-B localization and function in cardiomyocytes. Specifically, we identify a novel exon (exon 43′) in the ankyrin-B regulatory domain that mediates interaction with the Rho-GEF obscurin. Ankyrin-B transcripts harboring exon 43′ represent the primary cardiac isoform in human and mouse. We demonstrate that ankyrin-B and obscurin are co-localized at the M-line of myocytes and co-immunoprecipitate from heart. We define the structural requirements for ankyrin-B/obscurin interaction to two motifs in the ankyrin-B regulatory domain and demonstrate that both are critical for obscurin/ankyrin-B interaction. In addition, we demonstrate that interaction with obscurin is required for ankyrin-B M-line targeting. Specifically, both obscurin-binding motifs are required for the M-line targeting of a GFP-ankyrin-B regulatory domain. Moreover, this construct acts as a dominant-negative by competing with endogenous ankyrin-B for obscurin-binding at the M-line, thus providing a powerful new tool to evaluate the function of obscurin/ankyrin-B interactions. With this new tool, we demonstrate that the obscurin/ankyrin-B interaction is critical for recruitment of PP2A to the cardiac M-line. Together, these data provide the first evidence for the molecular basis of ankyrin-B and PP2A targeting and function at the cardiac M-line. Finally, we report that ankyrin-B R1788W is localized adjacent to the ankyrin-B obscurin-binding motif and increases binding activity for obscurin. In summary, our new findings demonstrate that ANK2 is subject to alternative splicing that gives rise to unique polypeptides with diverse roles in cardiac function. PMID:18782775

Regulation of NAD+ metabolism, signaling and compartmentalization in the yeast Saccharomyces cerevisiae.

PubMed

Kato, Michiko; Lin, Su-Ju

2014-11-01

Pyridine nucleotides are essential coenzymes in many cellular redox reactions in all living systems. In addition to functioning as a redox carrier, NAD(+) is also a required co-substrate for the conserved sirtuin deacetylases. Sirtuins regulate transcription, genome maintenance and metabolism and function as molecular links between cells and their environment. Maintaining NAD(+) homeostasis is essential for proper cellular function and aberrant NAD(+) metabolism has been implicated in a number of metabolic- and age-associated diseases. Recently, NAD(+) metabolism has been linked to the phosphate-responsive signaling pathway (PHO pathway) in the budding yeast Saccharomyces cerevisiae. Activation of the PHO pathway is associated with the production and mobilization of the NAD(+) metabolite nicotinamide riboside (NR), which is mediated in part by PHO-regulated nucleotidases. Cross-regulation between NAD(+) metabolism and the PHO pathway has also been reported; however, detailed mechanisms remain to be elucidated. The PHO pathway also appears to modulate the activities of common downstream effectors of multiple nutrient-sensing pathways (Ras-PKA, TOR, Sch9/AKT). These signaling pathways were suggested to play a role in calorie restriction-mediated beneficial effects, which have also been linked to Sir2 function and NAD(+) metabolism. Here, we discuss the interactions of these pathways and their potential roles in regulating NAD(+) metabolism. In eukaryotic cells, intracellular compartmentalization facilitates the regulation of enzymatic functions and also concentrates or sequesters specific metabolites. Various NAD(+)-mediated cellular functions such as mitochondrial oxidative phosphorylation are compartmentalized. Therefore, we also discuss several key players functioning in mitochondrial, cytosolic and vacuolar compartmentalization of NAD(+) intermediates, and their potential roles in NAD(+) homeostasis. To date, it remains unclear how NAD(+) and NAD(+) intermediates shuttle between different cellular compartments. Together, these studies provide a molecular basis for how NAD(+) homeostasis factors and the interacting signaling pathways confer metabolic flexibility and contribute to maintaining cell fitness and genome stability. Copyright © 2014 Elsevier B.V. All rights reserved.

Regulation of NAD+ metabolism, signaling and compartmentalization in the yeast Saccharomyces cerevisiae

PubMed Central

Kato, Michiko; Lin, Su-Ju

2014-01-01

Pyridine nucleotides are essential coenzymes in many cellular redox reactions in all living systems. In addition to functioning as a redox carrier, NAD+ is also a required co-substrate for the conserved sirtuin deacetylases. Sirtuins regulate transcription, genome maintenance and metabolism and function as molecular links between cells and their environment. Maintaining NAD+ homeostasis is essential for proper cellular function and aberrant NAD+ metabolism has been implicated in a number of metabolic- and age-associated diseases. Recently, NAD+ metabolism has been linked to the phosphate-responsive signaling pathway (PHO pathway) in the budding yeast Saccharomyces cerevisiae. Activation of the PHO pathway is associated with the production and mobilization of the NAD+ metabolite nicotinamide riboside (NR), which is mediated in part by PHO-regulated nucleotidases. Cross-regulation between NAD+ metabolism and the PHO pathway has also been reported; however, detailed mechanisms remain to be elucidated. The PHO pathway also appears to modulate the activities of common downstream effectors of multiple nutrient-sensing pathways (Ras-PKA, TOR, Sch9/AKT). These signaling pathways were suggested to play a role in calorie restriction-mediated beneficial effects, which have also been linked to Sir2 function and NAD+ metabolism. Here, we discuss the interactions of these pathways and their potential roles in regulating NAD+ metabolism. In eukaryotic cells, intracellular compartmentalization facilitates the regulation of enzymatic functions and also concentrates or sequesters specific metabolites. Various NAD+-mediated cellular functions such as mitochondrial oxidative phosphorylation are compartmentalized. Therefore, we also discuss several key players functioning in mitochondrial, cytosolic and vacuolar compartmentalization of NAD+ intermediates, and their potential roles in NAD+ homeostasis. To date, it remains unclear how NAD+ and NAD+ intermediates shuttle between different cellular compartments. Together, these studies provide a molecular basis for how NAD+ homeostasis factors and the interacting signaling pathways confer metabolic flexibility and contribute to maintaining cell fitness and genome stability. PMID:25096760

Structural and Functional Analysis of a Novel Interaction Motif within UFM1-activating Enzyme 5 (UBA5) Required for Binding to Ubiquitin-like Proteins and Ufmylation*

PubMed Central

Habisov, Sabrina; Huber, Jessica; Ichimura, Yoshinobu; Akutsu, Masato; Rogova, Natalia; Loehr, Frank; McEwan, David G.; Johansen, Terje; Dikic, Ivan; Doetsch, Volker; Komatsu, Masaaki; Rogov, Vladimir V.; Kirkin, Vladimir

2016-01-01

The covalent conjugation of ubiquitin-fold modifier 1 (UFM1) to proteins generates a signal that regulates transcription, response to cell stress, and differentiation. Ufmylation is initiated by ubiquitin-like modifier activating enzyme 5 (UBA5), which activates and transfers UFM1 to ubiquitin-fold modifier-conjugating enzyme 1 (UFC1). The details of the interaction between UFM1 and UBA5 required for UFM1 activation and its downstream transfer are however unclear. In this study, we described and characterized a combined linear LC3-interacting region/UFM1-interacting motif (LIR/UFIM) within the C terminus of UBA5. This single motif ensures that UBA5 binds both UFM1 and light chain 3/γ-aminobutyric acid receptor-associated proteins (LC3/GABARAP), two ubiquitin (Ub)-like proteins. We demonstrated that LIR/UFIM is required for the full biological activity of UBA5 and for the effective transfer of UFM1 onto UFC1 and a downstream protein substrate both in vitro and in cells. Taken together, our study provides important structural and functional insights into the interaction between UBA5 and Ub-like modifiers, improving the understanding of the biology of the ufmylation pathway. PMID:26929408

Hormone treatment enhances WT1 activation of Renilla luciferase constructs in LNCaP cells.

PubMed

Hanson, Julie; Reese, Jennifer; Gorman, Jacquelyn; Cash, Jennifer; Fraizer, Gail

2007-01-01

The zinc finger transcription factor, WT1, regulates many growth control genes, repressing or activating transcription depending on the gene and cell type. Based on earlier analyses of the effect of WT1 on androgen responsive genes, we hypothesized that there may be an interaction between the androgen signaling pathway and WT1, such that the commonly used Renilla luciferase control vectors were activated in LNCaP prostate cancer cells. Using cotransfection assays we tested the effects of WT1 and/or the androgen analog, R1881, on two Renilla luciferase vectors, pRL-SV40 and the promoter-less pRL-null. To determine whether the zinc finger DNA binding domain was required, the zinc finger mutant DDS-WT1 (R394W) was tested; but it had no significant effect on the Renilla luciferase vectors. To determine whether the androgen signaling pathway was required, WT1 was co-transfected with Renilla vectors in cells with varied hormone responsiveness. The WT1 effect on pRL-null varied from no significant effect in 293 and PC3 cells to very strong enhancement in LNCaP cells treated with 5 nM R1881. Overall, these results suggest that hormone enhanced WT1 mediated activation of Renilla luciferase and that these interactions require an intact WT1 zinc finger DNA binding domain.

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

The PDZ protein tax-interacting protein-1 inhibits beta-catenin transcriptional activity and growth of colorectal cancer cells.

PubMed

Kanamori, Mutsumi; Sandy, Peter; Marzinotto, Stefania; Benetti, Roberta; Kai, Chikatoshi; Hayashizaki, Yoshihide; Schneider, Claudio; Suzuki, Harukazu

2003-10-03

Wnt signaling is essential during development while deregulation of this pathway frequently leads to the formation of various tumors including colorectal carcinomas. A key component of the pathway is beta-catenin that, in association with TCF-4, directly regulates the expression of Wnt-responsive genes. To identify novel binding partners of beta-catenin that may control its transcriptional activity, we performed a mammalian two-hybrid screen and isolated the Tax-interacting protein (TIP-1). The in vivo complex formation between beta-catenin and TIP-1 was verified by coimmunoprecipitation, and a direct physical association was revealed by glutathione S-transferase pull-down experiments in vitro. By using a panel of deletion mutants of both proteins, we demonstrate that the interaction is mediated by the PDZ (PSD-95/DLG/ZO-1 homology) domain of TIP-1 and requires primarily the last four amino acids of beta-catenin. TIP-1 overexpression resulted in a dose-dependent decrease in the transcriptional activity of beta-catenin when tested on the TOP/FOPFLASH reporter system. Conversely, siRNA-mediated knock-down of endogenous TIP-1 slightly increased endogenous beta-catenin transactivation function. Moreover, we show that overexpression of TIP-1 reduced the proliferation and anchorage-independent growth of colorectal cancer cells. These data suggest that TIP-1 may represent a novel regulatory element in the Wnt/beta-catenin signaling pathway.

Pathway redundancy and protein essentiality revealed in the Saccharomyces cerevisiae interaction networks

PubMed Central

Ulitsky, Igor; Shamir, Ron

2007-01-01

The biological interpretation of genetic interactions is a major challenge. Recently, Kelley and Ideker proposed a method to analyze together genetic and physical networks, which explains many of the known genetic interactions as linking different pathways in the physical network. Here, we extend this method and devise novel analytic tools for interpreting genetic interactions in a physical context. Applying these tools on a large-scale Saccharomyces cerevisiae data set, our analysis reveals 140 between-pathway models that explain 3765 genetic interactions, roughly doubling those that were previously explained. Model genes tend to have short mRNA half-lives and many phosphorylation sites, suggesting that their stringent regulation is linked to pathway redundancy. We also identify ‘pivot' proteins that have many physical interactions with both pathways in our models, and show that pivots tend to be essential and highly conserved. Our analysis of models and pivots sheds light on the organization of the cellular machinery as well as on the roles of individual proteins. PMID:17437029

Insight into bacterial virulence mechanisms against host immune response via the Yersinia pestis-human protein-protein interaction network.

PubMed

Yang, Huiying; Ke, Yuehua; Wang, Jian; Tan, Yafang; Myeni, Sebenzile K; Li, Dong; Shi, Qinghai; Yan, Yanfeng; Chen, Hui; Guo, Zhaobiao; Yuan, Yanzhi; Yang, Xiaoming; Yang, Ruifu; Du, Zongmin

2011-11-01

A Yersinia pestis-human protein interaction network is reported here to improve our understanding of its pathogenesis. Up to 204 interactions between 66 Y. pestis bait proteins and 109 human proteins were identified by yeast two-hybrid assay and then combined with 23 previously published interactions to construct a protein-protein interaction network. Topological analysis of the interaction network revealed that human proteins targeted by Y. pestis were significantly enriched in the proteins that are central in the human protein-protein interaction network. Analysis of this network showed that signaling pathways important for host immune responses were preferentially targeted by Y. pestis, including the pathways involved in focal adhesion, regulation of cytoskeleton, leukocyte transendoepithelial migration, and Toll-like receptor (TLR) and mitogen-activated protein kinase (MAPK) signaling. Cellular pathways targeted by Y. pestis are highly relevant to its pathogenesis. Interactions with host proteins involved in focal adhesion and cytoskeketon regulation pathways could account for resistance of Y. pestis to phagocytosis. Interference with TLR and MAPK signaling pathways by Y. pestis reflects common characteristics of pathogen-host interaction that bacterial pathogens have evolved to evade host innate immune response by interacting with proteins in those signaling pathways. Interestingly, a large portion of human proteins interacting with Y. pestis (16/109) also interacted with viral proteins (Epstein-Barr virus [EBV] and hepatitis C virus [HCV]), suggesting that viral and bacterial pathogens attack common cellular functions to facilitate infections. In addition, we identified vasodilator-stimulated phosphoprotein (VASP) as a novel interaction partner of YpkA and showed that YpkA could inhibit in vitro actin assembly mediated by VASP.

Crosstalk between Caveolin-1/Extracellular Signal-regulated Kinase (ERK) and β-Catenin Survival Pathways in Osteocyte Mechanotransduction*

PubMed Central

Gortazar, Arancha R.; Martin-Millan, Marta; Bravo, Beatriz; Plotkin, Lilian I.; Bellido, Teresita

2013-01-01

Osteocyte viability is a critical determinant of bone strength and is promoted by both mechanical stimulation and activation of the Wnt signaling pathway. Earlier studies demonstrated that both stimuli promote survival of osteocytes by activating the ERKs. Here, we show that there is interaction between the caveolin-1/ERK and Wnt/β-catenin signaling pathways in the transduction of mechanical cues into osteocyte survival. Thus, ERK nuclear translocation and anti-apoptosis induced by mechanical stimulation are abolished by the Wnt antagonist Dkk1 and the β-catenin degradation stimulator Axin2. Conversely, GSK3β phosphorylation and β-catenin accumulation induced by mechanical stimulation are abolished by either pharmacologic inhibition of ERKs or silencing caveolin-1. In contrast, the canonical Wnt signaling inhibitor dominant-negative T cell factor does not alter ERK nuclear translocation or survival induced by mechanical stimulation. These findings demonstrate that β-catenin accumulation is an essential component of the mechanotransduction machinery in osteocytes, albeit β-catenin/T cell factor-mediated transcription is not required. The simultaneous requirement of β-catenin for ERK activation and of ERK activation for β-catenin accumulation suggests a bidirectional crosstalk between the caveolin-1/ERK and Wnt/β-catenin pathways in mechanotransduction leading to osteocyte survival. PMID:23362257

Regulation and function of DNA methylation in plants and animals

PubMed Central

He, Xin-Jian; Chen, Taiping; Zhu, Jian-Kang

2011-01-01

DNA methylation is an important epigenetic mark involved in diverse biological processes. In plants, DNA methylation can be established through the RNA-directed DNA methylation pathway, an RNA interference pathway for transcriptional gene silencing (TGS), which requires 24-nt small interfering RNAs. In mammals, de novo DNA methylation occurs primarily at two developmental stages: during early embryogenesis and during gametogenesis. While it is not clear whether establishment of DNA methylation patterns in mammals involves RNA interference in general, de novo DNA methylation and suppression of transposons in germ cells require 24-32-nt piwi-interacting small RNAs. DNA methylation status is dynamically regulated by DNA methylation and demethylation reactions. In plants, active DNA demethylation relies on the repressor of silencing 1 family of bifunctional DNA glycosylases, which remove the 5-methylcytosine base and then cleave the DNA backbone at the abasic site, initiating a base excision repair (BER) pathway. In animals, multiple mechanisms of active DNA demethylation have been proposed, including a deaminase- and DNA glycosylase-initiated BER pathway. New information concerning the effects of various histone modifications on the establishment and maintenance of DNA methylation has broadened our understanding of the regulation of DNA methylation. The function of DNA methylation in plants and animals is also discussed in this review. PMID:21321601

Platelets regulate lymphatic vascular development through CLEC-2-SLP-76 signaling.

PubMed

Bertozzi, Cara C; Schmaier, Alec A; Mericko, Patricia; Hess, Paul R; Zou, Zhiying; Chen, Mei; Chen, Chiu-Yu; Xu, Bin; Lu, Min-min; Zhou, Diane; Sebzda, Eric; Santore, Matthew T; Merianos, Demetri J; Stadtfeld, Matthias; Flake, Alan W; Graf, Thomas; Skoda, Radek; Maltzman, Jonathan S; Koretzky, Gary A; Kahn, Mark L

2010-07-29

Although platelets appear by embryonic day 10.5 in the developing mouse, an embryonic role for these cells has not been identified. The SYK-SLP-76 signaling pathway is required in blood cells to regulate embryonic blood-lymphatic vascular separation, but the cell type and molecular mechanism underlying this regulatory pathway are not known. In the present study we demonstrate that platelets regulate lymphatic vascular development by directly interacting with lymphatic endothelial cells through C-type lectin-like receptor 2 (CLEC-2) receptors. PODOPLANIN (PDPN), a transmembrane protein expressed on the surface of lymphatic endothelial cells, is required in nonhematopoietic cells for blood-lymphatic separation. Genetic loss of the PDPN receptor CLEC-2 ablates PDPN binding by platelets and confers embryonic lymphatic vascular defects like those seen in animals lacking PDPN or SLP-76. Platelet factor 4-Cre-mediated deletion of Slp-76 is sufficient to confer lymphatic vascular defects, identifying platelets as the cell type in which SLP-76 signaling is required to regulate lymphatic vascular development. Consistent with these genetic findings, we observe SLP-76-dependent platelet aggregate formation on the surface of lymphatic endothelial cells in vivo and ex vivo. These studies identify a nonhemostatic pathway in which platelet CLEC-2 receptors bind lymphatic endothelial PDPN and activate SLP-76 signaling to regulate embryonic vascular development.

Pre-irradiation testing and analysis to support the LWRS Hybrid SiC-CMC-Zircaloy-04 unfueled rodlet irradiation

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

Isabella J van Rooyen

2012-09-01

Nuclear fuel performance is a significant driver of nuclear power plant operational performance, safety, economics and waste disposal requirements. The Advanced Light Water Reactor (LWR) Nuclear Fuel Development Pathway focuses on improving the scientific knowledge basis to enable the development of high-performance, high burn-up fuels with improved safety and cladding integrity and improved nuclear fuel cycle economics. To achieve significant improvements, fundamental changes are required in the areas of nuclear fuel composition, cladding integrity, and fuel/cladding interaction.

Pre-irradiation testing and analysis to support the LWRS Hybrid SiC-CMC-Zircaloy-04 unfueled rodlet irradiation

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

Isabella J van Rooyen

2013-01-01

Nuclear fuel performance is a significant driver of nuclear power plant operational performance, safety, economics and waste disposal requirements. The Advanced Light Water Reactor (LWR) Nuclear Fuel Development Pathway focuses on improving the scientific knowledge basis to enable the development of high-performance, high burn-up fuels with improved safety and cladding integrity and improved nuclear fuel cycle economics. To achieve significant improvements, fundamental changes are required in the areas of nuclear fuel composition, cladding integrity, and fuel/cladding interaction.

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.

In silico study of protein to protein interaction analysis of AMP-activated protein kinase and mitochondrial activity in three different farm animal species

NASA Astrophysics Data System (ADS)

Prastowo, S.; Widyas, N.

2018-03-01

AMP-activated protein kinase (AMPK) is cellular energy censor which works based on ATP and AMP concentration. This protein interacts with mitochondria in determine its activity to generate energy for cell metabolism purposes. For that, this paper aims to compare the protein to protein interaction of AMPK and mitochondrial activity genes in the metabolism of known animal farm (domesticated) that are cattle (Bos taurus), pig (Sus scrofa) and chicken (Gallus gallus). In silico study was done using STRING V.10 as prominent protein interaction database, followed with biological function comparison in KEGG PATHWAY database. Set of genes (12 in total) were used as input analysis that are PRKAA1, PRKAA2, PRKAB1, PRKAB2, PRKAG1, PRKAG2, PRKAG3, PPARGC1, ACC, CPT1B, NRF2 and SOD. The first 7 genes belong to gene in AMPK family, while the last 5 belong to mitochondrial activity genes. The protein interaction result shows 11, 8 and 5 metabolism pathways in Bos taurus, Sus scrofa and Gallus gallus, respectively. The top pathway in Bos taurus is AMPK signaling pathway (10 genes), Sus scrofa is Adipocytokine signaling pathway (8 genes) and Gallus gallus is FoxO signaling pathway (5 genes). Moreover, the common pathways found in those 3 species are Adipocytokine signaling pathway, Insulin signaling pathway and FoxO signaling pathway. Genes clustered in Adipocytokine and Insulin signaling pathway are PRKAA2, PPARGC1A, PRKAB1 and PRKAG2. While, in FoxO signaling pathway are PRKAA2, PRKAB1, PRKAG2. According to that, we found PRKAA2, PRKAB1 and PRKAG2 are the common genes. Based on the bioinformatics analysis, we can demonstrate that protein to protein interaction shows distinct different of metabolism in different species. However, further validation is needed to give a clear explanation.

Mapping biological process relationships and disease perturbations within a pathway network.

PubMed

Stoney, Ruth; Robertson, David L; Nenadic, Goran; Schwartz, Jean-Marc

2018-01-01

Molecular interaction networks are routinely used to map the organization of cellular function. Edges represent interactions between genes, proteins, or metabolites. However, in living cells, molecular interactions are dynamic, necessitating context-dependent models. Contextual information can be integrated into molecular interaction networks through the inclusion of additional molecular data, but there are concerns about completeness and relevance of this data. We developed an approach for representing the organization of human cellular processes using pathways as the nodes in a network. Pathways represent spatial and temporal sets of context-dependent interactions, generating a high-level network when linked together, which incorporates contextual information without the need for molecular interaction data. Analysis of the pathway network revealed linked communities representing functional relationships, comparable to those found in molecular networks, including metabolism, signaling, immunity, and the cell cycle. We mapped a range of diseases onto this network and find that pathways associated with diseases tend to be functionally connected, highlighting the perturbed functions that result in disease phenotypes. We demonstrated that disease pathways cluster within the network. We then examined the distribution of cancer pathways and showed that cancer pathways tend to localize within the signaling, DNA processes and immune modules, although some cancer-associated nodes are found in other network regions. Altogether, we generated a high-confidence functional network, which avoids some of the shortcomings faced by conventional molecular models. Our representation provides an intuitive functional interpretation of cellular organization, which relies only on high-quality pathway and Gene Ontology data. The network is available at https://data.mendeley.com/datasets/3pbwkxjxg9/1.

Interactions between chemical and climate stressors: A role for mechanistic toxicology in assessing climate change risks

USGS Publications Warehouse

Hooper, Michael J.; Ankley, Gerald T.; Cristol, Daniel A.; Maryoung, Lindley A.; Noyes, Pamela D.; Pinkerton, Kent E.

2013-01-01

Incorporation of global climate change (GCC) effects into assessments of chemical risk and injury requires integrated examinations of chemical and nonchemical stressors. Environmental variables altered by GCC (temperature, precipitation, salinity, pH) can influence the toxicokinetics of chemical absorption, distribution, metabolism, and excretion as well as toxicodynamic interactions between chemicals and target molecules. In addition, GCC challenges processes critical for coping with the external environment (water balance, thermoregulation, nutrition, and the immune, endocrine, and neurological systems), leaving organisms sensitive to even slight perturbations by chemicals when pushed to the limits of their physiological tolerance range. In simplest terms, GCC can make organisms more sensitive to chemical stressors, while alternatively, exposure to chemicals can make organisms more sensitive to GCC stressors. One challenge is to identify potential interactions between nonchemical and chemical stressors affecting key physiological processes in an organism. We employed adverse outcome pathways, constructs depicting linkages between mechanism-based molecular initiating events and impacts on individuals or populations, to assess how chemical- and climate-specific variables interact to lead to adverse outcomes. Case examples are presented for prospective scenarios, hypothesizing potential chemical–GCC interactions, and retrospective scenarios, proposing mechanisms for demonstrated chemical–climate interactions in natural populations. Understanding GCC interactions along adverse outcome pathways facilitates extrapolation between species or other levels of organization, development of hypotheses and focal areas for further research, and improved inputs for risk and resource injury assessments.

Drosophila Neurexin IV Interacts with Roundabout and is Required for Repulsive Midline Axon Guidance

PubMed Central

Banerjee, Swati; Blauth, Kevin; Peters, Kimberly; Rogers, Stephen L.; Fanning, Alan S.; Bhat, Manzoor A.

2010-01-01

Slit/Roundabout (Robo) signaling controls midline repulsive axon guidance. However, proteins that interact with Slit/Robo at the cell surface remain largely uncharacterized. Here, we report that the Drosophila transmembrane septate junction-specific protein, Neurexin IV (Nrx IV), functions in midline repulsive axon guidance. Nrx IV is expressed in the neurons of the developing ventral nerve cord and nrx IV mutants show crossing and circling of ipsilateral axons and fused commissures. Interestingly, the axon guidance defects observed in nrx IV mutants seem independent of its other binding partners such as Contactin and Neuroglian and the midline glia protein Wrapper that interacts in trans with Nrx IV. nrx IV mutants show diffuse Robo localization and dose-dependent genetic interactions between nrx IV/robo and nrx IV/slit indicate that they function in a common pathway. In vivo biochemical studies reveal that Nrx IV associates with Robo, Slit and Syndecan, and interactions between Robo and Slit, or Nrx IV and Slit, are affected in nrx IV and robo mutants, respectively. Coexpression of Nrx IV and Robo in mammalian cells confirms that these proteins retain the ability to interact in a heterologous system. Furthermore, we demonstrate that the extracellular region of Nrx IV is sufficient to rescue Robo localization and axon guidance phenotypes in nrx IV mutants. Together our studies establish that Nrx IV is essential for proper Robo localization, and identify Nrx IV as a novel interacting partner of the Slit/Robo signaling pathway. PMID:20410118

Drosophila neurexin IV interacts with Roundabout and is required for repulsive midline axon guidance.

PubMed

Banerjee, Swati; Blauth, Kevin; Peters, Kimberly; Rogers, Stephen L; Fanning, Alan S; Bhat, Manzoor A

2010-04-21

Slit/Roundabout (Robo) signaling controls midline repulsive axon guidance. However, proteins that interact with Slit/Robo at the cell surface remain largely uncharacterized. Here, we report that the Drosophila transmembrane septate junction-specific protein Neurexin IV (Nrx IV) functions in midline repulsive axon guidance. Nrx IV is expressed in the neurons of the developing ventral nerve cord, and nrx IV mutants show crossing and circling of ipsilateral axons and fused commissures. Interestingly, the axon guidance defects observed in nrx IV mutants seem independent of its other binding partners, such as Contactin and Neuroglian and the midline glia protein Wrapper, which interacts in trans with Nrx IV. nrx IV mutants show diffuse Robo localization, and dose-dependent genetic interactions between nrx IV/robo and nrx IV/slit indicate that they function in a common pathway. In vivo biochemical studies reveal that Nrx IV associates with Robo, Slit, and Syndecan, and interactions between Robo and Slit, or Nrx IV and Slit, are affected in nrx IV and robo mutants, respectively. Coexpression of Nrx IV and Robo in mammalian cells confirms that these proteins retain the ability to interact in a heterologous system. Furthermore, we demonstrate that the extracellular region of Nrx IV is sufficient to rescue Robo localization and axon guidance phenotypes in nrx IV mutants. Together, our studies establish that Nrx IV is essential for proper Robo localization and identify Nrx IV as a novel interacting partner of the Slit/Robo signaling pathway.

INTERACTIONS BETWEEN CHEMICAL AND CLIMATE STRESSORS: A ROLE FOR MECHANISTIC TOXICOLOGY IN ASSESSING CLIMATE CHANGE RISKS

PubMed Central

Hooper, Michael J; Ankley, Gerald T; Cristol, Daniel A; Maryoung, Lindley A; Noyes, Pamela D; Pinkerton, Kent E

2013-01-01

Incorporation of global climate change (GCC) effects into assessments of chemical risk and injury requires integrated examinations of chemical and nonchemical stressors. Environmental variables altered by GCC (temperature, precipitation, salinity, pH) can influence the toxicokinetics of chemical absorption, distribution, metabolism, and excretion as well as toxicodynamic interactions between chemicals and target molecules. In addition, GCC challenges processes critical for coping with the external environment (water balance, thermoregulation, nutrition, and the immune, endocrine, and neurological systems), leaving organisms sensitive to even slight perturbations by chemicals when pushed to the limits of their physiological tolerance range. In simplest terms, GCC can make organisms more sensitive to chemical stressors, while alternatively, exposure to chemicals can make organisms more sensitive to GCC stressors. One challenge is to identify potential interactions between nonchemical and chemical stressors affecting key physiological processes in an organism. We employed adverse outcome pathways, constructs depicting linkages between mechanism-based molecular initiating events and impacts on individuals or populations, to assess how chemical- and climate-specific variables interact to lead to adverse outcomes. Case examples are presented for prospective scenarios, hypothesizing potential chemical–GCC interactions, and retrospective scenarios, proposing mechanisms for demonstrated chemical–climate interactions in natural populations. Understanding GCC interactions along adverse outcome pathways facilitates extrapolation between species or other levels of organization, development of hypotheses and focal areas for further research, and improved inputs for risk and resource injury assessments. Environ. Toxicol. Chem. 2013;32:32–48. © 2012 SETAC PMID:23136056

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

Multiple functions of BCL-2 family proteins.

PubMed

Hardwick, J Marie; Soane, Lucian

2013-02-01

BCL-2 family proteins are the regulators of apoptosis, but also have other functions. This family of interacting partners includes inhibitors and inducers of cell death. Together they regulate and mediate the process by which mitochondria contribute to cell death known as the intrinsic apoptosis pathway. This pathway is required for normal embryonic development and for preventing cancer. However, before apoptosis is induced, BCL-2 proteins have critical roles in normal cell physiology related to neuronal activity, autophagy, calcium handling, mitochondrial dynamics and energetics, and other processes of normal healthy cells. The relative importance of these physiological functions compared to their apoptosis functions in overall organismal physiology is difficult to decipher. Apoptotic and noncanonical functions of these proteins may be intertwined to link cell growth to cell death. Disentanglement of these functions may require delineation of biochemical activities inherent to the characteristic three-dimensional shape shared by distantly related viral and cellular BCL-2 family members.

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

ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling.

PubMed

Harris, Abigail; Siggers, Pam; Corrochano, Silvia; Warr, Nick; Sagar, Danielle; Grimes, Daniel T; Suzuki, Makoto; Burdine, Rebecca D; Cong, Feng; Koo, Bon-Kyoung; Clevers, Hans; Stévant, Isabelle; Nef, Serge; Wells, Sara; Brauner, Raja; Ben Rhouma, Bochra; Belguith, Neïla; Eozenou, Caroline; Bignon-Topalovic, Joelle; Bashamboo, Anu; McElreavey, Ken; Greenfield, Andy

2018-05-22

Mammalian sex determination is controlled by the antagonistic interactions of two genetic pathways: The SRY-SOX9-FGF9 network promotes testis determination partly by opposing proovarian pathways, while RSPO1/WNT-β-catenin/FOXL2 signals control ovary development by inhibiting SRY-SOX9-FGF9. The molecular basis of this mutual antagonism is unclear. Here we show that ZNRF3, a WNT signaling antagonist and direct target of RSPO1-mediated inhibition, is required for sex determination in mice. XY mice lacking ZNRF3 exhibit complete or partial gonadal sex reversal, or related defects. These abnormalities are associated with ectopic WNT/β-catenin activity and reduced Sox9 expression during fetal sex determination. Using exome sequencing of individuals with 46,XY disorders of sex development, we identified three human ZNRF3 variants in very rare cases of XY female presentation. We tested two missense variants and show that these disrupt ZNRF3 activity in both human cell lines and zebrafish embryo assays. Our data identify a testis-determining function for ZNRF3 and indicate a mechanism of direct molecular interaction between two mutually antagonistic organogenetic pathways. Copyright © 2018 the Author(s). Published by PNAS.

Autolysosomal β-catenin degradation regulates Wnt-autophagy-p62 crosstalk

PubMed Central

Petherick, Katy J; Williams, Ann C; Lane, Jon D; Ordóñez-Morán, Paloma; Huelsken, Joerg; Collard, Tracey J; Smartt, Helena JM; Batson, Jennifer; Malik, Karim; Paraskeva, Chris; Greenhough, Alexander

2013-01-01

The Wnt/β-catenin signalling and autophagy pathways each play important roles during development, adult tissue homeostasis and tumorigenesis. Here we identify the Wnt/β-catenin signalling pathway as a negative regulator of both basal and stress-induced autophagy. Manipulation of β-catenin expression levels in vitro and in vivo revealed that β-catenin suppresses autophagosome formation and directly represses p62/SQSTM1 (encoding the autophagy adaptor p62) via TCF4. Furthermore, we show that during nutrient deprivation β-catenin is selectively degraded via the formation of a β-catenin–LC3 complex, attenuating β-catenin/TCF-driven transcription and proliferation to favour adaptation during metabolic stress. Formation of the β-catenin–LC3 complex is mediated by a W/YXXI/L motif and LC3-interacting region (LIR) in β-catenin, which is required for interaction with LC3 and non-proteasomal degradation of β-catenin. Thus, Wnt/β-catenin represses autophagy and p62 expression, while β-catenin is itself targeted for autophagic clearance in autolysosomes upon autophagy induction. These findings reveal a regulatory feedback mechanism that place β-catenin at a key cellular integration point coordinating proliferation with autophagy, with implications for targeting these pathways for cancer therapy. PMID:23736261

ZNRF3 functions in mammalian sex determination by inhibiting canonical WNT signaling

PubMed Central

Harris, Abigail; Siggers, Pam; Warr, Nick; Sagar, Danielle; Grimes, Daniel T.; Cong, Feng; Koo, Bon-Kyoung; Clevers, Hans; Stévant, Isabelle; Nef, Serge; Wells, Sara; Brauner, Raja; Ben Rhouma, Bochra; Belguith, Neïla; Eozenou, Caroline; Bignon-Topalovic, Joelle; Bashamboo, Anu; McElreavey, Ken

2018-01-01

Mammalian sex determination is controlled by the antagonistic interactions of two genetic pathways: The SRY-SOX9-FGF9 network promotes testis determination partly by opposing proovarian pathways, while RSPO1/WNT-β-catenin/FOXL2 signals control ovary development by inhibiting SRY-SOX9-FGF9. The molecular basis of this mutual antagonism is unclear. Here we show that ZNRF3, a WNT signaling antagonist and direct target of RSPO1-mediated inhibition, is required for sex determination in mice. XY mice lacking ZNRF3 exhibit complete or partial gonadal sex reversal, or related defects. These abnormalities are associated with ectopic WNT/β-catenin activity and reduced Sox9 expression during fetal sex determination. Using exome sequencing of individuals with 46,XY disorders of sex development, we identified three human ZNRF3 variants in very rare cases of XY female presentation. We tested two missense variants and show that these disrupt ZNRF3 activity in both human cell lines and zebrafish embryo assays. Our data identify a testis-determining function for ZNRF3 and indicate a mechanism of direct molecular interaction between two mutually antagonistic organogenetic pathways. PMID:29735715

The identification and characterisation of a functional interaction between arginyl-tRNA-protein transferase and topoisomerase II.

PubMed

Barker, Catherine R; Mouchel, Nathalie A P; Jenkins, John R

2006-04-07

Topoisomerase II is required for the viability of all eukaryotic cells. It plays important roles in DNA replication, recombination, chromosome segregation, and the maintenance of the nuclear scaffold. Proteins that interact with and regulate this essential enzyme are of great interest. To investigate the role of proteins interacting with the N-terminal domain of the Saccharomyces cerevisiae topoisomerase II, we used a yeast two-hybrid protein interaction screen. We identified an interaction between arginyl-tRNA-protein transferase (Ate1) and the N-terminal domain of the S. cerevisiae topoisomerase II, including the potential site of interaction. Ate1 is a component of the N-end rule protein degradation pathway which targets proteins for degradation. We also propose a previously unidentified role for Ate1 in modulating the level of topoisomerase II through the cell cycle.

Groups: knowledge spreadsheets for symbolic biocomputing.

PubMed

Travers, Michael; Paley, Suzanne M; Shrager, Jeff; Holland, Timothy A; Karp, Peter D

2013-01-01

Knowledge spreadsheets (KSs) are a visual tool for interactive data analysis and exploration. They differ from traditional spreadsheets in that rather than being oriented toward numeric data, they work with symbolic knowledge representation structures and provide operations that take into account the semantics of the application domain. 'Groups' is an implementation of KSs within the Pathway Tools system. Groups allows Pathway Tools users to define a group of objects (e.g. groups of genes or metabolites) from a Pathway/Genome Database. Groups can be transformed (e.g. by transforming a metabolite group to the group of pathways in which those metabolites are substrates); combined through set operations; analysed (e.g. through enrichment analysis); and visualized (e.g. by painting onto a metabolic map diagram). Users of the Pathway Tools-based BioCyc.org website have made extensive use of Groups, and an informal survey of Groups users suggests that Groups has achieved the goal of allowing biologists themselves to perform some data manipulations that previously would have required the assistance of a programmer. Database URL: BioCyc.org.

A role for NRAGE in NF-κB activation through the non-canonical BMP pathway

PubMed Central

2010-01-01

Background Previous studies have linked neurotrophin receptor-interacting MAGE protein to the bone morphogenic protein signaling pathway and its effect on p38 mediated apoptosis of neural progenitor cells via the XIAP-Tak1-Tab1 complex. Its effect on NF-κB has yet to be explored. Results Herein we report that NRAGE, via the same XIAP-Tak1-Tab1 complex, is required for the phosphorylation of IKK -α/β and subsequent transcriptional activation of the p65 subunit of NF-κB. Ablation of endogenous NRAGE by siRNA inhibited NF-κB pathway activation, while ablation of Tak1 and Tab1 by morpholino inhibited overexpression of NRAGE from activating NF-κB. Finally, cytokine profiling of an NRAGE over-expressing stable line revealed the expression of macrophage migration inhibitory factor. Conclusion Modulation of NRAGE expression revealed novel roles in regulating NF-κB activity in the non-canonical bone morphogenic protein signaling pathway. The expression of macrophage migration inhibitory factor by bone morphogenic protein -4 reveals novel crosstalk between an immune cytokine and a developmental pathway. PMID:20100315

Inferring gene and protein interactions using PubMed citations and consensus Bayesian networks.

PubMed

Deeter, Anthony; Dalman, Mark; Haddad, Joseph; Duan, Zhong-Hui

2017-01-01

The PubMed database offers an extensive set of publication data that can be useful, yet inherently complex to use without automated computational techniques. Data repositories such as the Genomic Data Commons (GDC) and the Gene Expression Omnibus (GEO) offer experimental data storage and retrieval as well as curated gene expression profiles. Genetic interaction databases, including Reactome and Ingenuity Pathway Analysis, offer pathway and experiment data analysis using data curated from these publications and data repositories. We have created a method to generate and analyze consensus networks, inferring potential gene interactions, using large numbers of Bayesian networks generated by data mining publications in the PubMed database. Through the concept of network resolution, these consensus networks can be tailored to represent possible genetic interactions. We designed a set of experiments to confirm that our method is stable across variation in both sample and topological input sizes. Using gene product interactions from the KEGG pathway database and data mining PubMed publication abstracts, we verify that regardless of the network resolution or the inferred consensus network, our method is capable of inferring meaningful gene interactions through consensus Bayesian network generation with multiple, randomized topological orderings. Our method can not only confirm the existence of currently accepted interactions, but has the potential to hypothesize new ones as well. We show our method confirms the existence of known gene interactions such as JAK-STAT-PI3K-AKT-mTOR, infers novel gene interactions such as RAS- Bcl-2 and RAS-AKT, and found significant pathway-pathway interactions between the JAK-STAT signaling and Cardiac Muscle Contraction KEGG pathways.

Noncovalent interaction-assisted drug delivery system with highly efficient uptake and release of paclitaxel for anticancer therapy.

PubMed

Wei, Yuping; Ma, Liang; Zhang, Liang; Xu, Xia

2017-01-01

An effective drug delivery system requires efficient drug uptake and release inside cancer cells. Here, we report a novel drug delivery system, in which paclitaxel (PTX) interacts with a novel cell penetrating peptide (CPP) through noncovalent interaction designed based on molecular simulations. This CPP/PTX complex confers high efficiency in delivering PTX into cancer cells not by endocytosis but by an energy-independent pathway. Once inside cells, the noncovalent interaction between PTX and the CPP may allow fast release of PTX within cells due to the direct translocation of CPP/PTX. This drug delivery system exhibits strong capacity for inhibition of tumor growth and offers a new avenue for the development of advanced drug delivery systems for anticancer therapy.

Activated STAT5 proteins induce activation of the PI 3-kinase/Akt and Ras/MAPK pathways via the Gab2 scaffolding adapter.

PubMed

Nyga, Rémy; Pecquet, Christian; Harir, Noria; Gu, Haihua; Dhennin-Duthille, Isabelle; Régnier, Aline; Gouilleux-Gruart, Valérie; Lassoued, Kaïss; Gouilleux, Fabrice

2005-08-15

The active forms of STAT5A (signal transducer and activator of transcription 5A) and STAT5B are able to relieve the cytokine dependence of haematopoietic cells and to induce leukaemia in mice. We have demonstrated previously that activation of the PI3K (phosphoinositide 3-kinase) signalling cascade plays a major role in cell growth and survival induced by these proteins. Interaction between STAT5 and p85, the regulatory subunit of the PI3K, has been suggested to be required for this activation. We show in the present study that the scaffolding protein Gab2 [Grb2 (growth-factor-receptor-bound protein 2)-associated binder-2] is an essential component of this interaction. Gab2 is persistently tyrosine-phosphorylated in Ba/F3 cells expressing caSTAT5 (constitutively activated STAT5), independent of JAK2 (Janus kinase 2) activation where it interacts with STAT5, p85 and Grb2, but not with Shp2 [SH2 (Src homology 2)-domain-containing tyrosine phosphatase] proteins. Interaction of STAT5 with Gab2 was also observed in Ba/F3 cells stimulated with interleukin-3 or expressing the oncogenic fusion protein Tel-JAK2. The MAPKs (mitogen-activated protein kinases) ERK1 (extracellular-signal-regulated kinase 1) and ERK2 were constitutively activated in the caSTAT5-expressing cells and were found to be required for caSTAT5-induced cell proliferation. Overexpression of Gab2-3YF, a mutant of Gab2 incapable of binding PI3K, inhibited the proliferation and survival of caSTAT5-expressing cells as well as ERK1/2 and Akt/protein kinase B phosphorylation. Taken together, our results indicate that Gab2 is required for caSTAT5-induced cell proliferation by regulating both the PI3K/Akt and the Ras/MAPK pathways.

Activated STAT5 proteins induce activation of the PI 3-kinase/Akt and Ras/MAPK pathways via the Gab2 scaffolding adapter

PubMed Central

2005-01-01

The active forms of STAT5A (signal transducer and activator of transcription 5A) and STAT5B are able to relieve the cytokine dependence of haematopoietic cells and to induce leukaemia in mice. We have demonstrated previously that activation of the PI3K (phosphoinositide 3-kinase) signalling cascade plays a major role in cell growth and survival induced by these proteins. Interaction between STAT5 and p85, the regulatory subunit of the PI3K, has been suggested to be required for this activation. We show in the present study that the scaffolding protein Gab2 [Grb2 (growth-factor-receptor-bound protein 2)-associated binder-2] is an essential component of this interaction. Gab2 is persistently tyrosine-phosphorylated in Ba/F3 cells expressing caSTAT5 (constitutively activated STAT5), independent of JAK2 (Janus kinase 2) activation where it interacts with STAT5, p85 and Grb2, but not with Shp2 [SH2 (Src homology 2)-domain-containing tyrosine phosphatase] proteins. Interaction of STAT5 with Gab2 was also observed in Ba/F3 cells stimulated with interleukin-3 or expressing the oncogenic fusion protein Tel–JAK2. The MAPKs (mitogen-activated protein kinases) ERK1 (extracellular-signal-regulated kinase 1) and ERK2 were constitutively activated in the caSTAT5-expressing cells and were found to be required for caSTAT5-induced cell proliferation. Overexpression of Gab2-3YF, a mutant of Gab2 incapable of binding PI3K, inhibited the proliferation and survival of caSTAT5-expressing cells as well as ERK1/2 and Akt/protein kinase B phosphorylation. Taken together, our results indicate that Gab2 is required for caSTAT5-induced cell proliferation by regulating both the PI3K/Akt and the Ras/MAPK pathways. PMID:15833084

Thalamocortical and corticothalamic pathways differentially contribute to goal-directed behaviors in the rat

PubMed Central

Alcaraz, Fabien; Fresno, Virginie; Marchand, Alain R; Kremer, Eric J; Coutureau, Etienne

2018-01-01

Highly distributed neural circuits are thought to support adaptive decision-making in volatile and complex environments. Notably, the functional interactions between prefrontal and reciprocally connected thalamic nuclei areas may be important when choices are guided by current goal value or action-outcome contingency. We examined the functional involvement of selected thalamocortical and corticothalamic pathways connecting the dorsomedial prefrontal cortex (dmPFC) and the mediodorsal thalamus (MD) in the behaving rat. Using a chemogenetic approach to inhibit projection-defined dmPFC and MD neurons during an instrumental learning task, we show that thalamocortical and corticothalamic pathways differentially support goal attributes. Both pathways participate in adaptation to the current goal value, but only thalamocortical neurons are required to integrate current causal relationships. These data indicate that antiparallel flow of information within thalamocortical circuits may convey qualitatively distinct aspects of adaptive decision-making and highlight the importance of the direction of information flow within neural circuits. PMID:29405119

mTORC1 and p53

PubMed Central

Hasty, Paul; Sharp, Zelton Dave; Curiel, Tyler J.; Campisi, Judith

2013-01-01

A balance must be struck between cell growth and stress responses to ensure that cells proliferate without accumulating damaged DNA. This balance means that optimal cell proliferation requires the integration of pro-growth and stress-response pathways. mTOR (mechanistic target of rapamycin) is a pleiotropic kinase found in complex 1 (mTORC1). The mTORC1 pathway governs a response to mitogenic signals with high energy levels to promote protein synthesis and cell growth. In contrast, the p53 DNA damage response pathway is the arbiter of cell proliferation, restraining mTORC1 under conditions of genotoxic stress. Recent studies suggest a complicated integration of these pathways to ensure successful cell growth and proliferation without compromising genome maintenance. Deciphering this integration could be key to understanding the potential clinical usefulness of mTORC1 inhibitors like rapamycin. Here we discuss how these p53-mTORC1 interactions might play a role in the suppression of cancer and perhaps the development of cellular senescence and organismal aging. PMID:23255104

β-Catenin destruction complex-independent regulation of Hippo–YAP signaling by APC in intestinal tumorigenesis

PubMed Central

Cai, Jing; Maitra, Anirban; Anders, Robert A.; Taketo, Makoto M.; Pan, Duojia

2015-01-01

Mutations in Adenomatous polyposis coli (APC) underlie familial adenomatous polyposis (FAP), an inherited cancer syndrome characterized by the widespread development of colorectal polyps. APC is best known as a scaffold protein in the β-catenin destruction complex, whose activity is antagonized by canonical Wnt signaling. Whether other effector pathways mediate APC's tumor suppressor function is less clear. Here we report that activation of YAP, the downstream effector of the Hippo signaling pathway, is a general hallmark of tubular adenomas from FAP patients. We show that APC functions as a scaffold protein that facilitates the Hippo kinase cascade by interacting with Sav1 and Lats1. Consistent with the molecular link between APC and the Hippo signaling pathway, genetic analysis reveals that YAP is absolutely required for the development of APC-deficient adenomas. These findings establish Hippo–YAP signaling as a critical effector pathway downstream from APC, independent from its involvement in the β-catenin destruction complex. PMID:26193883

SpirPro: A Spirulina proteome database and web-based tools for the analysis of protein-protein interactions at the metabolic level in Spirulina (Arthrospira) platensis C1.

PubMed

Senachak, Jittisak; Cheevadhanarak, Supapon; Hongsthong, Apiradee

2015-07-29

Spirulina (Arthrospira) platensis is the only cyanobacterium that in addition to being studied at the molecular level and subjected to gene manipulation, can also be mass cultivated in outdoor ponds for commercial use as a food supplement. Thus, encountering environmental changes, including temperature stresses, is common during the mass production of Spirulina. The use of cyanobacteria as an experimental platform, especially for photosynthetic gene manipulation in plants and bacteria, is becoming increasingly important. Understanding the mechanisms and protein-protein interaction networks that underlie low- and high-temperature responses is relevant to Spirulina mass production. To accomplish this goal, high-throughput techniques such as OMICs analyses are used. Thus, large datasets must be collected, managed and subjected to information extraction. Therefore, databases including (i) proteomic analysis and protein-protein interaction (PPI) data and (ii) domain/motif visualization tools are required for potential use in temperature response models for plant chloroplasts and photosynthetic bacteria. A web-based repository was developed including an embedded database, SpirPro, and tools for network visualization. Proteome data were analyzed integrated with protein-protein interactions and/or metabolic pathways from KEGG. The repository provides various information, ranging from raw data (2D-gel images) to associated results, such as data from interaction and/or pathway analyses. This integration allows in silico analyses of protein-protein interactions affected at the metabolic level and, particularly, analyses of interactions between and within the affected metabolic pathways under temperature stresses for comparative proteomic analysis. The developed tool, which is coded in HTML with CSS/JavaScript and depicted in Scalable Vector Graphics (SVG), is designed for interactive analysis and exploration of the constructed network. SpirPro is publicly available on the web at http://spirpro.sbi.kmutt.ac.th . SpirPro is an analysis platform containing an integrated proteome and PPI database that provides the most comprehensive data on this cyanobacterium at the systematic level. As an integrated database, SpirPro can be applied in various analyses, such as temperature stress response networking analysis in cyanobacterial models and interacting domain-domain analysis between proteins of interest.

Manganese-Induced Neurotoxicity and Alterations in Gene Expression in Human Neuroblastoma SH-SY5Y Cells.

PubMed

Gandhi, Deepa; Sivanesan, Saravanadevi; Kannan, Krishnamurthi

2018-06-01

Manganese (Mn) is an essential trace element required for many physiological functions including proper biochemical and cellular functioning of the central nervous system (CNS). However, exposure to excess level of Mn through occupational settings or from environmental sources has been associated with neurotoxicity. The cellular and molecular mechanism of Mn-induced neurotoxicity remains unclear. In the current study, we investigated the effects of 30-day exposure to a sub-lethal concentration of Mn (100 μM) in human neuroblastoma cells (SH-SY5Y) using transcriptomic approach. Microarray analysis revealed differential expression of 1057 transcripts in Mn-exposed SH-SY5Y cells as compared to control cells. Gene functional annotation cluster analysis exhibited that the differentially expressed genes were associated with several biological pathways. Specifically, genes involved in neuronal pathways including neuron differentiation and development, regulation of neurogenesis, synaptic transmission, and neuronal cell death (apoptosis) were found to be significantly altered. KEGG pathway analysis showed upregulation of p53 signaling pathways and neuroactive ligand-receptor interaction pathways, and downregulation of neurotrophin signaling pathway. On the basis of the gene expression profile, possible molecular mechanisms underlying Mn-induced neuronal toxicity were predicted.

cPath: open source software for collecting, storing, and querying biological pathways

PubMed Central

Cerami, Ethan G; Bader, Gary D; Gross, Benjamin E; Sander, Chris

2006-01-01

Background Biological pathways, including metabolic pathways, protein interaction networks, signal transduction pathways, and gene regulatory networks, are currently represented in over 220 diverse databases. These data are crucial for the study of specific biological processes, including human diseases. Standard exchange formats for pathway information, such as BioPAX, CellML, SBML and PSI-MI, enable convenient collection of this data for biological research, but mechanisms for common storage and communication are required. Results We have developed cPath, an open source database and web application for collecting, storing, and querying biological pathway data. cPath makes it easy to aggregate custom pathway data sets available in standard exchange formats from multiple databases, present pathway data to biologists via a customizable web interface, and export pathway data via a web service to third-party software, such as Cytoscape, for visualization and analysis. cPath is software only, and does not include new pathway information. Key features include: a built-in identifier mapping service for linking identical interactors and linking to external resources; built-in support for PSI-MI and BioPAX standard pathway exchange formats; a web service interface for searching and retrieving pathway data sets; and thorough documentation. The cPath software is freely available under the LGPL open source license for academic and commercial use. Conclusion cPath is a robust, scalable, modular, professional-grade software platform for collecting, storing, and querying biological pathways. It can serve as the core data handling component in information systems for pathway visualization, analysis and modeling. PMID:17101041

Concept mapping One-Carbon Metabolism to model future ontologies for nutrient-gene-phenotype interactions.

PubMed

Joslin, A C; Green, R; German, J B; Lange, M C

2014-09-01

Advances in the development of bioinformatic tools continue to improve investigators' ability to interrogate, organize, and derive knowledge from large amounts of heterogeneous information. These tools often require advanced technical skills not possessed by life scientists. User-friendly, low-barrier-to-entry methods of visualizing nutrigenomics information are yet to be developed. We utilized concept mapping software from the Institute for Human and Machine Cognition to create a conceptual model of diet and health-related data that provides a foundation for future nutrigenomics ontologies describing published nutrient-gene/polymorphism-phenotype data. In this model, maps containing phenotype, nutrient, gene product, and genetic polymorphism interactions are visualized as triples of two concepts linked together by a linking phrase. These triples, or "knowledge propositions," contextualize aggregated data and information into easy-to-read knowledge maps. Maps of these triples enable visualization of genes spanning the One-Carbon Metabolism (OCM) pathway, their sequence variants, and multiple literature-mined associations including concepts relevant to nutrition, phenotypes, and health. The concept map development process documents the incongruity of information derived from pathway databases versus literature resources. This conceptual model highlights the importance of incorporating information about genes in upstream pathways that provide substrates, as well as downstream pathways that utilize products of the pathway under investigation, in this case OCM. Other genes and their polymorphisms, such as TCN2 and FUT2, although not directly involved in OCM, potentially alter OCM pathway functionality. These upstream gene products regulate substrates such as B12. Constellations of polymorphisms affecting the functionality of genes along OCM, together with substrate and cofactor availability, may impact resultant phenotypes. These conceptual maps provide a foundational framework for development of nutrient-gene/polymorphism-phenotype ontologies and systems visualization.

Plasmacytoid Dendritic Cells Require Direct Infection To Sustain the Pulmonary Influenza A Virus-Specific CD8 T Cell Response

PubMed Central

Hemann, Emily A.; Sjaastad, Louisa E.; Langlois, Ryan A.

2015-01-01

ABSTRACT Following influenza A virus (IAV) infection, development of a robust IAV-specific CD8 T cell response is required for clearance of primary infection and enhances memory protection. Following IAV infection, plasmacytoid dendritic cells (pDC) or CD8α+ DC regulate pulmonary effector CD8 T cell responses within the lung. Without this DC-T cell interaction, insufficient effector CD8 T cells are maintained in the lungs, leading to enhanced morbidity and mortality. Previous studies have demonstrated that pDC are capable of classical presentation or cross-presentation of IAV antigens and could potentially regulate IAV-specific CD8 T cell responses through either mechanism. Our results demonstrate that pDC from the lungs of donor mice infected with an IAV that is not able to replicate in hematopoietic cells (142t-IAV), unlike donor pDC isolated from the lungs of control infected mice, are not able to rescue the host IAV-specific CD8 T cell response from apoptosis. This indicates that pDC must utilize the direct presentation pathway for this rescue. This inability of pDC from 142t-IAV donors to rescue the IAV-specific CD8 T cell response is not due to differences in the overall ability of 142t-IAV to replicate within the lungs or generate defective viral genomes or to differences in levels of costimulatory molecules required for this interaction. We further demonstrate that bypassing the antigen presentation pathway by coating the 142t-IAV pDC with IAV peptide epitopes restores their ability to rescue the IAV-specific CD8 T cell response. IMPORTANCE IAV continues to be a global health burden, infecting 5 to 20% of the global population annually. Continued investigation into the mechanisms that mediate protective immune responses against IAV is important to improving current vaccination and antiviral strategies antagonistic toward IAV. Our findings presented herein demonstrate a key requirement for pDC promotion of effector CD8 T cell survival: that rather than utilizing cross-presentation, pDC must be infected and utilize the endogenous pathway for presentation of antigens to CD8 T cells during in vivo IAV infections. This suggests that targeting presentation via the endogenous pathway in pDC could be important for the development of unique antiviral cellular therapies. PMID:26719269

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

Trait- and density-mediated indirect interactions initiated by an exotic invasive plant autogenic ecosystem engineer

Treesearch

Dean E. Pearson

2010-01-01

Indirect interactions are important for structuring ecological systems. However, research on indirect effects has been heavily biased toward top-down trophic interactions, and less is known about other indirect-interaction pathways. As autogenic ecosystem engineers, plants can serve as initiators of nontrophic indirect interactions that, like top-down pathways, can...

HPIminer: A text mining system for building and visualizing human protein interaction networks and pathways.

PubMed

Subramani, Suresh; Kalpana, Raja; Monickaraj, Pankaj Moses; Natarajan, Jeyakumar

2015-04-01

The knowledge on protein-protein interactions (PPI) and their related pathways are equally important to understand the biological functions of the living cell. Such information on human proteins is highly desirable to understand the mechanism of several diseases such as cancer, diabetes, and Alzheimer's disease. Because much of that information is buried in biomedical literature, an automated text mining system for visualizing human PPI and pathways is highly desirable. In this paper, we present HPIminer, a text mining system for visualizing human protein interactions and pathways from biomedical literature. HPIminer extracts human PPI information and PPI pairs from biomedical literature, and visualize their associated interactions, networks and pathways using two curated databases HPRD and KEGG. To our knowledge, HPIminer is the first system to build interaction networks from literature as well as curated databases. Further, the new interactions mined only from literature and not reported earlier in databases are highlighted as new. A comparative study with other similar tools shows that the resultant network is more informative and provides additional information on interacting proteins and their associated networks. Copyright © 2015 Elsevier Inc. All rights reserved.

Inner nuclear envelope protein SUN1 plays a prominent role in mammalian mRNA export.

PubMed

Li, Ping; Noegel, Angelika A

2015-11-16

Nuclear export of messenger ribonucleoproteins (mRNPs) through the nuclear pore complex (NPC) can be roughly classified into two forms: bulk and specific export, involving an nuclear RNA export factor 1 (NXF1)-dependent pathway and chromosome region maintenance 1 (CRM1)-dependent pathway, respectively. SUN proteins constitute the inner nuclear envelope component of the l I: nker of N: ucleoskeleton and C: ytoskeleton (LINC) complex. Here, we show that mammalian cells require SUN1 for efficient nuclear mRNP export. The results indicate that both SUN1 and SUN2 interact with heterogeneous nuclear ribonucleoprotein (hnRNP) F/H and hnRNP K/J. SUN1 depletion inhibits the mRNP export, with accumulations of both hnRNPs and poly(A)+RNA in the nucleus. Leptomycin B treatment indicates that SUN1 functions in mammalian mRNA export involving the NXF1-dependent pathway. SUN1 mediates mRNA export through its association with mRNP complexes via a direct interaction with NXF1. Additionally, SUN1 associates with the NPC through a direct interaction with Nup153, a nuclear pore component involved in mRNA export. Taken together, our results reveal that the inner nuclear envelope protein SUN1 has additional functions aside from being a central component of the LINC complex and that it is an integral component of the mammalian mRNA export pathway suggesting a model whereby SUN1 recruits NXF1-containing mRNP onto the nuclear envelope and hands it over to Nup153. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Mechanisms of action of Coxiella burnetii effectors inferred from host-pathogen protein interactions.

PubMed

Wallqvist, Anders; Wang, Hao; Zavaljevski, Nela; Memišević, Vesna; Kwon, Keehwan; Pieper, Rembert; Rajagopala, Seesandra V; Reifman, Jaques

2017-01-01

Coxiella burnetii is an obligate Gram-negative intracellular pathogen and the etiological agent of Q fever. Successful infection requires a functional Type IV secretion system, which translocates more than 100 effector proteins into the host cytosol to establish the infection, restructure the intracellular host environment, and create a parasitophorous vacuole where the replicating bacteria reside. We used yeast two-hybrid (Y2H) screening of 33 selected C. burnetii effectors against whole genome human and murine proteome libraries to generate a map of potential host-pathogen protein-protein interactions (PPIs). We detected 273 unique interactions between 20 pathogen and 247 human proteins, and 157 between 17 pathogen and 137 murine proteins. We used orthology to combine the data and create a single host-pathogen interaction network containing 415 unique interactions between 25 C. burnetii and 363 human proteins. We further performed complementary pairwise Y2H testing of 43 out of 91 C. burnetii-human interactions involving five pathogen proteins. We used the combined data to 1) perform enrichment analyses of target host cellular processes and pathways, 2) examine effectors with known infection phenotypes, and 3) infer potential mechanisms of action for four effectors with uncharacterized functions. The host-pathogen interaction profiles supported known Coxiella phenotypes, such as adapting cell morphology through cytoskeletal re-arrangements, protein processing and trafficking, organelle generation, cholesterol processing, innate immune modulation, and interactions with the ubiquitin and proteasome pathways. The generated dataset of PPIs-the largest collection of unbiased Coxiella host-pathogen interactions to date-represents a rich source of information with respect to secreted pathogen effector proteins and their interactions with human host proteins.

Inhibition of Ku70 acetylation by INHAT subunit SET/TAF-Iβ regulates Ku70-mediated DNA damage response.

PubMed

Kim, Kee-Beom; Kim, Dong-Wook; Park, Jin Woo; Jeon, Young-Joo; Kim, Daehwan; Rhee, Sangmyung; Chae, Jung-Il; Seo, Sang-Beom

2014-07-01

DNA double-strand breaks (DSBs) can cause either cell death or genomic instability. The Ku heterodimer Ku70/80 is required for the NHEJ (non-homologous end-joining) DNA DSB repair pathway. The INHAT (inhibitor of histone acetyltransferases) complex subunit, SET/TAF-Iβ, can inhibit p300- and PCAF-mediated acetylation of both histone and p53, thereby repressing general transcription and that of p53 target genes. Here, we show that SET/TAF-Iβ interacts with Ku70/80, and that this interaction inhibits CBP- and PCAF-mediated Ku70 acetylation in an INHAT domain-dependent manner. Notably, DNA damage by UV disrupted the interaction between SET/TAF-Iβ and Ku70. Furthermore, we demonstrate that overexpressed SET/TAF-Iβ inhibits recruitment of Ku70/80 to DNA damage sites. We propose that dysregulation of SET/TAF-Iβ expression prevents repair of damaged DNA and also contributes to cellular proliferation. All together, our findings indicate that SET/TAF-Iβ interacts with Ku70/80 in the nucleus and inhibits Ku70 acetylation. Upon DNA damage, SET/TAF-Iβ dissociates from the Ku complex and releases Ku70/Ku80, which are then recruited to DNA DSB sites via the NHEJ DNA repair pathway.

Enteric neural crest cells regulate vertebrate stomach patterning and differentiation.

PubMed

Faure, Sandrine; McKey, Jennifer; Sagnol, Sébastien; de Santa Barbara, Pascal

2015-01-15

In vertebrates, the digestive tract develops from a uniform structure where reciprocal epithelial-mesenchymal interactions pattern this complex organ into regions with specific morphologies and functions. Concomitant with these early patterning events, the primitive GI tract is colonized by the vagal enteric neural crest cells (vENCCs), a population of cells that will give rise to the enteric nervous system (ENS), the intrinsic innervation of the GI tract. The influence of vENCCs on early patterning and differentiation of the GI tract has never been evaluated. In this study, we report that a crucial number of vENCCs is required for proper chick stomach development, patterning and differentiation. We show that reducing the number of vENCCs by performing vENCC ablations induces sustained activation of the BMP and Notch pathways in the stomach mesenchyme and impairs smooth muscle development. A reduction in vENCCs also leads to the transdifferentiation of the stomach into a stomach-intestinal mixed phenotype. In addition, sustained Notch signaling activity in the stomach mesenchyme phenocopies the defects observed in vENCC-ablated stomachs, indicating that inhibition of the Notch signaling pathway is essential for stomach patterning and differentiation. Finally, we report that a crucial number of vENCCs is also required for maintenance of stomach identity and differentiation through inhibition of the Notch signaling pathway. Altogether, our data reveal that, through the regulation of mesenchyme identity, vENCCs act as a new mediator in the mesenchymal-epithelial interactions that control stomach development. © 2015. Published by The Company of Biologists Ltd.

MG53 participates in ischaemic postconditioning through the RISK signalling pathway

PubMed Central

Zhang, Yan; Lv, Fengxiang; Jin, Li; Peng, Wei; Song, Ruisheng; Ma, Jianjie; Cao, Chun-Mei; Xiao, Rui-Ping

2011-01-01

Aims Recent studies show that ischaemic postconditioning (PostC), similar to the well-established ischaemic preconditioning (IPC), confers cardioprotection against ischaemia/reperfusion (IR) injury, and both IPC and PostC can activate the reperfusion injury salvage kinase (RISK) pathway and the survivor activating factor enhancement (SAFE) pathway. PostC is clinically more attractive because of its therapeutic application at the predictable onset of reperfusion. Our previous studies have demonstrated that MG53 is a primary component of the IPC machinery. Here, we investigated the potential role of MG53 in PostC-mediated myocardial protection and explored the underlying mechanism. Methods and results Using Langendorff perfusion, we investigated IR injury in wild-type (wt) and MG53-deficient (mg53−/−) mouse hearts with or without PostC. IR-induced myocardial damage was markedly exacerbated in mg53−/− hearts compared with wt controls. PostC protected wt hearts against IR-induced myocardial infarction, myocyte necrosis, and apoptosis, but failed to protect mg53−/− hearts. The loss of PostC protection in mg53−/− hearts was attributed to selectively impaired PostC-activated RISK signalling. Mechanistically, MG53 is required for the interaction between caveolin 3 (CaV3) and the p85 subunit of phosphoinositide 3-kinase (p85-PI3K) and PostC-mediated activation of the RISK pathway. Importantly, a structure–function study revealed that the MG53 tripartite motif (TRIM) domain (aa1–284) physically interacted with CaV3 but not p85-PI3K, whereas the MG53 SPRY domain (aa285–477) interacted with p85-PI3K but not CaV3, indicating that MG53 binds to CaV3 and p85 at its N- and C-terminus, respectively. Conclusions We conclude that MG53 participates in PostC-mediated cardioprotection largely through tethering CaV3 and PI3K and subsequent activation of the RISK pathway. PMID:21285295

In silico analysis of the potential mechanism of telocinobufagin on breast cancer MCF-7 cells.

PubMed

Dang, Yi-Wu; Lin, Peng; Liu, Li-Min; He, Rong-Quan; Zhang, Li-Jie; Peng, Zhi-Gang; Li, Xiao-Jiao; Chen, Gang

2018-05-01

The extractives from a ChanSu, traditional Chinese medicine, have been discovered to possess anti-inflammatory and tumor-suppressing abilities. However, the molecular mechanism of telocinobufagin, a compound extracted from ChanSu, on breast cancer cells has not been clarified. The aim of this study is to investigate the underlying mechanism of telocinobufagin on breast cancer cells. The differentially expressed genes after telocinobufagin treatment on breast cancer cells were searched and downloaded from Gene Expression Omnibus (GEO), ArrayExpress and literatures. Bioinformatics tools were applied to further explore the potential mechanism of telocinobufagin in breast cancer using the Kyoto Encyclopedia of genes and genomes (KEGG) pathway, Gene ontology (GO) enrichment, panther, and protein-protein interaction analyses. To better comprehend the role of telocinobufagin in breast cancer, we also queried the Connectivity Map using the gene expression profiles of telocinobufagin treatment. One GEO accession (GSE85871) provided 1251 differentially expressed genes after telocinobufagin treatment on MCF-7 cells. The pathway of neuroactive ligand-receptor interaction, cell adhesion molecules (CAMs), intestinal immune network for IgA production, hematopoietic cell lineage and calcium signaling pathway were the key pathways from KEGG analysis. IGF1 and KSR1, owning to higher protein levels in breast cancer tissues, IGF1 and KSR1 could be the hub genes related to telocinobufagin treatment. It was indicated that the molecular mechanism of telocinobufagin resembled that of fenspiride. Telocinobufagin might regulate neuroactive ligand-receptor interaction pathway to exert its influences in breast cancer MCF-7 cells, and its molecular mechanism might share some similarities with fenspiride. This study only presented a comprehensive picture of the role of telocinobufagin in breast cancer MCF-7 cells using big data. However, more thorough and deeper researches are required to add to the validity of this study. Copyright © 2018 Elsevier GmbH. All rights reserved.

Framing Ethnic Variations in Alcohol Outcomes from Biological Pathways to Neighborhood Context

PubMed Central

Chartier, Karen G.; Scott, Denise M.; Wall, Tamara L.; Covault, Jonathan; Karriker-Jaffe, Katherine J.; Mills, Britain A.; Luczak, Susan E.; Caetano, Raul; Arroyo, Judith A.

2013-01-01

Health disparities research seeks to eliminate disproportionate negative health outcomes experienced in some racial/ethnic minority groups. This brief review presents findings on factors associated with drinking and alcohol-related problems in racial/ethnic groups. Those discussed are: 1) biological pathways to alcohol problems, 2) gene by stress interactions, 3) neighborhood disadvantage, stress, and access to alcohol, and 4) drinking cultures and contexts. These factors and their interrelationships are complex, requiring a multi-level perspective. The use of interdisciplinary teams and an epigenetic focus are suggested to move the research forward. The application of multi-level research to policy, prevention, and intervention programs may help prioritize combinations of the most promising intervention targets. PMID:24483624

Inferring gene and protein interactions using PubMed citations and consensus Bayesian networks

PubMed Central

Dalman, Mark; Haddad, Joseph; Duan, Zhong-Hui

2017-01-01

The PubMed database offers an extensive set of publication data that can be useful, yet inherently complex to use without automated computational techniques. Data repositories such as the Genomic Data Commons (GDC) and the Gene Expression Omnibus (GEO) offer experimental data storage and retrieval as well as curated gene expression profiles. Genetic interaction databases, including Reactome and Ingenuity Pathway Analysis, offer pathway and experiment data analysis using data curated from these publications and data repositories. We have created a method to generate and analyze consensus networks, inferring potential gene interactions, using large numbers of Bayesian networks generated by data mining publications in the PubMed database. Through the concept of network resolution, these consensus networks can be tailored to represent possible genetic interactions. We designed a set of experiments to confirm that our method is stable across variation in both sample and topological input sizes. Using gene product interactions from the KEGG pathway database and data mining PubMed publication abstracts, we verify that regardless of the network resolution or the inferred consensus network, our method is capable of inferring meaningful gene interactions through consensus Bayesian network generation with multiple, randomized topological orderings. Our method can not only confirm the existence of currently accepted interactions, but has the potential to hypothesize new ones as well. We show our method confirms the existence of known gene interactions such as JAK-STAT-PI3K-AKT-mTOR, infers novel gene interactions such as RAS- Bcl-2 and RAS-AKT, and found significant pathway-pathway interactions between the JAK-STAT signaling and Cardiac Muscle Contraction KEGG pathways. PMID:29049295

MicroRNA-Dependent Transcriptional Silencing of Transposable Elements in Drosophila Follicle Cells.

PubMed

Mugat, Bruno; Akkouche, Abdou; Serrano, Vincent; Armenise, Claudia; Li, Blaise; Brun, Christine; Fulga, Tudor A; Van Vactor, David; Pélisson, Alain; Chambeyron, Séverine

2015-05-01

RNA interference-related silencing mechanisms concern very diverse and distinct biological processes, from gene regulation (via the microRNA pathway) to defense against molecular parasites (through the small interfering RNA and the Piwi-interacting RNA pathways). Small non-coding RNAs serve as specificity factors that guide effector proteins to ribonucleic acid targets via base-pairing interactions, to achieve transcriptional or post-transcriptional regulation. Because of the small sequence complementarity required for microRNA-dependent post-transcriptional regulation, thousands of microRNA (miRNA) putative targets have been annotated in Drosophila. In Drosophila somatic ovarian cells, genomic parasites, such as transposable elements (TEs), are transcriptionally repressed by chromatin changes induced by Piwi-interacting RNAs (piRNAs) that prevent them from invading the germinal genome. Here we show, for the first time, that a functional miRNA pathway is required for the piRNA-mediated transcriptional silencing of TEs in this tissue. Global miRNA depletion, caused by tissue- and stage-specific knock down of drosha (involved in miRNA biogenesis), AGO1 or gawky (both responsible for miRNA activity), resulted in loss of TE-derived piRNAs and chromatin-mediated transcriptional de-silencing of TEs. This specific TE de-repression was also observed upon individual titration (by expression of the complementary miRNA sponge) of two miRNAs (miR-14 and miR-34) as well as in a miR-14 loss-of-function mutant background. Interestingly, the miRNA defects differentially affected TE- and 3' UTR-derived piRNAs. To our knowledge, this is the first indication of possible differences in the biogenesis or stability of TE- and 3' UTR-derived piRNAs. This work is one of the examples of detectable phenotypes caused by loss of individual miRNAs in Drosophila and the first genetic evidence that miRNAs have a role in the maintenance of genome stability via piRNA-mediated TE repression.

Cytolethal distending toxins require components of the ER-associated degradation pathway for host cell entry.

PubMed

Eshraghi, Aria; Dixon, Shandee D; Tamilselvam, Batcha; Kim, Emily Jin-Kyung; Gargi, Amandeep; Kulik, Julia C; Damoiseaux, Robert; Blanke, Steven R; Bradley, Kenneth A

2014-07-01

Intracellular acting protein exotoxins produced by bacteria and plants are important molecular determinants that drive numerous human diseases. A subset of these toxins, the cytolethal distending toxins (CDTs), are encoded by several Gram-negative pathogens and have been proposed to enhance virulence by allowing evasion of the immune system. CDTs are trafficked in a retrograde manner from the cell surface through the Golgi apparatus and into the endoplasmic reticulum (ER) before ultimately reaching the host cell nucleus. However, the mechanism by which CDTs exit the ER is not known. Here we show that three central components of the host ER associated degradation (ERAD) machinery, Derlin-2 (Derl2), the E3 ubiquitin-protein ligase Hrd1, and the AAA ATPase p97, are required for intoxication by some CDTs. Complementation of Derl2-deficient cells with Derl2:Derl1 chimeras identified two previously uncharacterized functional domains in Derl2, the N-terminal 88 amino acids and the second ER-luminal loop, as required for intoxication by the CDT encoded by Haemophilus ducreyi (Hd-CDT). In contrast, two motifs required for Derlin-dependent retrotranslocation of ERAD substrates, a conserved WR motif and an SHP box that mediates interaction with the AAA ATPase p97, were found to be dispensable for Hd-CDT intoxication. Interestingly, this previously undescribed mechanism is shared with the plant toxin ricin. These data reveal a requirement for multiple components of the ERAD pathway for CDT intoxication and provide insight into a Derl2-dependent pathway exploited by retrograde trafficking toxins.

Cytolethal Distending Toxins Require Components of the ER-Associated Degradation Pathway for Host Cell Entry

PubMed Central

Eshraghi, Aria; Dixon, Shandee D.; Tamilselvam, Batcha; Kim, Emily Jin-Kyung; Gargi, Amandeep; Kulik, Julia C.; Damoiseaux, Robert; Blanke, Steven R.; Bradley, Kenneth A.

2014-01-01

Intracellular acting protein exotoxins produced by bacteria and plants are important molecular determinants that drive numerous human diseases. A subset of these toxins, the cytolethal distending toxins (CDTs), are encoded by several Gram-negative pathogens and have been proposed to enhance virulence by allowing evasion of the immune system. CDTs are trafficked in a retrograde manner from the cell surface through the Golgi apparatus and into the endoplasmic reticulum (ER) before ultimately reaching the host cell nucleus. However, the mechanism by which CDTs exit the ER is not known. Here we show that three central components of the host ER associated degradation (ERAD) machinery, Derlin-2 (Derl2), the E3 ubiquitin-protein ligase Hrd1, and the AAA ATPase p97, are required for intoxication by some CDTs. Complementation of Derl2-deficient cells with Derl2:Derl1 chimeras identified two previously uncharacterized functional domains in Derl2, the N-terminal 88 amino acids and the second ER-luminal loop, as required for intoxication by the CDT encoded by Haemophilus ducreyi (Hd-CDT). In contrast, two motifs required for Derlin-dependent retrotranslocation of ERAD substrates, a conserved WR motif and an SHP box that mediates interaction with the AAA ATPase p97, were found to be dispensable for Hd-CDT intoxication. Interestingly, this previously undescribed mechanism is shared with the plant toxin ricin. These data reveal a requirement for multiple components of the ERAD pathway for CDT intoxication and provide insight into a Derl2-dependent pathway exploited by retrograde trafficking toxins. PMID:25078082

Non-covalent Interactions with SUMO and Ubiquitin Orchestrate Distinct Functions of the SLX4 Complex in Genome Maintenance

PubMed Central

Ouyang, Jian; Garner, Elizabeth; Hallet, Alexander; Nguyen, Hai Dang; Rickman, Kimberly A.; Gill, Grace; Smogorzewska, Agata; Zou, Lee

2014-01-01

SLX4, a coordinator of multiple DNA structure-specific endonucleases, is important for several DNA repair pathways. Non-covalent interactions of SLX4 with ubiquitin are required for localizing SLX4 to DNA-interstrand crosslinks (ICLs), yet how SLX4 is targeted to other functional contexts remains unclear. Here, we show that SLX4 binds SUMO-2/3 chains via SUMO-interacting motifs (SIMs). The SIMs of SLX4 are dispensable for ICL repair, but important for processing CPT-induced replication intermediates, suppressing fragile site instability, and localizing SLX4 to ALT telomeres. The localization of SLX4 to laser-induced DNA damage also requires the SIMs, as well as DNA-end resection, UBC9 and MDC1. Furthermore, the SUMO binding of SLX4 enhances its interaction with specific DNA-damage sensors or telomere-binding proteins, including RPA, MRE11-RAD50-NBS1 and TRF2. Thus, the interactions of SLX4 with SUMO and ubiquitin increase its affinity for factors recognizing different DNA lesions or telomeres, helping to direct the SLX4 complex in distinct functional contexts. PMID:25533185

Wnt pathway curation using automated natural language processing: combining statistical methods with partial and full parse for knowledge extraction.

PubMed

Santos, Carlos; Eggle, Daniela; States, David J

2005-04-15

Wnt signaling is a very active area of research with highly relevant publications appearing at a rate of more than one per day. Building and maintaining databases describing signal transduction networks is a time-consuming and demanding task that requires careful literature analysis and extensive domain-specific knowledge. For instance, more than 50 factors involved in Wnt signal transduction have been identified as of late 2003. In this work we describe a natural language processing (NLP) system that is able to identify references to biological interaction networks in free text and automatically assembles a protein association and interaction map. A 'gold standard' set of names and assertions was derived by manual scanning of the Wnt genes website (http://www.stanford.edu/~rnusse/wntwindow.html) including 53 interactions involved in Wnt signaling. This system was used to analyze a corpus of peer-reviewed articles related to Wnt signaling including 3369 Pubmed and 1230 full text papers. Names for key Wnt-pathway associated proteins and biological entities are identified using a chi-squared analysis of noun phrases over-represented in the Wnt literature as compared to the general signal transduction literature. Interestingly, we identified several instances where generic terms were used on the website when more specific terms occur in the literature, and one typographic error on the Wnt canonical pathway. Using the named entity list and performing an exhaustive assertion extraction of the corpus, 34 of the 53 interactions in the 'gold standard' Wnt signaling set were successfully identified (64% recall). In addition, the automated extraction found several interactions involving key Wnt-related molecules which were missing or different from those in the canonical diagram, and these were confirmed by manual review of the text. These results suggest that a combination of NLP techniques for information extraction can form a useful first-pass tool for assisting human annotation and maintenance of signal pathway databases. The pipeline software components are freely available on request to the authors. dstates@umich.edu http://stateslab.bioinformatics.med.umich.edu/software.html.

Casein Kinase 2 Reverses Tail-Independent Inhibition of Kinesin-1

NASA Astrophysics Data System (ADS)

Xu, Jing; Shu, Zhanyong; Anand, Preetha; Reddy, Babu; Cermelli, Silvia; Whisenant, Thomas; King, Stephen; Bardwell, Lee; Huang, Lan; Gross, Steven

2011-03-01

Kinesin-1 is a plus-end microtubule-based molecular motor, and defects in kinesin transport are linked to diseases including neurodegeneration. Kinesin can auto-inhibit via a direct head-tail interaction, but is believed to be active otherwise. In contrast, this study uncovers a fast but reversible inhibition distinct from the canonical auto-inhibition pathway. The majority of the initially active kinesin (full-length or tail-less) loses its ability to bind/interact with microtubule, and Casein Kinase 2 (CK2) reverses this inactivation (up to 4-fold) without altering kinesin's single motor properties. Motor phosphorylation is not required for this CK2 -mediated kinesin activation. In cultured mammalian cells, knockdown of CK2 level, but not kinase activity, was sufficient to decrease the force required to stall lipid droplet transport, consistent with a reduction in the number of active motors. We propose that CK2 forms a positive regulating complex with the motor. This study provides the first direct evidence of a protein kinase positively regulating kinesin-transport, and uncovers a pathway whereby inactive cargo-bound kinesin can be activated. This work is supported by NIGMS grants GM64624 and GM079156 to SPG, GM-74830 to LH, NIH grants GM76516 and GM60366 to LB, and AHA grant 825278F to JX.

A Synthetic Interaction Screen Identifies Factors Selectively Required for Proliferation and TERT Transcription in p53-Deficient Human Cancer Cells

PubMed Central

Park, Sung Mi; Zhu, Lihua J.; Debily, Marie-anne; Kittler, Ellen L. W.; Zapp, Maria L.; Lapointe, David; Gobeil, Stephane; Virbasius, Ching-Man; Green, Michael R.

2012-01-01

Numerous genetic and epigenetic alterations render cancer cells selectively dependent on specific genes and regulatory pathways, and represent potential vulnerabilities that can be therapeutically exploited. Here we describe an RNA interference (RNAi)–based synthetic interaction screen to identify genes preferentially required for proliferation of p53-deficient (p53−) human cancer cells. We find that compared to p53-competent (p53+) human cancer cell lines, diverse p53− human cancer cell lines are preferentially sensitive to loss of the transcription factor ETV1 and the DNA damage kinase ATR. In p53− cells, RNAi–mediated knockdown of ETV1 or ATR results in decreased expression of the telomerase catalytic subunit TERT leading to growth arrest, which can be reversed by ectopic TERT expression. Chromatin immunoprecipitation analysis reveals that ETV1 binds to a region downstream of the TERT transcriptional start-site in p53− but not p53+ cells. We find that the role of ATR is to phosphorylate and thereby stabilize ETV1. Our collective results identify a regulatory pathway involving ETV1, ATR, and TERT that is preferentially important for proliferation of diverse p53− cancer cells. PMID:23284306

Drosophila melanogaster auxilin regulates the internalization of Delta to control activity of the Notch signaling pathway

PubMed Central

Hagedorn, Elliott J.; Bayraktar, Jennifer L.; Kandachar, Vasundhara R.; Bai, Ting; Englert, Dane M.; Chang, Henry C.

2006-01-01

We have isolated mutations in the Drosophila melanogaster homologue of auxilin, a J-domain–containing protein known to cooperate with Hsc70 in the disassembly of clathrin coats from clathrin-coated vesicles in vitro. Consistent with this biochemical role, animals with reduced auxilin function exhibit genetic interactions with Hsc70 and clathrin. Interestingly, the auxilin mutations interact specifically with Notch and disrupt several Notch-mediated processes. Genetic evidence places auxilin function in the signal-sending cells, upstream of Notch receptor activation, suggesting that the relevant cargo for this auxilin-mediated endocytosis is the Notch ligand Delta. Indeed, the localization of Delta protein is disrupted in auxilin mutant tissues. Thus, our data suggest that auxilin is an integral component of the Notch signaling pathway, participating in the ubiquitin-dependent endocytosis of Delta. Furthermore, the fact that auxilin is required for Notch signaling suggests that ligand endocytosis in the signal-sending cells needs to proceed past coat disassembly to activate Notch. PMID:16682530

A Protein Scaffold Coordinates SRC-Mediated JNK Activation in Response to Metabolic Stress.

PubMed

Kant, Shashi; Standen, Claire L; Morel, Caroline; Jung, Dae Young; Kim, Jason K; Swat, Wojciech; Flavell, Richard A; Davis, Roger J

2017-09-19

Obesity is a major risk factor for the development of metabolic syndrome and type 2 diabetes. How obesity contributes to metabolic syndrome is unclear. Free fatty acid (FFA) activation of a non-receptor tyrosine kinase (SRC)-dependent cJun NH 2 -terminal kinase (JNK) signaling pathway is implicated in this process. However, the mechanism that mediates SRC-dependent JNK activation is unclear. Here, we identify a role for the scaffold protein JIP1 in SRC-dependent JNK activation. SRC phosphorylation of JIP1 creates phosphotyrosine interaction motifs that bind the SH2 domains of SRC and the guanine nucleotide exchange factor VAV. These interactions are required for SRC-induced activation of VAV and the subsequent engagement of a JIP1-tethered JNK signaling module. The JIP1 scaffold protein, therefore, plays a dual role in FFA signaling by coordinating upstream SRC functions together with downstream effector signaling by the JNK pathway. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

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

Fields, Whitney; Kielian, Margaret, E-mail: margaret.kielian@einstein.yu.edu

The alphavirus membrane protein E1 mediates low pH-triggered fusion of the viral and endosome membranes during virus entry. During virus biogenesis E1 associates as a heterodimer with the transmembrane protein p62. Late in the secretory pathway, cellular furin cleaves p62 to the mature E2 protein and a peripheral protein E3. E3 remains bound to E2 at low pH, stabilizing the heterodimer and thus protecting E1 from the acidic pH of the secretory pathway. Release of E3 at neutral pH then primes the virus for fusion during entry. Here we used site-directed mutagenesis and revertant analysis to define residues important formore » the interactions at the E3–E2 interface. Our data identified a key residue, E2 W235, which was required for E1 pH protection and alphavirus production. Our data also suggest additional residues on E3 and E2 that affect their interacting surfaces and thus influence the pH protection of E1 during alphavirus exit.« less

Sestrin2 is a leucine sensor for the mTORC1 pathway.

PubMed

Wolfson, Rachel L; Chantranupong, Lynne; Saxton, Robert A; Shen, Kuang; Scaria, Sonia M; Cantor, Jason R; Sabatini, David M

2016-01-01

Leucine is a proteogenic amino acid that also regulates many aspects of mammalian physiology, in large part by activating the mTOR complex 1 (mTORC1) protein kinase, a master growth controller. Amino acids signal to mTORC1 through the Rag guanosine triphosphatases (GTPases). Several factors regulate the Rags, including GATOR1, aGTPase-activating protein; GATOR2, a positive regulator of unknown function; and Sestrin2, a GATOR2-interacting protein that inhibits mTORC1 signaling. We find that leucine, but not arginine, disrupts the Sestrin2-GATOR2 interaction by binding to Sestrin2 with a dissociation constant of 20 micromolar, which is the leucine concentration that half-maximally activates mTORC1. The leucine-binding capacity of Sestrin2 is required for leucine to activate mTORC1 in cells. These results indicate that Sestrin2 is a leucine sensor for the mTORC1 pathway. Copyright © 2016, American Association for the Advancement of Science.

Clathrin light chains are required for the gyrating-clathrin recycling pathway and thereby promote cell migration.

PubMed

Majeed, Sophia R; Vasudevan, Lavanya; Chen, Chih-Ying; Luo, Yi; Torres, Jorge A; Evans, Timothy M; Sharkey, Andrew; Foraker, Amy B; Wong, Nicole M L; Esk, Christopher; Freeman, Theresa A; Moffett, Ashley; Keen, James H; Brodsky, Frances M

2014-05-23

The clathrin light chain (CLC) subunits participate in several membrane traffic pathways involving both clathrin and actin, through binding the actin-organizing huntingtin-interacting proteins (Hip). However, CLCs are dispensable for clathrin-mediated endocytosis of many cargoes. Here we observe that CLC depletion affects cell migration through Hip binding and reduces surface expression of β1-integrin by interference with recycling following normal endocytosis of inactive β1-integrin. CLC depletion and expression of a modified CLC also inhibit the appearance of gyrating (G)-clathrin structures, known mediators of rapid recycling of transferrin receptor from endosomes. Expression of the modified CLC reduces β1-integrin and transferrin receptor recycling, as well as cell migration, implicating G-clathrin in these processes. Supporting a physiological role for CLC in migration, the CLCb isoform of CLC is upregulated in migratory human trophoblast cells during uterine invasion. Together, these studies establish CLCs as mediating clathrin-actin interactions needed for recycling by G-clathrin during migration.

Reconstruction of metabolic pathways by combining probabilistic graphical model-based and knowledge-based methods

PubMed Central

2014-01-01

Automatic reconstruction of metabolic pathways for an organism from genomics and transcriptomics data has been a challenging and important problem in bioinformatics. Traditionally, known reference pathways can be mapped into an organism-specific ones based on its genome annotation and protein homology. However, this simple knowledge-based mapping method might produce incomplete pathways and generally cannot predict unknown new relations and reactions. In contrast, ab initio metabolic network construction methods can predict novel reactions and interactions, but its accuracy tends to be low leading to a lot of false positives. Here we combine existing pathway knowledge and a new ab initio Bayesian probabilistic graphical model together in a novel fashion to improve automatic reconstruction of metabolic networks. Specifically, we built a knowledge database containing known, individual gene / protein interactions and metabolic reactions extracted from existing reference pathways. Known reactions and interactions were then used as constraints for Bayesian network learning methods to predict metabolic pathways. Using individual reactions and interactions extracted from different pathways of many organisms to guide pathway construction is new and improves both the coverage and accuracy of metabolic pathway construction. We applied this probabilistic knowledge-based approach to construct the metabolic networks from yeast gene expression data and compared its results with 62 known metabolic networks in the KEGG database. The experiment showed that the method improved the coverage of metabolic network construction over the traditional reference pathway mapping method and was more accurate than pure ab initio methods. PMID:25374614

Gene interactions in the DNA damage-response pathway identified by genome-wide RNA-interference analysis of synthetic lethality

PubMed Central

van Haaften, Gijs; Vastenhouw, Nadine L.; Nollen, Ellen A. A.; Plasterk, Ronald H. A.; Tijsterman, Marcel

2004-01-01

Here, we describe a systematic search for synthetic gene interactions in a multicellular organism, the nematode Caenorhabditis elegans. We established a high-throughput method to determine synthetic gene interactions by genome-wide RNA interference and identified genes that are required to protect the germ line against DNA double-strand breaks. Besides known DNA-repair proteins such as the C. elegans orthologs of TopBP1, RPA2, and RAD51, eight genes previously unassociated with a double-strand-break response were identified. Knockdown of these genes increased sensitivity to ionizing radiation and camptothecin and resulted in increased chromosomal nondisjunction. All genes have human orthologs that may play a role in human carcinogenesis. PMID:15326288

Electron Transfer Pathways Facilitating U(VI) Reduction by Fe(II) on Al- vs Fe-Oxides

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

Taylor, S. D.; Becker, U.; Rosso, K. M.

This study continues mechanistic development of heterogeneous electron transfer (ET) pathways at mineral surfaces in aquatic environments that enable the reduction U(VI) by surface-associated Fe(II). Using computational molecular simulation within the framework of Marcus Theory, our findings highlight the importance of the configurations and interaction of the electron donor and acceptor species with the substrate, with respect to influencing its electronic structure and thereby the ability of semiconducting minerals to facilitate ET. U(VI) reduction by surface-associated Fe(II) (adsorbed or structurally incorporated into the lattice) on an insulating, corundum (001) surface (α-Al2O3) occurs when proximal inner-sphere (IS) surface complexes are formed,more » such that ET occurs through a combination of direct exchange (i.e., Fe d- and U f-orbitals overlap through space) and superexchange via intervening surface oxygen atoms. U(VI) reduction by coadsorbed Fe(II) on the isostructural semiconducting hematite (α-Fe2O3) basal surface requires either their direct electronic interaction (e.g., IS complexation) or mediation of this interaction indirectly through the surface via an intrasurface pathway. Conceptually possible longer-range ET by charge-hopping through surface Fe atoms was investigated to determine whether this indirect pathway is competitive with direct ET. The calculations show that energy barriers are large for this conduction-based pathway; interfacial ET into the hematite surface is endothermic (+80.1 kJ/mol) and comprises the rate-limiting step (10–6 s–1). The presence of the IS adsorbates appears to weaken the electronic coupling between underlying Fe ions within the surface, resulting in slower intra-surface ET (10–5 s–1) than expected in the bulk basal plane. Our findings lay out first insights into donor-acceptor communication via a charge-hopping pathway through the surface for heterogeneous reduction of U(VI) by Fe(II) and help provide a basis for experimental interrogation of this important process at mineral-water interfaces.« less

Overexpression of hypoxia-inducible factor and metabolic pathways: possible targets of cancer.

PubMed

Singh, Davinder; Arora, Rohit; Kaur, Pardeep; Singh, Balbir; Mannan, Rahul; Arora, Saroj

2017-01-01

Cancer, the main cause of human deaths in the modern world is a group of diseases. Anticancer drug discovery is a challenge for scientists because of involvement of multiple survival pathways of cancer cells. An extensive study on the regulation of each step of these pathways may help find a potential cancer target. Up-regulated HIF-1 expression and altered metabolic pathways are two classical characteristics of cancer. Oxygen-dependent (through pVHL, PHDs, calcium-mediated) and independent (through growth factor signaling pathway, mdm2 pathway, HSP90) regulation of HIF-1α leads to angiogenesis, metastasis, and cell survival. The two subunits of HIF-1 regulates in the same fashion through different mechanisms. HIF-1α translation upregulates via mammalian target of rapamycin and mitogen-activated protein kinase signaling pathways, whereas HIF-1β through calmodulin kinase. Further, the stabilized interactions of these two subunits are important for proper functioning. Also, metabolic pathways crucial for the formation of building blocks (pentose phosphate pathway) and energy generation (glycolysis, TCA cycle and catabolism of glutamine) are altered in cancer cells to protect them from oxidative stress and to meet the reduced oxygen and nutrient supply. Up-regulated anaerobic metabolism occurs through enhanced expression of hexokinase, phosphofructokinase, triosephosphate isomerase, glucose 6-phosphate dehydrogenase and down-regulation of aerobic metabolism via pyruvate dehydrogenase kinase and lactate dehydrogenase which compensate energy requirements along with high glucose intake. Controlled expression of these two pathways through their common intermediate may serve as potent cancer target in future.

PAR1 participates in the ability of multidrug resistance and tumorigenesis by controlling Hippo-YAP pathway.

PubMed

Fujimoto, Daisuke; Ueda, Yuki; Hirono, Yasuo; Goi, Takanori; Yamaguchi, Akio

2015-10-27

The Hippo pathway significantly correlates with organ size control and tumorigenesis. The activity of YAP/TAZ, a transducer of the Hippo pathway, is required to sustain self-renewal and tumor-initiation capacities in cancer stem cells (CSCs). But, upstream signals that control the mammalian Hippo pathway have not been well understood. Here, we reveal a connection between the Protease-activated receptor 1 (PAR1) signaling pathway and the Hippo-YAP pathway in gastric cancer stem-like cells. The selective PAR1 agonist TFLLR-NH2 induces an increase in the fraction of side population cells which is enriched in CSCs, and promotes tumorigenesis, multi cancer drug resistance, cell morphological change, and cell invasion which are characteristics of CSCs. In addition, PAR1 activation inhibits the Hippo-YAP pathway kinase Lats via Rho GTPase. Lats kinase inhibition in turn results in increased nuclear localization of dephosphorylated YAP. Furthermore, PAR1 activation confers CSCs related traits via the Hippo-YAP pathway, and the Hippo-YAP pathway correlates with epithelial mesenchymal transition which is induced by PAR1 activation. Our research suggests that the PAR1 signaling deeply participates in the ability of multi drug resistance and tumorigenesis through interactions with the Hippo-YAP pathway signaling in gastric cancer stem-like cells. We presume that inhibited YAP is a new therapeutic target in the treatment human gastric cancer invasion and metastasis by dysregulated PAR1 or its agonists.

Inactivation of conserved genes induces microbial aversion, drug detoxification, and innate immunity in C.elegans

PubMed Central

Melo, Justine A.; Ruvkun, Gary

2012-01-01

Summary The nematode C. elegans consumes benign bacteria such as E. coli and is repelled by pathogens and toxins. Here we show that RNAi and toxin-mediated disruption of core cellular activities, including translation, respiration, and protein turnover, stimulates behavioral avoidance of attractive E. coli. RNAi of such essential processes also induces expression of detoxification and innate immune response genes in the absence of toxins or pathogens. Disruption of core processes in non-neuronal tissues can stimulate aversion behavior, revealing a neuroendocrine axis of control. Microbial avoidance requires serotonergic and Jnk kinase signaling. We propose that surveillance pathways oversee critical cellular activities to detect pathogens, many of which deploy toxins and virulence factors to disrupt these same host pathways. Variation in cellular surveillance and endocrine pathways controlling behavior, detoxification and immunity selected by past toxin or microbial interactions could underlie aberrant responses to foods, medicines, and microbes. PMID:22500807

LENS: web-based lens for enrichment and network studies of human proteins

PubMed Central

2015-01-01

Background Network analysis is a common approach for the study of genetic view of diseases and biological pathways. Typically, when a set of genes are identified to be of interest in relation to a disease, say through a genome wide association study (GWAS) or a different gene expression study, these genes are typically analyzed in the context of their protein-protein interaction (PPI) networks. Further analysis is carried out to compute the enrichment of known pathways and disease-associations in the network. Having tools for such analysis at the fingertips of biologists without the requirement for computer programming or curation of data would accelerate the characterization of genes of interest. Currently available tools do not integrate network and enrichment analysis and their visualizations, and most of them present results in formats not most conducive to human cognition. Results We developed the tool Lens for Enrichment and Network Studies of human proteins (LENS) that performs network and pathway and diseases enrichment analyses on genes of interest to users. The tool creates a visualization of the network, provides easy to read statistics on network connectivity, and displays Venn diagrams with statistical significance values of the network's association with drugs, diseases, pathways, and GWASs. We used the tool to analyze gene sets related to craniofacial development, autism, and schizophrenia. Conclusion LENS is a web-based tool that does not require and download or plugins to use. The tool is free and does not require login for use, and is available at http://severus.dbmi.pitt.edu/LENS. PMID:26680011

Electron tunneling through covalent and noncovalent pathways in proteins

NASA Technical Reports Server (NTRS)

Beratan, David N.; Onuchic, Jose Nelson; Hopfield, J. J.

1987-01-01

A model is presented for electron tunneling in proteins which allows the donor-acceptor interaction to be mediated by the covalent bonds between amino acids and noncovalent contacts between amino acid chains. The important tunneling pathways are predicted to include mostly bonded groups with less favorable nonbonded interactions being important when the through bond pathway is prohibitively long. In some cases, vibrational motion of nonbonded groups along the tunneling pathway strongly influences the temperature dependence of the rate. Quantitative estimates for the sizes of these noncovalent interactions are made and their role in protein mediated electron transport is discussed.

Factor X/Xa elicits protective signaling responses in endothelial cells directly via PAR-2 and indirectly via endothelial protein C receptor-dependent recruitment of PAR-1.

PubMed

Bae, Jong-Sup; Yang, Likui; Rezaie, Alireza R

2010-11-05

We recently demonstrated that the Gla domain-dependent interaction of protein C with endothelial protein C receptor (EPCR) leads to dissociation of the receptor from caveolin-1 and recruitment of PAR-1 to a protective signaling pathway. Thus, the activation of PAR-1 by either thrombin or PAR-1 agonist peptide elicited a barrier-protective response if endothelial cells were preincubated with protein C. In this study, we examined whether other vitamin K-dependent coagulation protease zymogens can modulate PAR-dependent signaling responses in endothelial cells. We discovered that the activation of both PAR-1 and PAR-2 in endothelial cells pretreated with factor FX (FX)-S195A, but not other procoagulant protease zymogens, also results in initiation of protective intracellular responses. Interestingly, similar to protein C, FX interaction with endothelial cells leads to dissociation of EPCR from caveolin-1 and recruitment of PAR-1 to a protective pathway. Further studies revealed that, FX activated by factor VIIa on tissue factor bearing endothelial cells also initiates protective signaling responses through the activation of PAR-2 independent of EPCR mobilization. All results could be recapitulated by the receptor agonist peptides to both PAR-1 and PAR-2. These results suggest that a cross-talk between EPCR and an unknown FX/FXa receptor, which does not require interaction with the Gla domain of FX, recruits PAR-1 to protective signaling pathways in endothelial cells.

Transition model for ricin-aptamer interactions with multiple pathways and energy barriers

NASA Astrophysics Data System (ADS)

Wang, Bin; Xu, Bingqian

2014-02-01

We develop a transition model to interpret single-molecule ricin-aptamer interactions with multiple unbinding pathways and energy barriers measured by atomic force microscopy dynamic force spectroscopy. Molecular simulations establish the relationship between binding conformations and the corresponding unbinding pathways. Each unbinding pathway follows a Bell-Evans multiple-barrier model. Markov-type transition matrices are developed to analyze the redistribution of unbinding events among the pathways under different loading rates. Our study provides detailed information about complex behaviors in ricin-aptamer unbinding events.

SLY1 and Syntaxin 18 specify a distinct pathway for procollagen VII export from the endoplasmic reticulum

PubMed Central

Nogueira, Cristina; Erlmann, Patrik; Villeneuve, Julien; Santos, António JM; Martínez-Alonso, Emma; Martínez-Menárguez, José Ángel; Malhotra, Vivek

2014-01-01

TANGO1 binds and exports Procollagen VII from the endoplasmic reticulum (ER). In this study, we report a connection between the cytoplasmic domain of TANGO1 and SLY1, a protein that is required for membrane fusion. Knockdown of SLY1 by siRNA arrested Procollagen VII in the ER without affecting the recruitment of COPII components, general protein secretion, and retrograde transport of the KDEL-containing protein BIP, and ERGIC53. SLY1 is known to interact with the ER-specific SNARE proteins Syntaxin 17 and 18, however only Syntaxin 18 was required for Procollagen VII export. Neither SLY1 nor Syntaxin 18 was required for the export of the equally bulky Procollagen I from the ER. Altogether, these findings reveal the sorting of bulky collagen family members by TANGO1 at the ER and highlight the existence of different export pathways for secretory cargoes one of which is mediated by the specific SNARE complex containing SLY1 and Syntaxin 18. DOI: http://dx.doi.org/10.7554/eLife.02784.001 PMID:24842878

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.

The Nkx5/HMX homeodomain protein MLS-2 is required for proper tube cell shape in the C.elegans excretory system

PubMed Central

Abdus-Saboor, Ishmail; Stone, Craig E.; Murray, John I.; Sundaram, Meera V.

2012-01-01

Cells perform wide varieties of functions that are facilitated, in part, by adopting unique shapes. Many of the genes and pathways that promote cell fate specification have been elucidated. However, relatively few transcription factors have been identified that promote shape acquisition after fate specification. Here we show that the Nkx5/HMX homeodomain protein MLS-2 is required for cellular elongation and shape maintenance of two tubular epithelial cells in the C.elegans excretory system, the duct and pore cells. The Nkx5/HMX family is highly conserved from sea urchins to humans, with known roles in neuronal and glial development. MLS-2 is expressed in the duct and pore, and defects in mls-2 mutants first arise when the duct and pore normally adopt unique shapes. MLS-2 cooperates with the EGF-Ras-ERK pathway to turn on the LIN-48/Ovo transcription factor in the duct cell during morphogenesis. These results reveal a novel interaction between the Nkx5/HMX family and the EGF-Ras pathway and implicate a transcription factor, MLS-2, as a regulator of cell shape. PMID:22537498

The Nkx5/HMX homeodomain protein MLS-2 is required for proper tube cell shape in the C. elegans excretory system.

PubMed

Abdus-Saboor, Ishmail; Stone, Craig E; Murray, John I; Sundaram, Meera V

2012-06-15

Cells perform wide varieties of functions that are facilitated, in part, by adopting unique shapes. Many of the genes and pathways that promote cell fate specification have been elucidated. However, relatively few transcription factors have been identified that promote shape acquisition after fate specification. Here we show that the Nkx5/HMX homeodomain protein MLS-2 is required for cellular elongation and shape maintenance of two tubular epithelial cells in the C. elegans excretory system, the duct and pore cells. The Nkx5/HMX family is highly conserved from sea urchins to humans, with known roles in neuronal and glial development. MLS-2 is expressed in the duct and pore, and defects in mls-2 mutants first arise when the duct and pore normally adopt unique shapes. MLS-2 cooperates with the EGF-Ras-ERK pathway to turn on the LIN-48/Ovo transcription factor in the duct cell during morphogenesis. These results reveal a novel interaction between the Nkx5/HMX family and the EGF-Ras pathway and implicate a transcription factor, MLS-2, as a regulator of cell shape. Copyright © 2012 Elsevier Inc. All rights reserved.

Investigating ego modules and pathways in osteosarcoma by integrating the EgoNet algorithm and pathway analysis.

PubMed

Chen, X Y; Chen, Y H; Zhang, L J; Wang, Y; Tong, Z C

2017-02-16

Osteosarcoma (OS) is the most common primary bone malignancy, but current therapies are far from effective for all patients. A better understanding of the pathological mechanism of OS may help to achieve new treatments for this tumor. Hence, the objective of this study was to investigate ego modules and pathways in OS utilizing EgoNet algorithm and pathway-related analysis, and reveal pathological mechanisms underlying OS. The EgoNet algorithm comprises four steps: constructing background protein-protein interaction (PPI) network (PPIN) based on gene expression data and PPI data; extracting differential expression network (DEN) from the background PPIN; identifying ego genes according to topological features of genes in reweighted DEN; and collecting ego modules using module search by ego gene expansion. Consequently, we obtained 5 ego modules (Modules 2, 3, 4, 5, and 6) in total. After applying the permutation test, all presented statistical significance between OS and normal controls. Finally, pathway enrichment analysis combined with Reactome pathway database was performed to investigate pathways, and Fisher's exact test was conducted to capture ego pathways for OS. The ego pathway for Module 2 was CLEC7A/inflammasome pathway, while for Module 3 a tetrasaccharide linker sequence was required for glycosaminoglycan (GAG) synthesis, and for Module 6 was the Rho GTPase cycle. Interestingly, genes in Modules 4 and 5 were enriched in the same pathway, the 2-LTR circle formation. In conclusion, the ego modules and pathways might be potential biomarkers for OS therapeutic index, and give great insight of the molecular mechanism underlying this tumor.

Investigating ego modules and pathways in osteosarcoma by integrating the EgoNet algorithm and pathway analysis

PubMed Central

Chen, X.Y.; Chen, Y.H.; Zhang, L.J.; Wang, Y.; Tong, Z.C.

2017-01-01

Osteosarcoma (OS) is the most common primary bone malignancy, but current therapies are far from effective for all patients. A better understanding of the pathological mechanism of OS may help to achieve new treatments for this tumor. Hence, the objective of this study was to investigate ego modules and pathways in OS utilizing EgoNet algorithm and pathway-related analysis, and reveal pathological mechanisms underlying OS. The EgoNet algorithm comprises four steps: constructing background protein-protein interaction (PPI) network (PPIN) based on gene expression data and PPI data; extracting differential expression network (DEN) from the background PPIN; identifying ego genes according to topological features of genes in reweighted DEN; and collecting ego modules using module search by ego gene expansion. Consequently, we obtained 5 ego modules (Modules 2, 3, 4, 5, and 6) in total. After applying the permutation test, all presented statistical significance between OS and normal controls. Finally, pathway enrichment analysis combined with Reactome pathway database was performed to investigate pathways, and Fisher's exact test was conducted to capture ego pathways for OS. The ego pathway for Module 2 was CLEC7A/inflammasome pathway, while for Module 3 a tetrasaccharide linker sequence was required for glycosaminoglycan (GAG) synthesis, and for Module 6 was the Rho GTPase cycle. Interestingly, genes in Modules 4 and 5 were enriched in the same pathway, the 2-LTR circle formation. In conclusion, the ego modules and pathways might be potential biomarkers for OS therapeutic index, and give great insight of the molecular mechanism underlying this tumor. PMID:28225867

A CAF-1–PCNA-Mediated Chromatin Assembly Pathway Triggered by Sensing DNA Damage

PubMed Central

Moggs, Jonathan G.; Grandi, Paola; Quivy, Jean-Pierre; Jónsson, Zophonías O.; Hübscher, Ulrich; Becker, Peter B.; Almouzni, Geneviève

2000-01-01

Sensing DNA damage is crucial for the maintenance of genomic integrity and cell cycle progression. The participation of chromatin in these events is becoming of increasing interest. We show that the presence of single-strand breaks and gaps, formed either directly or during DNA damage processing, can trigger the propagation of nucleosomal arrays. This nucleosome assembly pathway involves the histone chaperone chromatin assembly factor 1 (CAF-1). The largest subunit (p150) of this factor interacts directly with proliferating cell nuclear antigen (PCNA), and critical regions for this interaction on both proteins have been mapped. To isolate proteins specifically recruited during DNA repair, damaged DNA linked to magnetic beads was used. The binding of both PCNA and CAF-1 to this damaged DNA was dependent on the number of DNA lesions and required ATP. Chromatin assembly linked to the repair of single-strand breaks was disrupted by depletion of PCNA from a cell-free system. This defect was rescued by complementation with recombinant PCNA, arguing for role of PCNA in mediating chromatin assembly linked to DNA repair. We discuss the importance of the PCNA–CAF-1 interaction in the context of DNA damage processing and checkpoint control. PMID:10648606

A novel small molecule methyltransferase is important for virulence in Candida albicans.

PubMed

Lissina, Elena; Weiss, David; Young, Brian; Rella, Antonella; Cheung-Ong, Kahlin; Del Poeta, Maurizio; Clarke, Steven G; Giaever, Guri; Nislow, Corey

2013-12-20

Candida albicans is an opportunistic pathogen capable of causing life-threatening infections in immunocompromised individuals. Despite its significant health impact, our understanding of C. albicans pathogenicity is limited, particularly at the molecular level. One of the largely understudied enzyme families in C. albicans are small molecule AdoMet-dependent methyltransferases (smMTases), which are important for maintenance of cellular homeostasis by clearing toxic chemicals, generating novel cellular intermediates, and regulating intra- and interspecies interactions. In this study, we demonstrated that C. albicans Crg1 (CaCrg1) is a bona fide smMTase that interacts with the toxin in vitro and in vivo. We report that CaCrg1 is important for virulence-related processes such as adhesion, hyphal elongation, and membrane trafficking. Biochemical and genetic analyses showed that CaCrg1 plays a role in the complex sphingolipid pathway: it binds to exogenous short-chain ceramides in vitro and interacts genetically with genes of glucosylceramide pathway, and the deletion of CaCRG1 leads to significant changes in the abundance of phytoceramides. Finally we found that this novel lipid-related smMTase is required for virulence in the waxmoth Galleria mellonella, a model of infection.

Posttranslational Modifications Regulate the Postsynaptic Localization of PSD-95.

PubMed

Vallejo, Daniela; Codocedo, Juan F; Inestrosa, Nibaldo C

2017-04-01

The postsynaptic density (PSD) consists of a lattice-like array of interacting proteins that organizes and stabilizes synaptic receptors, ion channels, structural proteins, and signaling molecules required for normal synaptic transmission and synaptic function. The scaffolding and hub protein postsynaptic density protein-95 (PSD-95) is a major element of central chemical synapses and interacts with glutamate receptors, cell adhesion molecules, and cytoskeletal elements. In fact, PSD-95 can regulate basal synaptic stability as well as the activity-dependent structural plasticity of the PSD and, therefore, of the excitatory chemical synapse. Several studies have shown that PSD-95 is highly enriched at excitatory synapses and have identified multiple protein structural domains and protein-protein interactions that mediate PSD-95 function and trafficking to the postsynaptic region. PSD-95 is also a target of several signaling pathways that induce posttranslational modifications, including palmitoylation, phosphorylation, ubiquitination, nitrosylation, and neddylation; these modifications determine the synaptic stability and function of PSD-95 and thus regulate the fates of individual dendritic spines in the nervous system. In the present work, we review the posttranslational modifications that regulate the synaptic localization of PSD-95 and describe their functional consequences. We also explore the signaling pathways that induce such changes.

Evaluation of potential endocrine activity of 2,4-dichlorophenoxyacetic acid using in vitro assays.

PubMed

Coady, Katherine K; Kan, H Lynn; Schisler, Melissa R; Gollapudi, B Bhaskar; Neal, Barbara; Williams, Amy; LeBaron, Matthew J

2014-08-01

The herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) was evaluated in five in vitro screening assays to assess the potential for interaction with the androgen, estrogen and steroidogenesis pathways in the endocrine system. The assays were conducted to meet the requirements of the in vitro component of Tier 1 of the United States Environmental Protection Agency's Endocrine Disruptor Screening Program (EDSP), and included assays for estrogen receptor (ER) binding (rat uterine cytosol ER binding assay), ER-mediated transcriptional activation (HeLa-9903-ERα transactivation assay), androgen receptor (AR) binding (rat prostate cytosol AR binding assay), aromatase enzymatic activity inhibition (recombinant human CYP19 aromatase inhibition assay), and interference with steroidogenesis (H295R steroidogenesis assay). Results from these five assays demonstrated that 2,4-D does not have the potential to interact in vitro with the estrogen, androgen, or steroidogenesis pathways. These in vitro data are consistent with a corresponding lack of endocrine effects observed in apical in vivo animal studies, and thus provide important supporting data valuable in a comprehensive weight of evidence evaluation indicating a low potential of 2,4-D to interact with the endocrine system. Copyright © 2014 Elsevier Ltd. All rights reserved.

Phosphorylation of PPP(S/T)P motif of the free LRP6 intracellular domain is not required to activate the Wnt/beta-catenin pathway and attenuate GSK3beta activity.

PubMed

Beagle, Brandon; Mi, Kaihong; Johnson, Gail V W

2009-11-01

The canonical Wnt/beta-catenin signaling pathway plays a critical role in numerous physiological and pathological processes. LRP6 is an essential co-receptor for Wnt/beta-catenin signaling; as transduction of the Wnt signal is strongly dependent upon GSK3beta-mediated phosphorylation of multiple PPP(S/T)P motifs within the membrane-anchored LRP6 intracellular domain. Previously, we showed that the free LRP6 intracellular domain (LRP6-ICD) can activate the Wnt/beta-catenin pathway in a beta-catenin and TCF/LEF-1 dependent manner, as well as interact with and attenuate GSK3beta activity. However, it is unknown if the ability of LRP6-ICD to attenuate GSK3beta activity and modulate activation of the Wnt/beta-catenin pathway requires phosphorylation of the LRP6-ICD PPP(S/T)P motifs, in a manner similar to the membrane-anchored LRP6 intracellular domain. Here we provide evidence that the LRP6-ICD does not have to be phosphorylated at its PPP(S/T)P motif by GSK3beta to stabilize endogenous cytosolic beta-catenin resulting in activation of TCF/LEF-1 and the Wnt/beta-catenin pathway. LRP6-ICD and a mutant in which all 5 PPP(S/T)P motifs were changed to PPP(A)P motifs equivalently interacted with and attenuated GSK3beta activity in vitro, and both constructs inhibited the in situ GSK3beta-mediated phosphorylation of beta-catenin and tau to the same extent. These data indicate that the LRP6-ICD attenuates GSK3beta activity similar to other GSK3beta binding proteins, and is not a result of it being a GSK3beta substrate. Our findings suggest the functional and regulatory mechanisms governing the free LRP6-ICD may be distinct from membrane-anchored LRP6, and that release of the LRP6-ICD may provide a complimentary signaling cascade capable of modulating Wnt-dependent gene expression. (c) 2009 Wiley-Liss, Inc.

Spatially restricted G protein-coupled receptor activity via divergent endocytic compartments.

PubMed

Jean-Alphonse, Frederic; Bowersox, Shanna; Chen, Stanford; Beard, Gemma; Puthenveedu, Manojkumar A; Hanyaloglu, Aylin C

2014-02-14

Postendocytic sorting of G protein-coupled receptors (GPCRs) is driven by their interactions between highly diverse receptor sequence motifs with their interacting proteins, such as postsynaptic density protein (PSD95), Drosophila disc large tumor suppressor (Dlg1), zonula occludens-1 protein (zo-1) (PDZ) domain proteins. However, whether these diverse interactions provide an underlying functional specificity, in addition to driving sorting, is unknown. Here we identify GPCRs that recycle via distinct PDZ ligand/PDZ protein pairs that exploit their recycling machinery primarily for targeted endosomal localization and signaling specificity. The luteinizing hormone receptor (LHR) and β2-adrenergic receptor (B2AR), two GPCRs sorted to the regulated recycling pathway, underwent divergent trafficking to distinct endosomal compartments. Unlike B2AR, which traffics to early endosomes (EE), LHR internalizes to distinct pre-early endosomes (pre-EEs) for its recycling. Pre-EE localization required interactions of the LHR C-terminal tail with the PDZ protein GAIP-interacting protein C terminus, inhibiting its traffic to EEs. Rerouting the LHR to EEs, or EE-localized GPCRs to pre-EEs, spatially reprograms MAPK signaling. Furthermore, LHR-mediated activation of MAPK signaling requires internalization and is maintained upon loss of the EE compartment. We propose that combinatorial specificity between GPCR sorting sequences and interacting proteins dictates an unprecedented spatiotemporal control in GPCR signal activity.

Integration of growth factor signals at the c-fos serum response element.

PubMed

Price, M A; Hill, C; Treisman, R

1996-04-29

A transcription factor ternary complex composed of serum response factor (SRF) and a second factor, ternary complex factor (TCF), mediates the response of the c-fos Serum Response Element to growth factors and mitogens. In NIH3T3 fibroblasts, TCF binding is required for transcriptional activation by the SRE in response to activation of the Ras-Raf-ERK pathway. We compared the properties of three members of the TCF family, Elk-1, SAP-1 and SAP-2 (ERP/NET). Although all the proteins contain sequences required for ternary complex formation with SRF, only Elk-1 and SAP-1 appear to interact with the c-fos SRE efficiently in vivo. Each TCF contains a C-terminal activation domain capable of transcriptional activation in response to activation of the Ras-Raf-ERK pathway, and this is dependent on the integrity of S/T-P motifs conserved between all the TCF family members. In contrast, activation of the SRE by whole serum and the mitogenic phospholipid LPA requires SRF binding alone. Constitutively activated members of the Rho subfamily of Ras-like GTPases are also capable of inducing activation of the SRE in the absence of TCF; unlike activated Ras itself, these proteins do not activate the TCFs in NIH3T3 cells. At the SRE, SRF- and TCF-linked signalling pathways act synergistically to potentiate transcription.

The shortest path is not the one you know: application of biological network resources in precision oncology research.

PubMed

Kuperstein, Inna; Grieco, Luca; Cohen, David P A; Thieffry, Denis; Zinovyev, Andrei; Barillot, Emmanuel

2015-03-01

Several decades of molecular biology research have delivered a wealth of detailed descriptions of molecular interactions in normal and tumour cells. This knowledge has been functionally organised and assembled into dedicated biological pathway resources that serve as an invaluable tool, not only for structuring the information about molecular interactions but also for making it available for biological, clinical and computational studies. With the advent of high-throughput molecular profiling of tumours, close to complete molecular catalogues of mutations, gene expression and epigenetic modifications are available and require adequate interpretation. Taking into account the information about biological signalling machinery in cells may help to better interpret molecular profiles of tumours. Making sense out of these descriptions requires biological pathway resources for functional interpretation of the data. In this review, we describe the available biological pathway resources, their characteristics in terms of construction mode, focus, aims and paradigms of biological knowledge representation. We present a new resource that is focused on cancer-related signalling, the Atlas of Cancer Signalling Networks. We briefly discuss current approaches for data integration, visualisation and analysis, using biological networks, such as pathway scoring, guilt-by-association and network propagation. Finally, we illustrate with several examples the added value of data interpretation in the context of biological networks and demonstrate that it may help in analysis of high-throughput data like mutation, gene expression or small interfering RNA screening and can guide in patients stratification. Finally, we discuss perspectives for improving precision medicine using biological network resources and tools. Taking into account the information about biological signalling machinery in cells may help to better interpret molecular patterns of tumours and enable to put precision oncology into general clinical practice. © The Author 2015. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Combination therapy in a xenograft model of glioblastoma: enhancement of the antitumor activity of temozolomide by an MDM2 antagonist.

PubMed

Wang, Haiyan; Cai, Shanbao; Bailey, Barbara J; Reza Saadatzadeh, M; Ding, Jixin; Tonsing-Carter, Eva; Georgiadis, Taxiarchis M; Zachary Gunter, T; Long, Eric C; Minto, Robert E; Gordon, Kevin R; Sen, Stephanie E; Cai, Wenjing; Eitel, Jacob A; Waning, David L; Bringman, Lauren R; Wells, Clark D; Murray, Mary E; Sarkaria, Jann N; Gelbert, Lawrence M; Jones, David R; Cohen-Gadol, Aaron A; Mayo, Lindsey D; Shannon, Harlan E; Pollok, Karen E

2017-02-01

OBJECTIVE Improvement in treatment outcome for patients with glioblastoma multiforme (GBM) requires a multifaceted approach due to dysregulation of numerous signaling pathways. The murine double minute 2 (MDM2) protein may fulfill this requirement because it is involved in the regulation of growth, survival, and invasion. The objective of this study was to investigate the impact of modulating MDM2 function in combination with front-line temozolomide (TMZ) therapy in GBM. METHODS The combination of TMZ with the MDM2 protein-protein interaction inhibitor nutlin3a was evaluated for effects on cell growth, p53 pathway activation, expression of DNA repair proteins, and invasive properties. In vivo efficacy was assessed in xenograft models of human GBM. RESULTS In combination, TMZ/nutlin3a was additive to synergistic in decreasing growth of wild-type p53 GBM cells. Pharmacodynamic studies demonstrated that inhibition of cell growth following exposure to TMZ/nutlin3a correlated with: 1) activation of the p53 pathway, 2) downregulation of DNA repair proteins, 3) persistence of DNA damage, and 4) decreased invasion. Pharmacokinetic studies indicated that nutlin3a was detected in human intracranial tumor xenografts. To assess therapeutic potential, efficacy studies were conducted in a xenograft model of intracranial GBM by using GBM cells derived from a recurrent wild-type p53 GBM that is highly TMZ resistant (GBM10). Three 5-day cycles of TMZ/nutlin3a resulted in a significant increase in the survival of mice with GBM10 intracranial tumors compared with single-agent therapy. CONCLUSIONS Modulation of MDM2/p53-associated signaling pathways is a novel approach for decreasing TMZ resistance in GBM. To the authors' knowledge, this is the first study in a humanized intracranial patient-derived xenograft model to demonstrate the efficacy of combining front-line TMZ therapy and an inhibitor of MDM2 protein-protein interactions.

Are common symbiosis genes required for endophytic rice-rhizobial interactions?

PubMed

Chen, Caiyan; Zhu, Hongyan

2013-09-01

Legume plants are able to establish root nodule symbioses with nitrogen-fixing bacteria, called rhizobia. Recent studies revealed that the root nodule symbiosis has co-opted the signaling pathway that mediates the ancestral mycorrhizal symbiosis that occurs in most land plants. Despite being unable to induce nodulation, rhizobia have been shown to be able to infect and colonize the roots of non-legumes such as rice. One fascinating question is whether establishment of such associations requires the common symbiosis (Sym) genes that are essential for infection of plant cells by mycorrhizal fungi and rhizobia in legumes. Here, we demonstrated that the common Sym genes are not required for endophytic colonization of rice roots by nitrogen-fixing rhizobia.

The visual orientation memory of Drosophila requires Foraging (PKG) upstream of Ignorant (RSK2) in ring neurons of the central complex

PubMed Central

Kuntz, Sara; Poeck, Burkhard; Sokolowski, Marla B.; Strauss, Roland

2012-01-01

Orientation and navigation in a complex environment requires path planning and recall to exert goal-driven behavior. Walking Drosophila flies possess a visual orientation memory for attractive targets which is localized in the central complex of the adult brain. Here we show that this type of working memory requires the cGMP-dependent protein kinase encoded by the foraging gene in just one type of ellipsoid-body ring neurons. Moreover, genetic and epistatic interaction studies provide evidence that Foraging functions upstream of the Ignorant Ribosomal-S6 Kinase 2, thus revealing a novel neuronal signaling pathway necessary for this type of memory in Drosophila. PMID:22815538

A network model of genomic hormone interactions underlying dementia and its translational validation through serendipitous off-target effect

PubMed Central

2013-01-01

Background While the majority of studies have focused on the association between sex hormones and dementia, emerging evidence supports the role of other hormone signals in increasing dementia risk. However, due to the lack of an integrated view on mechanistic interactions of hormone signaling pathways associated with dementia, molecular mechanisms through which hormones contribute to the increased risk of dementia has remained unclear and capacity of translating hormone signals to potential therapeutic and diagnostic applications in relation to dementia has been undervalued. Methods Using an integrative knowledge- and data-driven approach, a global hormone interaction network in the context of dementia was constructed, which was further filtered down to a model of convergent hormone signaling pathways. This model was evaluated for its biological and clinical relevance through pathway recovery test, evidence-based analysis, and biomarker-guided analysis. Translational validation of the model was performed using the proposed novel mechanism discovery approach based on ‘serendipitous off-target effects’. Results Our results reveal the existence of a well-connected hormone interaction network underlying dementia. Seven hormone signaling pathways converge at the core of the hormone interaction network, which are shown to be mechanistically linked to the risk of dementia. Amongst these pathways, estrogen signaling pathway takes the major part in the model and insulin signaling pathway is analyzed for its association to learning and memory functions. Validation of the model through serendipitous off-target effects suggests that hormone signaling pathways substantially contribute to the pathogenesis of dementia. Conclusions The integrated network model of hormone interactions underlying dementia may serve as an initial translational platform for identifying potential therapeutic targets and candidate biomarkers for dementia-spectrum disorders such as Alzheimer’s disease. PMID:23885764

Metabolic pathways of Pseudomonas aeruginosa involved in competition with respiratory bacterial pathogens

PubMed Central

Beaume, Marie; Köhler, Thilo; Fontana, Thierry; Tognon, Mikael; Renzoni, Adriana; van Delden, Christian

2015-01-01

Background: Chronic airway infection by Pseudomonas aeruginosa considerably contributes to lung tissue destruction and impairment of pulmonary function in cystic-fibrosis (CF) patients. Complex interplays between P. aeruginosa and other co-colonizing pathogens including Staphylococcus aureus, Burkholderia sp., and Klebsiella pneumoniae may be crucial for pathogenesis and disease progression. Methods: We generated a library of PA14 transposon insertion mutants to identify P. aeruginosa genes required for exploitative and direct competitions with S. aureus, Burkholderia cenocepacia, and K. pneumoniae. Results: Whereas wild-type PA14 inhibited S. aureus growth, two transposon insertions located in pqsC and carB, resulted in reduced growth inhibition. PqsC is involved in the synthesis of 4-hydroxy-2-alkylquinolines (HAQs), a family of molecules having antibacterial properties, while carB is a key gene in pyrimidine biosynthesis. The carB mutant was also unable to grow in the presence of B. cepacia and K. pneumoniae but not Escherichia coli and S. epidermidis. We further identified a transposon insertion in purF, encoding a key enzyme of purine metabolism. This mutant displayed a severe growth deficiency in the presence of Gram-negative but not of Gram-positive bacteria. We identified a beneficial interaction in a bioA transposon mutant, unable to grow on rich medium. This growth defect could be restored either by addition of biotin or by co-culturing the mutant in the presence of K. pneumoniae or E. coli. Conclusion: Complex interactions take place between the various bacterial species colonizing CF-lungs. This work identified both detrimental and beneficial interactions occurring between P. aeruginosa and three other respiratory pathogens involving several major metabolic pathways. Manipulating these pathways could be used to interfere with bacterial interactions and influence the colonization by respiratory pathogens. PMID:25954256

SUMOylation of ATRIP potentiates DNA damage signaling by boosting multiple protein interactions in the ATR pathway

PubMed Central

Wu, Ching-Shyi; Ouyang, Jian; Mori, Eiichiro; Nguyen, Hai Dang; Maréchal, Alexandre; Hallet, Alexander; Chen, David J.; Zou, Lee

2014-01-01

The ATR (ATM [ataxia telangiectasia-mutated]- and Rad3-related) checkpoint is a crucial DNA damage signaling pathway. While the ATR pathway is known to transmit DNA damage signals through the ATR–Chk1 kinase cascade, whether post-translational modifications other than phosphorylation are important for this pathway remains largely unknown. Here, we show that protein SUMOylation plays a key role in the ATR pathway. ATRIP, the regulatory partner of ATR, is modified by SUMO2/3 at K234 and K289. An ATRIP mutant lacking the SUMOylation sites fails to localize to DNA damage and support ATR activation efficiently. Surprisingly, the ATRIP SUMOylation mutant is compromised in the interaction with a protein group, rather than a single protein, in the ATR pathway. Multiple ATRIP-interacting proteins, including ATR, RPA70, TopBP1, and the MRE11–RAD50–NBS1 complex, exhibit reduced binding to the ATRIP SUMOylation mutant in cells and display affinity for SUMO2 chains in vitro, suggesting that they bind not only ATRIP but also SUMO. Fusion of a SUMO2 chain to the ATRIP SUMOylation mutant enhances its interaction with the protein group and partially suppresses its localization and functional defects, revealing that ATRIP SUMOylation promotes ATR activation by providing a unique type of protein glue that boosts multiple protein interactions along the ATR pathway. PMID:24990965

Choline Metabolites: Gene by Diet Interactions

PubMed Central

Smallwood, Tangi; Allayee, Hooman; Bennett, Brian J.

2015-01-01

Purpose of review This review highlights recent advances in our understanding of the interactions between genetic polymorphisms in genes that metabolize choline and the dietary requirements of choline and how these interactions relate to human health and disease. Recent findings The importance of choline as an essential nutrient has been well established but our appreciation of the interaction between our underlying genetic architecture and dietary choline requirements is only beginning. It has been shown in both human and animal studies that choline deficiencies contribute to diseases such as non-alcoholic fatty liver disease and various neurodegenerative diseases. An adequate supply of dietary choline is important for optimum development, highlighted by the increased maternal requirements during fetal development and in breast-fed infants. We discuss recent studies investigating variants in PEMT and MTHFR1 that are associated with a variety of birth defects. In addition to genetic interactions, we discuss several recent studies that uncover changes in fetal global methylation patterns in response to maternal dietary choline intake that result in changes in gene expression in the offspring. In contrast to the developmental role of adequate choline, there is now an appreciation of the role choline has in cardiovascular disease through the gut microbiota-mediated metabolite trimethylamine N-oxide. This pathway highlights some of our understanding of how the microbiome affects nutrient processing and bioavailability. Finally, in order to better characterize the genetic architecture regulating choline requirements, we discuss recent results focused on identifying polymorphisms that regulate choline and its derivative products. Summary Here we discuss recent studies that have advanced our understanding of how specific alleles in key choline metabolism genes are related to dietary choline requirements and human disease. PMID:26655287

Biogenesis pathways of piRNAs loaded onto AGO3 in the Drosophila testis.

PubMed

Nagao, Akihiro; Mituyama, Toutai; Huang, Haidong; Chen, Dahua; Siomi, Mikiko C; Siomi, Haruhiko

2010-12-01

PIWI-interacting RNAs (piRNAs) silence transposable elements in animal germ cells. In Drosophila ovaries, piRNAs are produced by two distinct pathways: the "ping-pong" amplification cycle that operates in germ cells and a ping-pong-independent pathway termed the primary pathway that mainly operates in somatic cells. AGO3, one of three PIWI proteins in flies, is involved in the ping-pong cycle in ovaries. We characterized AGO3-associated piRNAs in fly testes and found that like in ovaries, AGO3 functions in the ping-pong cycle with Aubergine (Aub) for piRNA production from transposon transcripts. In contrast, most AGO3-associated piRNAs corresponding to Suppressor of Stellate [Su(Ste)] genes are antisense-oriented and bound to Aub. In addition, the vast majority of AGO3-bound piRNAs derived from the AT-chX locus on chromosome X are antisense-oriented and are also found among Aub-associated piRNAs. The presence of very few sense Su(Ste) and AT-chX piRNAs suggests that biogenesis of both Su(Ste) and AT-chX piRNAs by a ping-pong mechanism only is highly unlikely. Nevertheless, the mutual interdependence of AGO3 and Aub for the accumulation of these piRNAs shows that their production relies on both AGO3 and Aub. Analysis of piRNA pathway mutants revealed that although the requirements for piRNA factors for Su(Ste)- and AT-chX-piRNA levels mostly overlap and resemble those for the ping-pong mechanism in the ovaries, Armitage (armi) is not required for the accumulation of AT-chX-1 piRNA. These findings suggest that the impacts of armi mutants on the operation of the piRNA pathway are variable in germ cells of fly testes.

Cooperation between STAT5 and phosphatidylinositol 3-kinase in the IL-3-dependent survival of a bone marrow derived cell line.

PubMed

Rosa Santos, S C; Dumon, S; Mayeux, P; Gisselbrecht, S; Gouilleux, F

2000-02-24

Cytokine-dependent activation of distinct signaling pathways is a common scheme thought to be required for the subsequent programmation into cell proliferation and survival. The PI 3-kinase/Akt, Ras/MAP kinase, Ras/NFIL3 and JAK/STAT pathways have been shown to participate in cytokine mediated suppression of apoptosis in various cell types. However the relative importance of these signaling pathways seems to depend on the cellular context. In several cases, individual inhibition of each pathway is not sufficient to completely abrogate cytokine mediated cell survival suggesting that cooperation between these pathways is required. Here we showed that individual inhibition of STAT5, PI 3-kinase or MEK activities did not or weakly affected the IL-3 dependent survival of the bone marrow derived Ba/F3 cell line. However, the simultaneous inhibition of STAT5 and PI 3-kinase activities but not that of STAT5 and MEK reduced the IL-3 dependent survival of Ba/F3. Analysis of the expression of the Bcl-2 members indicated that phosphorylation of Bad and Bcl-x expression which are respectively regulated by the PI 3-kinase/Akt pathway and STAT5 probably explain this cooperation. Furthermore, we showed by co-immunoprecipitation studies and pull down experiments with fusion proteins encoding the GST-SH2 domains of p85 that STAT5 in its phosphorylated form interacts with the p85 subunit of the PI 3-kinase. These results indicate that the activations of STAT5 and the PI 3-kinase by IL-3 in Ba/F3 cells are tightly connected and cooperate to mediate IL-3-dependent suppression of apoptosis by modulating Bad phosphorylation and Bcl-x expression.

Functional reconstitution of mitochondrial Fe/S cluster synthesis on Isu1 reveals the involvement of ferredoxin.

PubMed

Webert, Holger; Freibert, Sven-Andreas; Gallo, Angelo; Heidenreich, Torsten; Linne, Uwe; Amlacher, Stefan; Hurt, Ed; Mühlenhoff, Ulrich; Banci, Lucia; Lill, Roland

2014-10-31

Maturation of iron-sulphur (Fe/S) proteins involves complex biosynthetic machinery. In vivo synthesis of [2Fe-2S] clusters on the mitochondrial scaffold protein Isu1 requires the cysteine desulphurase complex Nfs1-Isd11, frataxin, ferredoxin Yah1 and its reductase Arh1. The roles of Yah1-Arh1 have remained enigmatic, because they are not required for in vitro Fe/S cluster assembly. Here, we reconstitute [2Fe-2S] cluster synthesis on Isu1 in a reaction depending on Nfs1-Isd11, frataxin, Yah1, Arh1 and NADPH. Unlike in the bacterial system, frataxin is an essential part of Fe/S cluster biosynthesis and is required simultaneously and stoichiometrically to Yah1. Reduced but not oxidized Yah1 tightly interacts with apo-Isu1 indicating a dynamic interaction between Yah1-apo-Isu1. Nuclear magnetic resonance structural studies identify the Yah1-apo-Isu1 interaction surface and suggest a pathway for electron flow from reduced ferredoxin to Isu1. Together, our study defines the molecular function of the ferredoxin Yah1 and its human orthologue FDX2 in mitochondrial Fe/S cluster synthesis.

Short-Chain 3-Hydroxyacyl-Coenzyme A Dehydrogenase Associates with a Protein Super-Complex Integrating Multiple Metabolic Pathways

PubMed Central

Narayan, Srinivas B.; Master, Stephen R.; Sireci, Anthony N.; Bierl, Charlene; Stanley, Paige E.; Li, Changhong; Stanley, Charles A.; Bennett, Michael J.

2012-01-01

Proteins involved in mitochondrial metabolic pathways engage in functionally relevant multi-enzyme complexes. We previously described an interaction between short-chain 3-hydroxyacyl-coenzyme A dehydrogenase (SCHAD) and glutamate dehydrogenase (GDH) explaining the clinical phenotype of hyperinsulinism in SCHAD-deficient patients and adding SCHAD to the list of mitochondrial proteins capable of forming functional, multi-pathway complexes. In this work, we provide evidence of SCHAD's involvement in additional interactions forming tissue-specific metabolic super complexes involving both membrane-associated and matrix-dwelling enzymes and spanning multiple metabolic pathways. As an example, in murine liver, we find SCHAD interaction with aspartate transaminase (AST) and GDH from amino acid metabolic pathways, carbamoyl phosphate synthase I (CPS-1) from ureagenesis, other fatty acid oxidation and ketogenesis enzymes and fructose-bisphosphate aldolase, an extra-mitochondrial enzyme of the glycolytic pathway. Most of the interactions appear to be independent of SCHAD's role in the penultimate step of fatty acid oxidation suggesting an organizational, structural or non-enzymatic role for the SCHAD protein. PMID:22496890

Interactive Sonification Exploring Emergent Behavior Applying Models for Biological Information and Listening

PubMed Central

Choi, Insook

2018-01-01

Sonification is an open-ended design task to construct sound informing a listener of data. Understanding application context is critical for shaping design requirements for data translation into sound. Sonification requires methodology to maintain reproducibility when data sources exhibit non-linear properties of self-organization and emergent behavior. This research formalizes interactive sonification in an extensible model to support reproducibility when data exhibits emergent behavior. In the absence of sonification theory, extensibility demonstrates relevant methods across case studies. The interactive sonification framework foregrounds three factors: reproducible system implementation for generating sonification; interactive mechanisms enhancing a listener's multisensory observations; and reproducible data from models that characterize emergent behavior. Supramodal attention research suggests interactive exploration with auditory feedback can generate context for recognizing irregular patterns and transient dynamics. The sonification framework provides circular causality as a signal pathway for modeling a listener interacting with emergent behavior. The extensible sonification model adopts a data acquisition pathway to formalize functional symmetry across three subsystems: Experimental Data Source, Sound Generation, and Guided Exploration. To differentiate time criticality and dimensionality of emerging dynamics, tuning functions are applied between subsystems to maintain scale and symmetry of concurrent processes and temporal dynamics. Tuning functions accommodate sonification design strategies that yield order parameter values to render emerging patterns discoverable as well as rehearsable, to reproduce desired instances for clinical listeners. Case studies are implemented with two computational models, Chua's circuit and Swarm Chemistry social agent simulation, generating data in real-time that exhibits emergent behavior. Heuristic Listening is introduced as an informal model of a listener's clinical attention to data sonification through multisensory interaction in a context of structured inquiry. Three methods are introduced to assess the proposed sonification framework: Listening Scenario classification, data flow Attunement, and Sonification Design Patterns to classify sound control. Case study implementations are assessed against these methods comparing levels of abstraction between experimental data and sound generation. Outcomes demonstrate the framework performance as a reference model for representing experimental implementations, also for identifying common sonification structures having different experimental implementations, identifying common functions implemented in different subsystems, and comparing impact of affordances across multiple implementations of listening scenarios. PMID:29755311

Cooperative control between AtRGS1 and AtHXK1 in a WD40-repeat protein pathway in Arabidopsis thaliana

DOE PAGES

Huang, Jian -Ping; Tunc-Ozdemir, Meral; Chang, Ying; ...

2015-10-13

HEXOKINASE 1 (AtHXK1) and Regulator of G-protein Signaling 1 (AtRGS1) pathways, mediate D-glucose signaling in Arabidopsis. However, it is not known the degree, if any, that these pathways overlap and how. We show modest signaling crosstalk between these pathways, albeit complex with both epistatic interactions and additive effects that may be indirect. The action of HXK1 on AtRGS1 signaling lies downstream of the primary step in G protein-mediated sugar signaling in which the WD-repeat protein, AGB1, is the propelling signaling element. RHIP1, a previously unknown protein predicted here to have a 3-stranded helical structure, interacts with both AtRGS1 and AtHXK1more » in planta and is required for some glucose-regulated gene expression, providing a physical connection between these two proteins in sugar signaling. The rhip1 null mutant displays similar seedling growth phenotypes as rgs1-2 in response to glucose, further suggesting a role for RHIP1 in glucose signaling. Lastly, glucose signaling is a complex hierarchical relationship which is specific to the target gene and sugar phenotype and suggests that there are two glycolysis-independent glucose signaling sensors: AtRGS1 and AtHXK1 that weakly communicate with each other via feed-back and feed-forward loops to fine tune the response to glucose.« less

Examination of Association to Autism of Common Genetic Variation in Genes Related to Dopamine

PubMed Central

Anderson, B.M.; Schnetz-Boutaud, N.; Bartlett, J.; Wright, H.H.; Abramson, R.K.; Cuccaro, M.L.; Gilbert, J.R.; Pericak-Vance, M.A.; Haines, J.L.

2010-01-01

Autism is a severe neurodevelopmental disorder characterized by a triad of complications. Autistic individuals display significant disturbances in language and reciprocal social interactions, combined with repetitive and stereotypic behaviors. Prevalence studies suggest that autism is more common than originally believed, with recent estimates citing a rate of one in 150. Although this genomic approach has yielded multiple suggestive regions, a specific risk locus has yet to be identified and widely confirmed. Because many etiologies have been suggested for this complex syndrome, we hypothesize that one of the difficulties in identifying autism genes is that multiple genetic variants may be required to significantly increase the risk of developing autism. Thus we took the alternative approach of examining 14 prominent dopamine pathway candidate genes for detailed study by genotyping 28 SNPs. Although we did observe a nominally significant association for rs2239535 (p=.008) on chromosome 20, single locus analysis did not reveal any results as significant after correction for multiple comparisons. No significant interaction was identified when Multifactor Dimensionality Reduction (MDR) was employed to test specifically for multilocus effects. Although genome-wide linkage scans in autism have provided support for linkage to various loci along the dopamine pathway, our study does not provide strong evidence of linkage or association to any specific gene or combination of genes within the pathway. These results demonstrate that common genetic variation within the tested genes located within this pathway at most play a minor to moderate role in overall autism pathogenesis. PMID:19360691

Discovery of functional interactions among actin regulators by analysis of image fluctuations in an unperturbed motile cell system.

PubMed

Isogai, Tadamoto; Danuser, Gaudenz

2018-05-26

Cell migration is driven by propulsive forces derived from polymerizing actin that pushes and extends the plasma membrane. The underlying actin network is constantly undergoing adaptation to new mechano-chemical environments and intracellular conditions. As such, mechanisms that regulate actin dynamics inherently contain multiple feedback loops and redundant pathways. Given the highly adaptable nature of such a system, studies that use only perturbation experiments (e.g. knockdowns, overexpression, pharmacological activation/inhibition, etc.) are challenged by the nonlinearity and redundancy of the pathway. In these pathway configurations, perturbation experiments at best describe the function(s) of a molecular component in an adapting (e.g. acutely drug-treated) or fully adapted (e.g. permanent gene silenced) cell system, where the targeted component now resides in a non-native equilibrium. Here, we propose how quantitative live-cell imaging and analysis of constitutive fluctuations of molecular activities can overcome these limitations. We highlight emerging actin filament barbed-end biology as a prime example of a complex, nonlinear molecular process that requires a fluctuation analytic approach, especially in an unperturbed cellular system, to decipher functional interactions of barbed-end regulators, actin polymerization and membrane protrusion.This article is part of the theme issue 'Self-organization in cell biology'. © 2018 The Author(s).

The role of PACT in the RNA silencing pathway

PubMed Central

Lee, Yoontae; Hur, Inha; Park, Seong-Yeon; Kim, Young-Kook; Suh, Mi Ra; Kim, V Narry

2006-01-01

Small RNA-mediated gene silencing (RNA silencing) has emerged as a major regulatory pathway in eukaryotes. Identification of the key factors involved in this pathway has been a subject of rigorous investigation in recent years. In humans, small RNAs are generated by Dicer and assembled into the effector complex known as RNA-induced silencing complex (RISC) by multiple factors including hAgo2, the mRNA-targeting endonuclease, and TRBP (HIV-1 TAR RNA-binding protein), a dsRNA-binding protein that interacts with both Dicer and hAgo2. Here we describe an additional dsRNA-binding protein known as PACT, which is significant in RNA silencing. PACT is associated with an ∼500 kDa complex that contains Dicer, hAgo2, and TRBP. The interaction with Dicer involves the third dsRNA-binding domain (dsRBD) of PACT and the N-terminal region of Dicer containing the helicase motif. Like TRBP, PACT is not required for the pre-microRNA (miRNA) cleavage reaction step. However, the depletion of PACT strongly affects the accumulation of mature miRNA in vivo and moderately reduces the efficiency of small interfering RNA-induced RNA interference. Our study indicates that, unlike other RNase III type proteins, human Dicer may employ two different dsRBD-containing proteins that facilitate RISC assembly. PMID:16424907

Hippo signaling is required for Notch-dependent smooth muscle differentiation of neural crest.

PubMed

Manderfield, Lauren J; Aghajanian, Haig; Engleka, Kurt A; Lim, Lillian Y; Liu, Feiyan; Jain, Rajan; Li, Li; Olson, Eric N; Epstein, Jonathan A

2015-09-01

Notch signaling has well-defined roles in the assembly of arterial walls and in the development of the endothelium and smooth muscle of the vasculature. Hippo signaling regulates cellular growth in many tissues, and contributes to regulation of organ size, in addition to other functions. Here, we show that the Notch and Hippo pathways converge to regulate smooth muscle differentiation of the neural crest, which is crucial for normal development of the aortic arch arteries and cranial vasculature during embryonic development. Neural crest-specific deletion of the Hippo effectors Yap and Taz produces neural crest precursors that migrate normally, but fail to produce vascular smooth muscle, and Notch target genes such as Jagged1 fail to activate normally. We show that Yap is normally recruited to a tissue-specific Jagged1 enhancer by directly interacting with the Notch intracellular domain (NICD). The Yap-NICD complex is recruited to chromatin by the DNA-binding protein Rbp-J in a Tead-independent fashion. Thus, Hippo signaling can modulate Notch signaling outputs, and components of the Hippo and Notch pathways physically interact. Convergence of Hippo and Notch pathways by the mechanisms described here might be relevant for the function of these signaling cascades in many tissues and in diseases such as cancer. © 2015. Published by The Company of Biologists Ltd.

DELAY OF GERMINATION1 requires PP2C phosphatases of the ABA signalling pathway to control seed dormancy.

PubMed

Née, Guillaume; Kramer, Katharina; Nakabayashi, Kazumi; Yuan, Bingjian; Xiang, Yong; Miatton, Emma; Finkemeier, Iris; Soppe, Wim J J

2017-07-13

The time of seed germination is a major decision point in the life of plants determining future growth and development. This timing is controlled by seed dormancy, which prevents germination under favourable conditions. The plant hormone abscisic acid (ABA) and the protein DELAY OF GERMINATION 1 (DOG1) are essential regulators of dormancy. The function of ABA in dormancy is rather well understood, but the role of DOG1 is still unknown. Here, we describe four phosphatases that interact with DOG1 in seeds. Two of them belong to clade A of type 2C protein phosphatases: ABA-HYPERSENSITIVE GERMINATION 1 (AHG1) and AHG3. These phosphatases have redundant but essential roles in the release of seed dormancy epistatic to DOG1. We propose that the ABA and DOG1 dormancy pathways converge at clade A of type 2C protein phosphatases.The DOG1 protein is a major regulator of seed dormancy in Arabidopsis. Here, Née et al. provide evidence that DOG1 can interact with the type 2C protein phosphatases AHG1 and AHG3 and that this represents the convergence point of the DOG1-regulated dormancy pathway and signalling by the plant hormone abscisic acid.

Deregulated expression of TANK in glioblastomas triggers pro-tumorigenic ERK1/2 and AKT signaling pathways.

PubMed

Stellzig, J; Chariot, A; Shostak, K; Ismail Göktuna, S; Renner, F; Acker, T; Pagenstecher, A; Schmitz, M L

2013-11-11

Signal transmission by the noncanonical IkappaB kinases (IKKs), TANK-binding kinase 1 (TBK1) and IKKɛ, requires interaction with adapter proteins such as TRAF associated NF-κB activator (TANK). Although increased expression or dysregulation of both kinases has been described for a variety of human cancers, this study shows that deregulated expression of the TANK protein is frequently occurring in glioblastomas (GBMs). The functional relevance of TANK was analyzed in a panel of GBM-derived cell lines and revealed that knockdown of TANK arrests cells in the S-phase and prohibits tumor cell migration. Deregulated TANK expression affects several signaling pathways controlling cell proliferation and the inflammatory response. Interference with stoichiometrically assembled signaling complexes by overexpression or silencing of TANK prevented constitutive interferon-regulatory factor 3 (IRF3) phosphorylation. Knockdown of TANK frequently prevents constitutive activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2). TANK-mediated ERK1/2 activation is independent from the canonical MAP kinase or ERK kinase (MEK) 1/2-mediated pathway and utilizes an alternative pathway that uses a TBK1/IKKɛ/Akt signaling axis, thus identifying a novel pathway suitable to block constitutive ERK1/2 activity.

The dorsal "action" pathway.

PubMed

Gallivan, Jason P; Goodale, Melvyn A

2018-01-01

In 1992, Goodale and Milner proposed a division of labor in the visual pathways of the primate cerebral cortex. According to their account, the ventral pathway, which projects to occipitotemporal cortex, constructs our visual percepts, while the dorsal pathway, which projects to posterior parietal cortex, mediates the visual control of action. Although the framing of the two-visual-system hypothesis has not been without controversy, it is clear that vision for action and vision for perception have distinct computational requirements, and significant support for the proposed neuroanatomic division has continued to emerge over the last two decades from human neuropsychology, neuroimaging, behavioral psychophysics, and monkey neurophysiology. In this chapter, we review much of this evidence, with a particular focus on recent findings from human neuroimaging and monkey neurophysiology, demonstrating a specialized role for parietal cortex in visually guided behavior. But even though the available evidence suggests that dedicated circuits mediate action and perception, in order to produce adaptive goal-directed behavior there must be a close coupling and seamless integration of information processing across these two systems. We discuss such ventral-dorsal-stream interactions and argue that the two pathways play different, yet complementary, roles in the production of skilled behavior. Copyright © 2018 Elsevier B.V. All rights reserved.

Deregulated expression of TANK in glioblastomas triggers pro-tumorigenic ERK1/2 and AKT signaling pathways

PubMed Central

Stellzig, J; Chariot, A; Shostak, K; Ismail Göktuna, S; Renner, F; Acker, T; Pagenstecher, A; Schmitz, M L

2013-01-01

Signal transmission by the noncanonical IkappaB kinases (IKKs), TANK-binding kinase 1 (TBK1) and IKKɛ, requires interaction with adapter proteins such as TRAF associated NF-κB activator (TANK). Although increased expression or dysregulation of both kinases has been described for a variety of human cancers, this study shows that deregulated expression of the TANK protein is frequently occurring in glioblastomas (GBMs). The functional relevance of TANK was analyzed in a panel of GBM-derived cell lines and revealed that knockdown of TANK arrests cells in the S-phase and prohibits tumor cell migration. Deregulated TANK expression affects several signaling pathways controlling cell proliferation and the inflammatory response. Interference with stoichiometrically assembled signaling complexes by overexpression or silencing of TANK prevented constitutive interferon-regulatory factor 3 (IRF3) phosphorylation. Knockdown of TANK frequently prevents constitutive activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2). TANK-mediated ERK1/2 activation is independent from the canonical MAP kinase or ERK kinase (MEK) 1/2-mediated pathway and utilizes an alternative pathway that uses a TBK1/IKKɛ/Akt signaling axis, thus identifying a novel pathway suitable to block constitutive ERK1/2 activity. PMID:24217713

The Use of Gene Ontology Term and KEGG Pathway Enrichment for Analysis of Drug Half-Life

PubMed Central

Chen, Lei; Lu, Jing; Kong, XiangYin; Huang, Tao; Li, HaiPeng

2016-01-01

A drug’s biological half-life is defined as the time required for the human body to metabolize or eliminate 50% of the initial drug dosage. Correctly measuring the half-life of a given drug is helpful for the safe and accurate usage of the drug. In this study, we investigated which gene ontology (GO) terms and biological pathways were highly related to the determination of drug half-life. The investigated drugs, with known half-lives, were analyzed based on their enrichment scores for associated GO terms and KEGG pathways. These scores indicate which GO terms or KEGG pathways the drug targets. The feature selection method, minimum redundancy maximum relevance, was used to analyze these GO terms and KEGG pathways and to identify important GO terms and pathways, such as sodium-independent organic anion transmembrane transporter activity (GO:0015347), monoamine transmembrane transporter activity (GO:0008504), negative regulation of synaptic transmission (GO:0050805), neuroactive ligand-receptor interaction (hsa04080), serotonergic synapse (hsa04726), and linoleic acid metabolism (hsa00591), among others. This analysis confirmed our results and may show evidence for a new method in studying drug half-lives and building effective computational methods for the prediction of drug half-lives. PMID:27780226

Western and Chinese antirheumatic drug-induced T cell apoptotic DNA damage uses different caspase cascades and is independent of Fas/Fas ligand interaction.

PubMed

Lai, J H; Ho, L J; Lu, K C; Chang, D M; Shaio, M F; Han, S H

2001-06-01

Spontaneous or therapeutic induction of T cell apoptosis plays a critical role in establishing transplantation tolerance and maintaining remission of autoimmune diseases. We investigated the mechanisms of apoptosis induced by Chinese and Western antirheumatic drugs (ARDs) in human T cells. We found that hydroxychloroquine, Tripterygium wilfordii hook F, and tetrandrine (Tet), but not methotrexate, at therapeutic concentrations can cause T cell death. In addition, Tet selectively killed T cells, especially activated T cells. Although ARD-induced cytotoxicity was mediated through apoptotic mechanisms, Fas/Fas ligand interaction was not required. We further demonstrated that the processes of phosphatidylserine externalization and DNA damage along the ARD-induced T cell apoptotic pathway could operate independently, and that selective inhibition of DNA damage by caspase inhibitors did not prevent T cells from undergoing cell death. Moreover, we found that Tet- and Tripterygium wilfordii hook F-induced T cell DNA damage required caspase-3 activity, and hydroxychloroquine-induced T cell DNA damage was mediated through a caspase-3- and caspase-8-independent, but Z-Asp-Glu-Val-Asp-fluomethyl ketone-sensitive, signaling pathway. Finally, the observation that ARD-induced activation of caspase-3 in both Fas-sensitive and Fas-resistant Jurkat T cells indicates that Fas/Fas ligand interaction plays no role in ARD-induced T cell apoptosis. Our observations provide new information about the complex apoptotic mechanisms of ARDs, and have implications for combining Western and Chinese ARDs that have different immunomodulatory mechanisms in the therapy of autoimmune diseases and transplantation rejection.

Utilization of Integrated Assessment Modeling for determining geologic CO2 storage security

NASA Astrophysics Data System (ADS)

Pawar, R.

2017-12-01

Geologic storage of carbon dioxide (CO2) has been extensively studied as a potential technology to mitigate atmospheric concentration of CO2. Multiple international research & development efforts, large-scale demonstration and commercial projects are helping advance the technology. One of the critical areas of active investigation is prediction of long-term CO2 storage security and risks. A quantitative methodology for predicting a storage site's long-term performance is critical for making key decisions necessary for successful deployment of commercial scale projects where projects will require quantitative assessments of potential long-term liabilities. These predictions are challenging given that they require simulating CO2 and in-situ fluid movements as well as interactions through the primary storage reservoir, potential leakage pathways (such as wellbores, faults, etc.) and shallow resources such as groundwater aquifers. They need to take into account the inherent variability and uncertainties at geologic sites. This talk will provide an overview of an approach based on integrated assessment modeling (IAM) to predict long-term performance of a geologic storage site including, storage reservoir, potential leakage pathways and shallow groundwater aquifers. The approach utilizes reduced order models (ROMs) to capture the complex physical/chemical interactions resulting due to CO2 movement and interactions but are computationally extremely efficient. Applicability of the approach will be demonstrated through examples that are focused on key storage security questions such as what is the probability of leakage of CO2 from a storage reservoir? how does storage security vary for different geologic environments and operational conditions? how site parameter variability and uncertainties affect storage security, etc.

Dual chromatin recognition by the histone deacetylase complex HCHC is required for proper DNA methylation in Neurospora crassa

PubMed Central

Honda, Shinji; Bicocca, Vincent T.; Gessaman, Jordan D.; Rountree, Michael R.; Yokoyama, Ayumi; Yu, Eun Y.; Selker, Jeanne M. L.; Selker, Eric U.

2016-01-01

DNA methylation, heterochromatin protein 1 (HP1), histone H3 lysine 9 (H3K9) methylation, histone deacetylation, and highly repeated sequences are prototypical heterochromatic features, but their interrelationships are not fully understood. Prior work showed that H3K9 methylation directs DNA methylation and histone deacetylation via HP1 in Neurospora crassa and that the histone deacetylase complex HCHC is required for proper DNA methylation. The complex consists of the chromodomain proteins HP1 and chromodomain protein 2 (CDP-2), the histone deacetylase HDA-1, and the AT-hook motif protein CDP-2/HDA-1–associated protein (CHAP). We show that the complex is required for proper chromosome segregation, dissect its function, and characterize interactions among its components. Our analyses revealed the existence of an HP1-based DNA methylation pathway independent of its chromodomain. The pathway partially depends on CHAP but not on the CDP-2 chromodomain. CDP-2 serves as a bridge between the recognition of H3K9 trimethylation (H3K9me3) by HP1 and the histone deacetylase activity of HDA-1. CHAP is also critical for HDA-1 localization to heterochromatin. Specifically, the CHAP zinc finger interacts directly with the HDA-1 argonaute-binding protein 2 (Arb2) domain, and the CHAP AT-hook motifs recognize heterochromatic regions by binding to AT-rich DNA. Our data shed light on the interrelationships among the prototypical heterochromatic features and support a model in which dual recognition by the HP1 chromodomain and the CHAP AT-hooks are required for proper heterochromatin formation. PMID:27681634

Framing ethnic variations in alcohol outcomes from biological pathways to neighborhood context.

PubMed

Chartier, Karen G; Scott, Denise M; Wall, Tamara L; Covault, Jonathan; Karriker-Jaffe, Katherine J; Mills, Britain A; Luczak, Susan E; Caetano, Raul; Arroyo, Judith A

2014-03-01

Health disparities research seeks to eliminate disproportionate negative health outcomes experienced in some racial/ethnic minority groups. This brief review presents findings on factors associated with drinking and alcohol-related problems in racial/ethnic groups. Those discussed are as follows: (i) biological pathways to alcohol problems, (ii) gene × stress interactions, (iii) neighborhood disadvantage, stress, and access to alcohol, and (iv) drinking cultures and contexts. These factors and their interrelationships are complex, requiring a multilevel perspective. The use of interdisciplinary teams and an epigenetic focus are suggested to move the research forward. The application of multilevel research to policy, prevention, and intervention programs may help prioritize combinations of the most promising intervention targets. Copyright © 2014 by the Research Society on Alcoholism.

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

PubMed

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

2016-09-01

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

Kinase Pathway Database: An Integrated Protein-Kinase and NLP-Based Protein-Interaction Resource

PubMed Central

Koike, Asako; Kobayashi, Yoshiyuki; Takagi, Toshihisa

2003-01-01

Protein kinases play a crucial role in the regulation of cellular functions. Various kinds of information about these molecules are important for understanding signaling pathways and organism characteristics. We have developed the Kinase Pathway Database, an integrated database involving major completely sequenced eukaryotes. It contains the classification of protein kinases and their functional conservation, ortholog tables among species, protein–protein, protein–gene, and protein–compound interaction data, domain information, and structural information. It also provides an automatic pathway graphic image interface. The protein, gene, and compound interactions are automatically extracted from abstracts for all genes and proteins by natural-language processing (NLP).The method of automatic extraction uses phrase patterns and the GENA protein, gene, and compound name dictionary, which was developed by our group. With this database, pathways are easily compared among species using data with more than 47,000 protein interactions and protein kinase ortholog tables. The database is available for querying and browsing at http://kinasedb.ontology.ims.u-tokyo.ac.jp/. PMID:12799355

Brain Transcriptomic Response to Social Eavesdropping in Zebrafish (Danio rerio)

PubMed Central

Oliveira, Rui F.

2015-01-01

Public information is widely available at low cost to animals living in social groups. For instance, bystanders may eavesdrop on signaling interactions between conspecifics and use it to adapt their subsequent behavior towards the observed individuals. This social eavesdropping ability is expected to require specialized mechanisms such as social attention, which selects social information available for learning. To begin exploring the genetic basis of social eavesdropping, we used a previously established attention paradigm in the lab to study the brain gene expression profile of male zebrafish (Danio rerio) in relation to the attention they paid towards conspecifics involved or not involved in agonistic interactions. Microarray gene chips were used to characterize their brain transcriptomes based on differential expression of single genes and gene sets. These analyses were complemented by promoter region-based techniques. Using data from both approaches, we further drafted protein interaction networks. Our results suggest that attentiveness towards conspecifics, whether interacting or not, activates pathways linked to neuronal plasticity and memory formation. The network analyses suggested that fos and jun are key players on this response, and that npas4a, nr4a1 and egr4 may also play an important role. Furthermore, specifically observing fighting interactions further triggered pathways associated to a change in the alertness status (dnajb5) and to other genes related to memory formation (btg2, npas4b), which suggests that the acquisition of eavesdropped information about social relationships activates specific processes on top of those already activated just by observing conspecifics. PMID:26713440

[Mechanistic modelling allows to assess pathways of DNA lesion interactions underlying chromosome aberration formation].

PubMed

Eĭdel'man, Iu A; Slanina, S V; Sal'nikov, I V; Andreev, S G

2012-12-01

The knowledge of radiation-induced chromosomal aberration (CA) mechanisms is required in many fields of radiation genetics, radiation biology, biodosimetry, etc. However, these mechanisms are yet to be quantitatively characterised. One of the reasons is that the relationships between primary lesions of DNA/chromatin/chromosomes and dose-response curves for CA are unknown because the pathways of lesion interactions in an interphase nucleus are currently inaccessible for direct experimental observation. This article aims for the comparative analysis of two principally different scenarios of formation of simple and complex interchromosomal exchange aberrations: by lesion interactions at chromosome territories' surface vs. in the whole space of the nucleus. The analysis was based on quantitative mechanistic modelling of different levels of structures and processes involved in CA formation: chromosome structure in an interphase nucleus, induction, repair and interactions of DNA lesions. It was shown that the restricted diffusion of chromosomal loci, predicted by computational modelling of chromosome organization, results in lesion interactions in the whole space of the nucleus being impossible. At the same time, predicted features of subchromosomal dynamics agrees well with in vivo observations and does not contradict the mechanism of CA formation at the surface of chromosome territories. On the other hand, the "surface mechanism" of CA formation, despite having certain qualities, proved to be insufficient to explain high frequency of complex exchange aberrations observed by mFISH technique. The alternative mechanism, CA formation on nuclear centres is expected to be sufficient to explain frequent complex exchanges.

CDE-1 affects chromosome segregation through uridylation of CSR-1-bound siRNAs.

PubMed

van Wolfswinkel, Josien C; Claycomb, Julie M; Batista, Pedro J; Mello, Craig C; Berezikov, Eugene; Ketting, René F

2009-10-02

We have studied the function of a conserved germline-specific nucleotidyltransferase protein, CDE-1, in RNAi and chromosome segregation in C. elegans. CDE-1 localizes specifically to mitotic chromosomes in embryos. This localization requires the RdRP EGO-1, which physically interacts with CDE-1, and the Argonaute protein CSR-1. We found that CDE-1 is required for the uridylation of CSR-1 bound siRNAs, and that in the absence of CDE-1 these siRNAs accumulate to inappropriate levels, accompanied by defects in both meiotic and mitotic chromosome segregation. Elevated siRNA levels are associated with erroneous gene silencing, most likely through the inappropriate loading of CSR-1 siRNAs into other Argonaute proteins. We propose a model in which CDE-1 restricts specific EGO-1-generated siRNAs to the CSR-1 mediated, chromosome associated RNAi pathway, thus separating it from other endogenous RNAi pathways. The conserved nature of CDE-1 suggests that similar sorting mechanisms may operate in other animals, including mammals.

SMZ/SNZ and gibberellin signaling are required for nitrate-elicited delay of flowering time in Arabidopsis thaliana.

PubMed

Gras, Diana E; Vidal, Elena A; Undurraga, Soledad F; Riveras, Eleodoro; Moreno, Sebastián; Dominguez-Figueroa, José; Alabadi, David; Blázquez, Miguel A; Medina, Joaquín; Gutiérrez, Rodrigo A

2018-01-23

The reproductive success of plants largely depends on the correct programming of developmental phase transitions, particularly the shift from vegetative to reproductive growth. The timing of this transition is finely regulated by the integration of an array of environmental and endogenous factors. Nitrogen is the mineral macronutrient that plants require in the largest amount, and as such its availability greatly impacts on many aspects of plant growth and development, including flowering time. We found that nitrate signaling interacts with the age-related and gibberellic acid pathways to control flowering time in Arabidopsis thaliana. We revealed that repressors of flowering time belonging to the AP2-type transcription factor family including SCHLAFMUTZE (SMZ) and SCHNARCHZAPFEN (SNZ) are important regulators of flowering time in response to nitrate. Our results support a model whereby nitrate activates SMZ and SNZ via the gibberellin pathway to repress flowering time in Arabidopsis thaliana. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Opposing activities of Notch and Wnt signaling regulate intestinal stem cells and gut homeostasis

PubMed Central

Tian, Hua; Biehs, Brian; Chiu, Cecilia; Siebel, Chris; Wu, Yan; Costa, Mike; de Sauvage, Frederic J.; Klein, Ophir D.

2015-01-01

Summary Proper organ homeostasis requires tight control of adult stem cells and differentiation through integration of multiple inputs. In the mouse small intestine, Notch and Wnt signaling are required both for stem cell maintenance and for a proper balance of differentiation between secretory and absorptive cell lineages. In the absence of Notch signaling, stem cells preferentially generate secretory cells at the expense of absorptive cells. Here, we use function-blocking antibodies against Notch receptors to demonstrate that Notch blockade perturbs intestinal stem cell function by causing a de-repression of the Wnt signaling pathway, leading to mis-expression of prosecretory genes. Importantly, attenuation of the Wnt pathway rescued the phenotype associated with Notch blockade. These studies bring to light a negative regulatory mechanism that maintains stem cell activity and balanced differentiation, and we propose that the interaction between Wnt and Notch signaling described here represents a common theme in adult stem cell biology. PMID:25818302

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.

Guanosine triphosphatase activation occurs downstream of calcineurin in cardiac hypertrophy*.

PubMed

Richardson, Kenneth E; Tannous, Paul; Berenji, Kambeez; Nolan, Bridgid; Bayless, Kayla J; Davis, George E; Rothermel, Beverly A; Hill, Joseph A

2005-12-01

There is great interest in deciphering mechanisms of maladaptive remodeling in cardiac hypertrophy in the hope of affording clinical benefit. Potential targets of therapeutic intervention include the cytoplasmic phosphatase calcineurin and small guanosine triphosphate-binding proteins, such as Rac1 and RhoA, all of which have been implicated in maladaptive hypertrophy. However, little is known about the interaction-if any-between these important signaling molecules in hypertrophic heart disease. In this study, we examined the molecular interplay among these molecules, finding that Rho family guanosine triphosphatase signaling occurs either downstream of calcineurin or as a required, parallel pathway. It has been shown that 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibition blocks hypertrophy, and we report here that "statin" therapy effectively suppresses small G protein activation and blunts hypertrophic growth in vitro and in vivo. Importantly, despite significant suppression of hypertrophy, clinical and hemodynamic markers remained compensated, suggesting that the hypertrophic growth induced by this pathway is not required to maintain circulatory performance.

A network approach to predict pathogenic genes for Fusarium graminearum.

PubMed

Liu, Xiaoping; Tang, Wei-Hua; Zhao, Xing-Ming; Chen, Luonan

2010-10-04

Fusarium graminearum is the pathogenic agent of Fusarium head blight (FHB), which is a destructive disease on wheat and barley, thereby causing huge economic loss and health problems to human by contaminating foods. Identifying pathogenic genes can shed light on pathogenesis underlying the interaction between F. graminearum and its plant host. However, it is difficult to detect pathogenic genes for this destructive pathogen by time-consuming and expensive molecular biological experiments in lab. On the other hand, computational methods provide an alternative way to solve this problem. Since pathogenesis is a complicated procedure that involves complex regulations and interactions, the molecular interaction network of F. graminearum can give clues to potential pathogenic genes. Furthermore, the gene expression data of F. graminearum before and after its invasion into plant host can also provide useful information. In this paper, a novel systems biology approach is presented to predict pathogenic genes of F. graminearum based on molecular interaction network and gene expression data. With a small number of known pathogenic genes as seed genes, a subnetwork that consists of potential pathogenic genes is identified from the protein-protein interaction network (PPIN) of F. graminearum, where the genes in the subnetwork are further required to be differentially expressed before and after the invasion of the pathogenic fungus. Therefore, the candidate genes in the subnetwork are expected to be involved in the same biological processes as seed genes, which imply that they are potential pathogenic genes. The prediction results show that most of the pathogenic genes of F. graminearum are enriched in two important signal transduction pathways, including G protein coupled receptor pathway and MAPK signaling pathway, which are known related to pathogenesis in other fungi. In addition, several pathogenic genes predicted by our method are verified in other pathogenic fungi, which demonstrate the effectiveness of the proposed method. The results presented in this paper not only can provide guidelines for future experimental verification, but also shed light on the pathogenesis of the destructive fungus F. graminearum.

Coordination of Rad1-Rad10 interactions with Msh2-Msh3, Saw1 and RPA is essential for functional 3' non-homologous tail removal.

PubMed

Eichmiller, Robin; Medina-Rivera, Melisa; DeSanto, Rachel; Minca, Eugen; Kim, Christopher; Holland, Cory; Seol, Ja-Hwan; Schmit, Megan; Oramus, Diane; Smith, Jessica; Gallardo, Ignacio F; Finkelstein, Ilya J; Lee, Sang Eun; Surtees, Jennifer A

2018-06-01

Double strand DNA break repair (DSBR) comprises multiple pathways. A subset of DSBR pathways, including single strand annealing, involve intermediates with 3' non-homologous tails that must be removed to complete repair. In Saccharomyces cerevisiae, Rad1-Rad10 is the structure-specific endonuclease that cleaves the tails in 3' non-homologous tail removal (3' NHTR). Rad1-Rad10 is also an essential component of the nucleotide excision repair (NER) pathway. In both cases, Rad1-Rad10 requires protein partners for recruitment to the relevant DNA intermediate. Msh2-Msh3 and Saw1 recruit Rad1-Rad10 in 3' NHTR; Rad14 recruits Rad1-Rad10 in NER. We created two rad1 separation-of-function alleles, rad1R203A,K205A and rad1R218A; both are defective in 3' NHTR but functional in NER. In vitro, rad1R203A,K205A was impaired at multiple steps in 3' NHTR. The rad1R218A in vivo phenotype resembles that of msh2- or msh3-deleted cells; recruitment of rad1R218A-Rad10 to recombination intermediates is defective. Interactions among rad1R218A-Rad10 and Msh2-Msh3 and Saw1 are altered and rad1R218A-Rad10 interactions with RPA are compromised. We propose a model in which Rad1-Rad10 is recruited and positioned at the recombination intermediate through interactions, between Saw1 and DNA, Rad1-Rad10 and Msh2-Msh3, Saw1 and Msh2-Msh3 and Rad1-Rad10 and RPA. When any of these interactions is altered, 3' NHTR is impaired.

Fibroblast growth factor and cyclic AMP (cAMP) synergistically activate gene expression at a cAMP response element.

PubMed Central

Tan, Y; Low, K G; Boccia, C; Grossman, J; Comb, M J

1994-01-01

Growth factors and cyclic AMP (cAMP) are known to activate distinct intracellular signaling pathways. Fibroblast growth factor (FGF) activates ras-dependent kinase cascades, resulting in the activation of MAP kinases, whereas cAMP activates protein kinase A. In this study, we report that growth factors and cAMP act synergistically to stimulate proenkephalin gene expression. Positive synergy between growth factor- and cAMP-activated signaling pathways on gene expression has not been previously reported, and we suggest that these synergistic interactions represent a useful model for analyzing interactions between these pathways. Transfection and mutational studies indicate that both FGF-dependent gene activation and cAMP-dependent gene activation require cAMP response element 2 (CRE-2), a previously characterized cAMP-dependent regulatory element. Furthermore, multiple copies of this element are sufficient to confer FGF regulation upon a minimal promoter, indicating that FGF and cAMP signaling converge upon transcription factors acting at CRE-2. Among many different ATF/AP-1 factors tested, two factors, ATF-3 and c-Jun, stimulate proenkephalin transcription in an FGF- or Ras-dependent fashion. Finally, we show that ATF-3 and c-Jun form heterodimeric complexes in SK-N-MC cells and that the levels of both proteins are increased in response to FGF but not cAMP. Together, these results indicate that growth factor- and cAMP-dependent signaling pathways converge at CRE-2 to synergistically stimulate gene expression and that ATF-3 and c-Jun regulate proenkephalin transcription in response to both growth factor- and cAMP-dependent intracellular signaling pathways. Images PMID:7935470

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

PubMed Central

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

2016-01-01

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

Control of the mitotic exit network during meiosis

PubMed Central

Attner, Michelle A.; Amon, Angelika

2012-01-01

The mitotic exit network (MEN) is an essential GTPase signaling pathway that triggers exit from mitosis in budding yeast. We show here that during meiosis, the MEN is dispensable for exit from meiosis I but contributes to the timely exit from meiosis II. Consistent with a role for the MEN during meiosis II, we find that the signaling pathway is active only during meiosis II. Our analysis further shows that MEN signaling is modulated during meiosis in several key ways. Whereas binding of MEN components to spindle pole bodies (SPBs) is necessary for MEN signaling during mitosis, during meiosis MEN signaling occurs off SPBs and does not require the SPB recruitment factor Nud1. Furthermore, unlike during mitosis, MEN signaling is controlled through the regulated interaction between the MEN kinase Dbf20 and its activating subunit Mob1. Our data lead to the conclusion that a pathway essential for vegetative growth is largely dispensable for the specialized meiotic divisions and provide insights into how cell cycle regulatory pathways are modulated to accommodate different modes of cell division. PMID:22718910

Simulations of polymorphic icosahedral shells assembling around many cargo molecules

NASA Astrophysics Data System (ADS)

Mohajerani, Farzaneh; Perlmutter, Jason; Hagan, Michael

Bacterial microcompartments (BMCs) are large icosahedral shells that sequester the enzymes and reactants responsible for particular metabolic pathways in bacteria. Although different BMCs vary in size and encapsulate different cargoes, they are constructed from similar pentameric and hexameric shell proteins. Despite recent groundbreaking experiments which visualized the formation of individual BMCs, the detailed assembly pathways and the factors which control shell size remain unclear. In this talk, we describe theoretical and computational models that describe the dynamical encapsulation of hundreds of cargo molecules by self-assembling icosahedral shells. We present phase diagrams and analysis of dynamical simulation trajectories showing how the thermodynamics, assembly pathways, and emergent structures depend on the interactions among shell proteins and cargo molecules. Our model suggests a mechanism for controlling insertion of the 12 pentamers required for a closed shell topology, and the relationship between assembly pathway and BMC size polydispersity. In addition to elucidating how native BMCs assemble,our results establish principles for reengineering BMCs or viral capsids as customizable nanoreactors that can assemble around a programmable set of enzymes and reactants. Supported by NIH R01GM108021 and Brandeis MRSEC DMR-1420382.

Long noncoding ribonucleic acid NKILA induces the endoplasmic reticulum stress/autophagy pathway and inhibits the nuclear factor-k-gene binding pathway in rats after intracerebral hemorrhage.

PubMed

Jia, Jiaoying; Zhang, Mingming; Li, Qi; Zhou, Qian; Jiang, Yugang

2018-06-12

Long noncoding RNAs (lncRNAs) have emerged as an important class of molecules that have been associated with brain function and neurological disease, but the expression profiles of lncRNAs after intracerebral hemorrhage (ICH) remain to be elucidated. In this study, we determined the expression pattern of nuclear factor-k-gene binding (NF-kB) interacting lncRNA (NKILA) after ICH and examined its respective effects on the endoplasmic reticulum stress (ERS)/autophagy pathway, hippocampal neuron loss, and the NF-kB pathway after type VII collagenase-induced ICH in rats. The regulatory mechanisms of NKILA were investigated by an intraperitoneal injection of small interfering (siRNA) against NKILA into rats after ICH. NKILA inhibition mediated by siRNA against NKILA was shown to significantly reduce ERS and autophagy, activate the NF-kB pathway, decrease neurological deficits, brain edema, and injury, and induce blood-brain barrier breakdown, further leading to hippocampal neuron loss and the production of inflammation cytokines. Taken together, the demonstration that NKILA induces the ERS/autophagy pathway and inhibits the NF-kB pathway after ICH supports the concept that NKILA functions as a novel target that is required for the attenuation of brain injuries after ICH. © 2018 Wiley Periodicals, Inc.

Volatile communication in plant-aphid interactions.

PubMed

de Vos, Martin; Jander, Georg

2010-08-01

Volatile communication plays an important role in mediating the interactions between plants, aphids, and other organisms in the environment. In response to aphid infestation, many plants initiate indirect defenses through the release of volatiles that attract ladybugs, parasitoid wasps, and other aphid-consuming predators. Aphid-induced volatile release in the model plant Arabidopsis thaliana requires the jasmonate signaling pathway. Volatile release is also induced by infection with aphid-transmitted viruses. Consistent with mathematical models of optimal transmission, viruses that are acquired rapidly by aphids induce volatile release to attract migratory aphids, but discourage long-term aphid feeding. Although the ecology of these interactions is well-studied, further research is needed to identify the molecular basis of aphid-induced and virus-induced changes in plant volatile release. Copyright 2010 Elsevier Ltd. All rights reserved.

Prickle and Strabismus form a functional complex to generate a correct axis during planar cell polarity signaling

PubMed Central

Jenny, Andreas; Darken, Rachel S.; Wilson, Paul A.; Mlodzik, Marek

2003-01-01

Frizzled (Fz) signaling regulates the establishment of planar cell polarity (PCP). The PCP genes prickle (pk) and strabismus (stbm) are thought to antagonize Fz signaling. We show that they act in the same cell, R4, adjacent to that in which the Fz/PCP pathway is required in the Drosophila eye. We demonstrate that Stbm and Pk interact physically and that Stbm recruits Pk to the cell membrane. Through this interaction, Pk affects Stbm membrane localization and can cause clustering of Stbm. Pk is also known to interact with Dsh and is thought to antagonize Dsh by affecting its membrane localization. Thus our data suggest that the Stbm/Pk complex modulates Fz/Dsh activity, resulting in a symmetry-breaking step during polarity signaling. PMID:12941693

MYB75 Phosphorylation by MPK4 Is Required for Light-Induced Anthocyanin Accumulation in Arabidopsis[OPEN

PubMed Central

Li, Shengnan; Wang, Wenyi; Gao, Jinlan; Yin, Kangquan; Wang, Rui; Wang, Chengcheng; Mundy, John

2016-01-01

Light is a major environmental cue affecting various physiological and metabolic processes in plants. Although plant photoreceptors are well characterized, the mechanisms by which light regulates downstream responses are less clear. In Arabidopsis thaliana, the accumulation of photoprotective anthocyanin pigments is light dependent, and the R2R3 MYB transcription factor MYB75/PAP1 regulates anthocyanin accumulation. Here, we report that MYB75 interacts with and is phosphorylated by MAP KINASE4 (MPK4). Their interaction is dependent on MPK4 kinase activity and is required for full function of MYB75. MPK4 can be activated in response to light and is involved in the light-induced accumulation of anthocyanins. We show that MPK4 phosphorylation of MYB75 increases its stability and is essential for light-induced anthocyanin accumulation. Our findings reveal an important role for a MAPK pathway in light signal transduction. PMID:27811015

Identification of Genes That Interact With Drosophila liquid facets

PubMed Central

Eun, Suk Ho; Lea, Kristi; Overstreet, Erin; Stevens, Samuel; Lee, Ji-Hoon; Fischer, Janice A.

2007-01-01

We have performed mutagenesis screens of the Drosophila X chromosome and the autosomes for dominant enhancers of the rough eye resulting from overexpression of liquid facets. The liquid facets gene encodes the homolog of vertebrate endocytic Epsin, an endocytic adapter protein. In Drosophila, Liquid facets is a core component of the Notch signaling pathway required in the signaling cells for ligand endocytosis and signaling. Why ligand internalization by the signaling cells is essential for signaling is a mystery. The requirement for Liquid facets is a hint at the answer, and the genes identified in this screen provide further clues. Mutant alleles of clathrin heavy chain, Rala, split ends, and auxilin were identified as enhancers. We describe the mutant alleles and mutant phenotypes of Rala and aux. We discuss the relevance of all of these genetic interactions to the function of Liquid facets in Notch signaling. PMID:17179082

Topological, functional, and dynamic properties of the protein interaction networks rewired by benzo(a)pyrene

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

Ba, Qian; Key Laboratory of Food Safety Risk Assessment, Ministry of Health, Beijing; Li, Junyang

2015-03-01

Benzo(a)pyrene is a common environmental and foodborne pollutant that has been identified as a human carcinogen. Although the carcinogenicity of benzo(a)pyrene has been extensively reported, its precise molecular mechanisms and the influence on system-level protein networks are not well understood. To investigate the system-level influence of benzo(a)pyrene on protein interactions and regulatory networks, a benzo(a)pyrene-rewired protein interaction network was constructed based on 769 key proteins derived from more than 500 literature reports. The protein interaction network rewired by benzo(a)pyrene was a scale-free, highly-connected biological system. Ten modules were identified, and 25 signaling pathways were enriched, most of which belong tomore » the human diseases category, especially cancer and infectious disease. In addition, two lung-specific and two liver-specific pathways were identified. Three pathways were specific in short and medium-term networks (< 48 h), and five pathways were enriched only in the medium-term network (6 h–48 h). Finally, the expression of linker genes in the network was validated by Western blotting. These findings establish the overall, tissue- and time-specific benzo(a)pyrene-rewired protein interaction networks and provide insights into the biological effects and molecular mechanisms of action of benzo(a)pyrene. - Highlights: • Benzo(a)pyrene induced scale-free, highly-connected protein interaction networks. • 25 signaling pathways were enriched through modular analysis. • Tissue- and time-specific pathways were identified.« less

Metabotropic Glutamate Receptors and Interacting Proteins in Epileptogenesis

PubMed Central

Qian, Feng; Tang, Feng-Ru

2016-01-01

Neurotransmitter and receptor systems are involved in different neurological and neuropsychological disorders such as Parkinson's disease, depression, Alzheimer’s disease and epilepsy. Recent advances in studies of signal transduction pathways or interacting proteins of neurotransmitter receptor systems suggest that different receptor systems may share the common signal transduction pathways or interacting proteins which may be better therapeutic targets for development of drugs to effectively control brain diseases. In this paper, we reviewed metabotropic glutamate receptors (mGluRs) and their related signal transduction pathways or interacting proteins in status epilepticus and temporal lobe epilepsy, and proposed some novel therapeutical drug targets for controlling epilepsy and epileptogenesis. PMID:27030135

Urothelium update: how the bladder mucosa measures bladder filling.

PubMed

Janssen, D A W; Schalken, J A; Heesakkers, J P F A

2017-06-01

This review critically evaluates the evidence on mechanoreceptors and pathways in the bladder urothelium that are involved in normal bladder filling signalling. Evidence from in vitro and in vivo studies on (i) signalling pathways like the adenosine triphosphate pathway, cholinergic pathway and nitric oxide and adrenergic pathway, and (ii) different urothelial receptors that are involved in bladder filling signalling like purinergic receptors, sodium channels and TRP channels will be evaluated. Other potential pathways and receptors will also be discussed. Bladder filling results in continuous changes in bladder wall stretch and exposure to urine. Both barrier and afferent signalling functions in the urothelium are constantly adapting to cope with these dynamics. Current evidence shows that the bladder mucosa hosts essential pathways and receptors that mediate bladder filling signalling. Intracellular calcium ion increase is a dominant factor in this signalling process. However, there is still no complete understanding how interacting receptors and pathways create a bladder filling signal. Currently, there are still novel receptors investigated that could also be participating in bladder filling signalling. Normal bladder filling sensation is dependent on multiple interacting mechanoreceptors and signalling pathways. Research efforts need to focus on how these pathways and receptors interact to fully understand normal bladder filling signalling. © 2016 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Pathway collages: personalized multi-pathway diagrams.

PubMed

Paley, Suzanne; O'Maille, Paul E; Weaver, Daniel; Karp, Peter D

2016-12-13

Metabolic pathway diagrams are a classical way of visualizing a linked cascade of biochemical reactions. However, to understand some biochemical situations, viewing a single pathway is insufficient, whereas viewing the entire metabolic network results in information overload. How do we enable scientists to rapidly construct personalized multi-pathway diagrams that depict a desired collection of interacting pathways that emphasize particular pathway interactions? We define software for constructing personalized multi-pathway diagrams called pathway-collages using a combination of manual and automatic layouts. The user specifies a set of pathways of interest for the collage from a Pathway/Genome Database. Layouts for the individual pathways are generated by the Pathway Tools software, and are sent to a Javascript Pathway Collage application implemented using Cytoscape.js. That application allows the user to re-position pathways; define connections between pathways; change visual style parameters; and paint metabolomics, gene expression, and reaction flux data onto the collage to obtain a desired multi-pathway diagram. We demonstrate the use of pathway collages in two application areas: a metabolomics study of pathogen drug response, and an Escherichia coli metabolic model. Pathway collages enable facile construction of personalized multi-pathway diagrams.

Clinical evidence on the magnitude of change in growth pathway activity in relation to Tamoxifen resistance is required.

PubMed

Mansouri, Sepideh; Farahmand, Leila; Teymourzadeh, Azin; Majidzadeh-A, Keivan

2017-08-08

Despite prolonged disease-free survival and overall survival rates in estrogen receptor (ER)-positive patients undergoing adjuvant treatment, Tamoxifen therapy tends to fail due to eventual acquisition of resistance. Although numerous studies have emphasized the role of receptor tyrosine kinases (RTKs) in the development of Tamoxifen resistance, inadequate clinical evidence is available regarding the alteration of biomarker expression during acquired resistance, thus undermining the validity of the findings. Results of two meta-analyses investigating the effect of HER2 status on the prognosis of Tamoxifen-receiving patients have demonstrated that despite HER2-negative patients having longer disease-free survival; there is no difference in overhaul survival between the two groups. Furthermore, due to the intricate molecular interactions among estrogen receptors including ERα36, ERα66, and also RTKs, it is not surprising that RTK suppression does not restore Tamoxifen sensitivity. In considering such a complex network, we speculate that by the time HER2/EGFR is suppressed via targeted therapies, activation of ERα66 and ERα36 initiate molecular signaling pathways downstream of RTKs, thereby enhancing cell proliferation even in the presence of both Tamoxifen and RTK inhibitors. Although clinical findings regarding the molecular pathways downstream of RTKs have been thoroughly discussed in this review, further clinical studies are required in determining a consistency between preclinical and clinical findings. Discovering the best targets in preventing tumor progression requires thorough comprehension of estrogen-dependent and estrogen-independent pathways during Tamoxifen resistance development. Indeed, exploring additional clinically-proven targets would allow for better characterized treatments being available for breast cancer patients. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

FANC Pathway Promotes UV-Induced Stalled Replication Forks Recovery by Acting Both Upstream and Downstream Polη and Rev1

PubMed Central

Renaud, Emilie; Rosselli, Filippo

2013-01-01

To cope with ultraviolet C (UVC)-stalled replication forks and restart DNA synthesis, cells either undergo DNA translesion synthesis (TLS) by specialised DNA polymerases or tolerate the lesions using homologous recombination (HR)-based mechanisms. To gain insight into how cells manage UVC-induced stalled replication forks, we analysed the molecular crosstalk between the TLS DNA polymerases Polη and Rev1, the double-strand break repair (DSB)-associated protein MDC1 and the FANC pathway. We describe three novel functional interactions that occur in response to UVC-induced DNA lesions. First, Polη and Rev1, whose optimal expression and/or relocalisation depend on the FANC core complex, act upstream of FANCD2 and are required for the proper relocalisation of monoubiquitinylated FANCD2 (Ub-FANCD2) to subnuclear foci. Second, during S-phase, Ub-FANCD2 and MDC1 relocalise to UVC-damaged nuclear areas or foci simultaneously but independently of each other. Third, Ub-FANCD2 and MDC1 are independently required for optimal BRCA1 relocalisation. While RPA32 phosphorylation (p-RPA32) and RPA foci formation were reduced in parallel with increasing levels of H2AX phosphorylation and MDC1 foci in UVC-irradiated FANC pathway-depleted cells, MDC1 depletion was associated with increased UVC-induced Ub-FANCD2 and FANCD2 foci as well as p-RPA32 levels and p-RPA32 foci. On the basis of the previous observations, we propose that the FANC pathway participates in the rescue of UVC-stalled replication forks in association with TLS by maintaining the integrity of ssDNA regions and by preserving genome stability and preventing the formation of DSBs, the resolution of which would require the intervention of MDC1. PMID:23365640

SPATA2 promotes CYLD activity and regulates TNF-induced NF-κB signaling and cell death.

PubMed

Schlicher, Lisa; Wissler, Manuela; Preiss, Florian; Brauns-Schubert, Prisca; Jakob, Celia; Dumit, Veronica; Borner, Christoph; Dengjel, Joern; Maurer, Ulrich

2016-10-01

K63- and Met1-linked ubiquitylation are crucial posttranslational modifications for TNF receptor signaling. These non-degradative ubiquitylations are counteracted by deubiquitinases (DUBs), such as the enzyme CYLD, resulting in an appropriate signal strength, but the regulation of this process remains incompletely understood. Here, we describe an interaction partner of CYLD, SPATA2, which we identified by a mass spectrometry screen. We find that SPATA2 interacts via its PUB domain with CYLD, while a PUB interaction motif (PIM) of SPATA2 interacts with the PUB domain of the LUBAC component HOIP SPATA2 is required for the recruitment of CYLD to the TNF receptor signaling complex upon TNFR stimulation. Moreover, SPATA2 acts as an allosteric activator for the K63- and M1-deubiquitinase activity of CYLD In consequence, SPATA2 substantially attenuates TNF-induced NF-κB and MAPK signaling. Conversely, SPATA2 is required for TNF-induced complex II formation, caspase activation, and apoptosis. Thus, this study identifies SPATA2 as an important factor in the TNF signaling pathway with a substantial role for the effects mediated by the cytokine. © 2016 The Authors.

Receptor-Like Kinase RUPO Interacts with Potassium Transporters to Regulate Pollen Tube Growth and Integrity in Rice

PubMed Central

Liu, Lingtong; Zheng, Canhui; Kuang, Baijan; Wei, Liqin; Yan, Longfeng; Wang, Tai

2016-01-01

During sexual reproduction of flowering plants, the pollen tube grows fast and over a long distance within the pistil to deliver two sperms for double fertilization. Growing plant cells need to communicate constantly with external stimuli as well as monitor changes in surface tension of the cell wall and plasma membrane to coordinate these signals and internal growth machinery; however, the underlying mechanisms remain largely unknown. Here we show that the rice member of plant-specific receptor-like kinase CrRLK1Ls subfamily, Ruptured Pollen tube (RUPO), is specifically expressed in rice pollen. RUPO localizes to the apical plasma membrane and vesicle of pollen tubes and is required for male gamete transmission. K+ levels were greater in pollen of homozygous CRISPR-knockout lines than wild-type plants, and pollen tubes burst shortly after germination. We reveal the interaction of RUPO with high-affinity potassium transporters. Phosphorylation of RUPO established and dephosphorylation abolished the interaction. These results have revealed the receptor-like kinase as a regulator of high-affinity potassium transporters via phosphorylation-dependent interaction, and demonstrated a novel receptor-like kinase signaling pathway that mediates K+ homeostasis required for pollen tube growth and integrity. PMID:27447945

Activation of PI3K, Akt, and ERK during early rotavirus infection leads to V-ATPase-dependent endosomal acidification required for uncoating

PubMed Central

Kim, Deok-Song; Kim, Ji-Yun; Park, Jun-Gyu; Alfajaro, Mia Madel; Baek, Yeong-Bin; Cho, Eun-Hyo; Kwon, Joseph; Choi, Jong-Soon; Kang, Mun-Il; Park, Sang-Ik; Cho, Kyoung-Oh

2018-01-01

The cellular PI3K/Akt and/or MEK/ERK signaling pathways mediate the entry process or endosomal acidification during infection of many viruses. However, their roles in the early infection events of group A rotaviruses (RVAs) have remained elusive. Here, we show that late-penetration (L-P) human DS-1 and bovine NCDV RVA strains stimulate these signaling pathways very early in the infection. Inhibition of both signaling pathways significantly reduced production of viral progeny due to blockage of virus particles in the late endosome, indicating that neither of the two signaling pathways is involved in virus trafficking. However, immunoprecipitation assays using antibodies specific for pPI3K, pAkt, pERK and the subunit E of the V-ATPase co-immunoprecipitated the V-ATPase in complex with pPI3K, pAkt, and pERK. Moreover, Duolink proximity ligation assay revealed direct association of the subunit E of the V-ATPase with the molecules pPI3K, pAkt, and pERK, indicating that both signaling pathways are involved in V-ATPase-dependent endosomal acidification. Acidic replenishment of the medium restored uncoating of the RVA strains in cells pretreated with inhibitors specific for both signaling pathways, confirming the above results. Isolated components of the outer capsid proteins, expressed as VP4-VP8* and VP4-VP5* domains, and VP7, activated the PI3K/Akt and MEK/ERK pathways. Furthermore, psoralen-UV-inactivated RVA and CsCl-purified RVA triple-layered particles triggered activation of the PI3K/Akt and MEK/ERK pathways, confirming the above results. Our data demonstrate that multistep binding of outer capsid proteins of L-P RVA strains with cell surface receptors phosphorylates PI3K, Akt, and ERK, which in turn directly interact with the subunit E of the V-ATPase to acidify the late endosome for uncoating of RVAs. This study provides a better understanding of the RVA-host interaction during viral uncoating, which is of importance for the development of strategies aiming at controlling or preventing RVA infections. PMID:29352319

Comparative study on gene set and pathway topology-based enrichment methods.

PubMed

Bayerlová, Michaela; Jung, Klaus; Kramer, Frank; Klemm, Florian; Bleckmann, Annalen; Beißbarth, Tim

2015-10-22

Enrichment analysis is a popular approach to identify pathways or sets of genes which are significantly enriched in the context of differentially expressed genes. The traditional gene set enrichment approach considers a pathway as a simple gene list disregarding any knowledge of gene or protein interactions. In contrast, the new group of so called pathway topology-based methods integrates the topological structure of a pathway into the analysis. We comparatively investigated gene set and pathway topology-based enrichment approaches, considering three gene set and four topological methods. These methods were compared in two extensive simulation studies and on a benchmark of 36 real datasets, providing the same pathway input data for all methods. In the benchmark data analysis both types of methods showed a comparable ability to detect enriched pathways. The first simulation study was conducted with KEGG pathways, which showed considerable gene overlaps between each other. In this study with original KEGG pathways, none of the topology-based methods outperformed the gene set approach. Therefore, a second simulation study was performed on non-overlapping pathways created by unique gene IDs. Here, methods accounting for pathway topology reached higher accuracy than the gene set methods, however their sensitivity was lower. We conducted one of the first comprehensive comparative works on evaluating gene set against pathway topology-based enrichment methods. The topological methods showed better performance in the simulation scenarios with non-overlapping pathways, however, they were not conclusively better in the other scenarios. This suggests that simple gene set approach might be sufficient to detect an enriched pathway under realistic circumstances. Nevertheless, more extensive studies and further benchmark data are needed to systematically evaluate these methods and to assess what gain and cost pathway topology information introduces into enrichment analysis. Both types of methods for enrichment analysis require further improvements in order to deal with the problem of pathway overlaps.

Redesigning metabolism based on orthogonality principles

PubMed Central

Pandit, Aditya Vikram; Srinivasan, Shyam; Mahadevan, Radhakrishnan

2017-01-01

Modifications made during metabolic engineering for overproduction of chemicals have network-wide effects on cellular function due to ubiquitous metabolic interactions. These interactions, that make metabolic network structures robust and optimized for cell growth, act to constrain the capability of the cell factory. To overcome these challenges, we explore the idea of an orthogonal network structure that is designed to operate with minimal interaction between chemical production pathways and the components of the network that produce biomass. We show that this orthogonal pathway design approach has significant advantages over contemporary growth-coupled approaches using a case study on succinate production. We find that natural pathways, fundamentally linked to biomass synthesis, are less orthogonal in comparison to synthetic pathways. We suggest that the use of such orthogonal pathways can be highly amenable for dynamic control of metabolism and have other implications for metabolic engineering. PMID:28555623

Determining the elastic properties of aptamer-ricin single molecule multiple pathway interactions

NASA Astrophysics Data System (ADS)

Wang, Bin; Park, Bosoon; Kwon, Yongkuk; Xu, Bingqian

2014-05-01

We report on the elastic properties of ricin and anti-ricin aptamer interactions, which showed three stable binding conformations, each of which has its special elastic properties. These different unbinding pathways were investigated by the dynamic force spectroscopy. A series-spring model combining the worm-like-chain model and Hook's law was used to estimate the apparent spring constants of the aptamer and linker molecule polyethylene glycol. The aptamer in its three different unbinding pathways showed different apparent spring constants. The two reaction barriers in the unbinding pathways also influence the apparent spring constant of the aptamer. This special elastic behavior of aptamer was used to distinguish its three unbinding pathways under different loading rates. This method also offered a way to distinguish and discard the non-specific interactions in single molecule experiments.

Nonlinear analysis of saccade speed fluctuations during combined action and perception tasks

PubMed Central

Stan, C.; Astefanoaei, C.; Pretegiani, E.; Optican, L.; Creanga, D.; Rufa, A.; Cristescu, C.P.

2014-01-01

Background: Saccades are rapid eye movements used to gather information about a scene which requires both action and perception. These are usually studied separately, so that how perception influences action is not well understood. In a dual task, where the subject looks at a target and reports a decision, subtle changes in the saccades might be caused by action-perception interactions. Studying saccades might provide insight into how brain pathways for action and for perception interact. New method: We applied two complementary methods, multifractal detrended fluctuation analysis and Lempel-Ziv complexity index to eye peak speed recorded in two experiments, a pure action task and a combined action-perception task. Results: Multifractality strength is significantly different in the two experiments, showing smaller values for dual decision task saccades compared to simple-task saccades. The normalized Lempel-Ziv complexity index behaves similarly i.e. is significantly smaller in the decision saccade task than in the simple task. Comparison with existing methods: Compared to the usual statistical and linear approaches, these analyses emphasize the character of the dynamics involved in the fluctuations and offer a sensitive tool for quantitative evaluation of the multifractal features and of the complexity measure in the saccades peak speeds when different brain circuits are involved. Conclusion: Our results prove that the peak speed fluctuations have multifractal characteristics with lower magnitude for the multifractality strength and for the complexity index when two neural pathways are simultaneously activated, demonstrating the nonlinear interaction in the brain pathways for action and perception. PMID:24854830

The receptor kinase CERK1 has dual functions in symbiosis and immunity signalling.

PubMed

Zhang, Xiaowei; Dong, Wentao; Sun, Jongho; Feng, Feng; Deng, Yiwen; He, Zuhua; Oldroyd, Giles E D; Wang, Ertao

2015-01-01

The establishment of symbiotic interactions between mycorrhizal fungi, rhizobial bacteria and their legume hosts involves a common symbiosis signalling pathway. This signalling pathway is activated by Nod factors produced by rhizobia and these are recognised by the Nod factor receptors NFR1/LYK3 and NFR5/NFP. Mycorrhizal fungi produce lipochitooligosaccharides (LCOs) similar to Nod factors, as well as short-chain chitin oligomers (CO4/5), implying commonalities in signalling during mycorrhizal and rhizobial associations. Here we show that NFR1/LYK3, but not NFR5/NFP, is required for the establishment of the mycorrhizal interaction in legumes. NFR1/LYK3 is necessary for the recognition of mycorrhizal fungi and the activation of the symbiosis signalling pathway leading to induction of calcium oscillations and gene expression. Chitin oligosaccharides also act as microbe associated molecular patterns that promote plant immunity via similar LysM receptor-like kinases. CERK1 in rice has the highest homology to NFR1 and we show that this gene is also necessary for the establishment of the mycorrhizal interaction as well as for resistance to the rice blast fungus. Our results demonstrate that NFR1/LYK3/OsCERK1 represents a common receptor for chitooligosaccharide-based signals produced by mycorrhizal fungi, rhizobial bacteria (in legumes) and fungal pathogens. It would appear that mycorrhizal recognition has been conserved in multiple receptors across plant species, but additional diversification in certain plant species has defined other signals that this class of receptors can perceive. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Interaction of AIP with protein kinase A (cAMP-dependent protein kinase).

PubMed

Schernthaner-Reiter, Marie Helene; Trivellin, Giampaolo; Stratakis, Constantine A

2018-05-02

Germline mutations in the aryl hydrocarbon receptor-interacting protein (AIP) gene cause mostly somatotropinomas and/or prolactinomas in a subset of familial isolated pituitary adenomas (FIPA). AIP has been shown to interact with phosphodiesterases (PDEs) and G proteins, suggesting a link to the cyclic AMP (cAMP)-dependent protein kinase (PKA) pathway. Upregulation of PKA is seen in sporadic somatotropinomas that carry GNAS1 mutations, and those in Carney complex that are due to PRKAR1A mutations. To elucidate the mechanism of AIP-dependent pituitary tumorigenesis, we studied potential functional and physical interactions of AIP with PKA's main subunits PRKAR1A (R1α) and PRKACA (Cα). We found that AIP physically interacts with both R1α and Cα; this interaction is enhanced when all three components are present, but maintained during Cα-R1α dissociation by PKA pathway activation, indicating that AIP binds Cα/R1α both in complex and separately. The interaction between AIP and R1α/Cα is reduced when the frequent AIP pathogenic mutation p.R304* is present. AIP protein levels are regulated both by translation and the ubiquitin/proteasome pathway and Cα stabilizes both AIP and R1α protein levels. AIP reduction by siRNA leads to an increase of PKA pathway activity, which is disproportionately enhanced during PDE4-inhibition. We show that AIP interacts with the PKA pathway on multiple levels, including a physical interaction with both the main regulatory (R1α) and catalytic (Cα) PKA subunits and a functional interaction with PDE4-dependent PKA activation. These findings provide novel insights on the mechanisms of AIP-dependent pituitary tumorigenesis.

Bacterial discrimination by Dictyostelid amoebae reveals the complexity of ancient interspecies interactions

PubMed Central

Nasser, Waleed; Santhanam, Balaji; Miranda, Edward Roshan; Parikh, Anup; Juneja, Kavina; Rot, Gregor; Dinh, Chris; Chen, Rui; Zupan, Blaz; Shaulsky, Gad; Kuspa, Adam

2014-01-01

Background Amoebae and bacteria interact within predator/prey and host/pathogen relationships, but the general response of amoeba to bacteria is not well understood. The amoeba Dictyostelium discoideum feeds on, and is colonized by diverse bacterial species including Gram-positive [Gram(+)] and Gram-negative [Gram(−)] bacteria, two major groups of bacteria that differ in structure and macromolecular composition. Results Transcriptional profiling of D. discoideum revealed sets of genes whose expression is enriched in amoebae interacting with different species of bacteria, including sets that appear specific to amoebae interacting with Gram(+), or with Gram(−) bacteria. In a genetic screen utilizing the growth of mutant amoebae on a variety of bacteria as a phenotypic readout, we identified amoebal genes that are only required for growth on Gram(+) bacteria, including one that encodes the cell surface protein gp130, as well as several genes that are only required for growth on Gram(−) bacteria including one that encodes a putative lysozyme, AlyL. These genes are required for parts of the transcriptional response of wild-type amoebae, and this allowed their classification into potential response pathways. Conclusions We have defined genes that are critical for amoebal survival during feeding on Gram(+), or Gram(−), bacteria which we propose form part of a regulatory network that allows D. discoideum to elicit specific cellular responses to different species of bacteria in order to optimize survival. PMID:23664307

Discovery of multiple interacting partners of gankyrin, a proteasomal chaperone and an oncoprotein--evidence for a common hot spot site at the interface and its functional relevance.

PubMed

Nanaware, Padma P; Ramteke, Manoj P; Somavarapu, Arun K; Venkatraman, Prasanna

2014-07-01

Gankyrin, a non-ATPase component of the proteasome and a chaperone of proteasome assembly, is also an oncoprotein. Gankyrin regulates a variety of oncogenic signaling pathways in cancer cells and accelerates degradation of tumor suppressor proteins p53 and Rb. Therefore gankyrin may be a unique hub integrating signaling networks with the degradation pathway. To identify new interactions that may be crucial in consolidating its role as an oncogenic hub, crystal structure of gankyrin-proteasome ATPase complex was used to predict novel interacting partners. EEVD, a four amino acid linear sequence seems a hot spot site at this interface. By searching for EEVD in exposed regions of human proteins in PDB database, we predicted 34 novel interactions. Eight proteins were tested and seven of them were found to interact with gankyrin. Affinity of four interactions is high enough for endogenous detection. Others require gankyrin overexpression in HEK 293 cells or occur endogenously in breast cancer cell line- MDA-MB-435, reflecting lower affinity or presence of a deregulated network. Mutagenesis and peptide inhibition confirm that EEVD is the common hot spot site at these interfaces and therefore a potential polypharmacological drug target. In MDA-MB-231 cells in which the endogenous CLIC1 is silenced, trans-expression of Wt protein (CLIC1_EEVD) and not the hot spot site mutant (CLIC1_AAVA) resulted in significant rescue of the migratory potential. Our approach can be extended to identify novel functionally relevant protein-protein interactions, in expansion of oncogenic networks and in identifying potential therapeutic targets. © 2013 Wiley Periodicals, Inc.

Modeling biological pathway dynamics with timed automata.

PubMed

Schivo, Stefano; Scholma, Jetse; Wanders, Brend; Urquidi Camacho, Ricardo A; van der Vet, Paul E; Karperien, Marcel; Langerak, Rom; van de Pol, Jaco; Post, Janine N

2014-05-01

Living cells are constantly subjected to a plethora of environmental stimuli that require integration into an appropriate cellular response. This integration takes place through signal transduction events that form tightly interconnected networks. The understanding of these networks requires capturing their dynamics through computational support and models. ANIMO (analysis of Networks with Interactive Modeling) is a tool that enables the construction and exploration of executable models of biological networks, helping to derive hypotheses and to plan wet-lab experiments. The tool is based on the formalism of Timed Automata, which can be analyzed via the UPPAAL model checker. Thanks to Timed Automata, we can provide a formal semantics for the domain-specific language used to represent signaling networks. This enforces precision and uniformity in the definition of signaling pathways, contributing to the integration of isolated signaling events into complex network models. We propose an approach to discretization of reaction kinetics that allows us to efficiently use UPPAAL as the computational engine to explore the dynamic behavior of the network of interest. A user-friendly interface hides the use of Timed Automata from the user, while keeping the expressive power intact. Abstraction to single-parameter kinetics speeds up construction of models that remain faithful enough to provide meaningful insight. The resulting dynamic behavior of the network components is displayed graphically, allowing for an intuitive and interactive modeling experience.

Plasmacytoid Dendritic Cells Require Direct Infection To Sustain the Pulmonary Influenza A Virus-Specific CD8 T Cell Response.

PubMed

Hemann, Emily A; Sjaastad, Louisa E; Langlois, Ryan A; Legge, Kevin L

2015-12-30

Following influenza A virus (IAV) infection, development of a robust IAV-specific CD8 T cell response is required for clearance of primary infection and enhances memory protection. Following IAV infection, plasmacytoid dendritic cells (pDC) or CD8α(+) DC regulate pulmonary effector CD8 T cell responses within the lung. Without this DC-T cell interaction, insufficient effector CD8 T cells are maintained in the lungs, leading to enhanced morbidity and mortality. Previous studies have demonstrated that pDC are capable of classical presentation or cross-presentation of IAV antigens and could potentially regulate IAV-specific CD8 T cell responses through either mechanism. Our results demonstrate that pDC from the lungs of donor mice infected with an IAV that is not able to replicate in hematopoietic cells (142t-IAV), unlike donor pDC isolated from the lungs of control infected mice, are not able to rescue the host IAV-specific CD8 T cell response from apoptosis. This indicates that pDC must utilize the direct presentation pathway for this rescue. This inability of pDC from 142t-IAV donors to rescue the IAV-specific CD8 T cell response is not due to differences in the overall ability of 142t-IAV to replicate within the lungs or generate defective viral genomes or to differences in levels of costimulatory molecules required for this interaction. We further demonstrate that bypassing the antigen presentation pathway by coating the 142t-IAV pDC with IAV peptide epitopes restores their ability to rescue the IAV-specific CD8 T cell response. IAV continues to be a global health burden, infecting 5 to 20% of the global population annually. Continued investigation into the mechanisms that mediate protective immune responses against IAV is important to improving current vaccination and antiviral strategies antagonistic toward IAV. Our findings presented herein demonstrate a key requirement for pDC promotion of effector CD8 T cell survival: that rather than utilizing cross-presentation, pDC must be infected and utilize the endogenous pathway for presentation of antigens to CD8 T cells during in vivo IAV infections. This suggests that targeting presentation via the endogenous pathway in pDC could be important for the development of unique antiviral cellular therapies. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Protein Kinase A Modulates Transforming Growth Factor-β Signaling through a Direct Interaction with Smad4 Protein*

PubMed Central

Yang, Huibin; Li, Gangyong; Wu, Jing-Jiang; Wang, Lidong; Uhler, Michael; Simeone, Diane M.

2013-01-01

Transforming growth factor β (TGFβ) signaling normally functions to regulate embryonic development and cellular homeostasis. It is increasingly recognized that TGFβ signaling is regulated by cross-talk with other signaling pathways. We previously reported that TGFβ activates protein kinase A (PKA) independent of cAMP through an interaction of an activated Smad3-Smad4 complex and the regulatory subunit of the PKA holoenzyme (PKA-R). Here we define the interaction domains of Smad4 and PKA-R and the functional consequences of this interaction. Using a series of Smad4 and PKA-R truncation mutants, we identified amino acids 290–300 of the Smad4 linker region as critical for the specific interaction of Smad4 and PKA-R. Co-immunoprecipitation assays showed that the B cAMP binding domain of PKA-R was sufficient for interaction with Smad4. Targeting of B domain regions conserved among all PKA-R isoforms and exposed on the molecular surface demonstrated that amino acids 281–285 and 320–329 were required for complex formation with Smad4. Interactions of these specific regions of Smad4 and PKA-R were necessary for TGFβ-mediated increases in PKA activity, CREB (cAMP-response element-binding protein) phosphorylation, induction of p21, and growth inhibition. Moreover, this Smad4-PKA interaction was required for TGFβ-induced epithelial mesenchymal transition, invasion of pancreatic tumor cells, and regulation of tumor growth in vivo. PMID:23362281

Interactions of the Human LIP5 Regulatory Protein with Endosomal Sorting Complexes Required for Transport*♦

PubMed Central

Skalicky, Jack J.; Arii, Jun; Wenzel, Dawn M.; Stubblefield, William-May B.; Katsuyama, Angela; Uter, Nathan T.; Bajorek, Monika; Myszka, David G.; Sundquist, Wesley I.

2012-01-01

The endosomal sorting complex required for transport (ESCRT) pathway remodels membranes during multivesicular body biogenesis, the abscission stage of cytokinesis, and enveloped virus budding. The ESCRT-III and VPS4 ATPase complexes catalyze the membrane fission events associated with these processes, and the LIP5 protein helps regulate their interactions by binding directly to a subset of ESCRT-III proteins and to VPS4. We have investigated the biochemical and structural basis for different LIP5-ligand interactions and show that the first microtubule-interacting and trafficking (MIT) module of the tandem LIP5 MIT domain binds CHMP1B (and other ESCRT-III proteins) through canonical type 1 MIT-interacting motif (MIM1) interactions. In contrast, the second LIP5 MIT module binds with unusually high affinity to a novel MIM element within the ESCRT-III protein CHMP5. A solution structure of the relevant LIP5-CHMP5 complex reveals that CHMP5 helices 5 and 6 and adjacent linkers form an amphipathic “leucine collar” that wraps almost completely around the second LIP5 MIT module but makes only limited contacts with the first MIT module. LIP5 binds MIM1-containing ESCRT-III proteins and CHMP5 and VPS4 ligands independently in vitro, but these interactions are coupled within cells because formation of stable VPS4 complexes with both LIP5 and CHMP5 requires LIP5 to bind both a MIM1-containing ESCRT-III protein and CHMP5. Our studies thus reveal how the tandem MIT domain of LIP5 binds different types of ESCRT-III proteins, promoting assembly of active VPS4 enzymes on the polymeric ESCRT-III substrate. PMID:23105106

Toward Omics-Based, Systems Biomedicine, and Path and Drug Discovery Methodologies for Depression-Inflammation Research.

PubMed

Maes, Michael; Nowak, Gabriel; Caso, Javier R; Leza, Juan Carlos; Song, Cai; Kubera, Marta; Klein, Hans; Galecki, Piotr; Noto, Cristiano; Glaab, Enrico; Balling, Rudi; Berk, Michael

2016-07-01

Meta-analyses confirm that depression is accompanied by signs of inflammation including increased levels of acute phase proteins, e.g., C-reactive protein, and pro-inflammatory cytokines, e.g., interleukin-6. Supporting the translational significance of this, a meta-analysis showed that anti-inflammatory drugs may have antidepressant effects. Here, we argue that inflammation and depression research needs to get onto a new track. Firstly, the choice of inflammatory biomarkers in depression research was often too selective and did not consider the broader pathways. Secondly, although mild inflammatory responses are present in depression, other immune-related pathways cannot be disregarded as new drug targets, e.g., activation of cell-mediated immunity, oxidative and nitrosative stress (O&NS) pathways, autoimmune responses, bacterial translocation, and activation of the toll-like receptor and neuroprogressive pathways. Thirdly, anti-inflammatory treatments are sometimes used without full understanding of their effects on the broader pathways underpinning depression. Since many of the activated immune-inflammatory pathways in depression actually confer protection against an overzealous inflammatory response, targeting these pathways may result in unpredictable and unwanted results. Furthermore, this paper discusses the required improvements in research strategy, i.e., path and drug discovery processes, omics-based techniques, and systems biomedicine methodologies. Firstly, novel methods should be employed to examine the intracellular networks that control and modulate the immune, O&NS and neuroprogressive pathways using omics-based assays, including genomics, transcriptomics, proteomics, metabolomics, epigenomics, immunoproteomics and metagenomics. Secondly, systems biomedicine analyses are essential to unravel the complex interactions between these cellular networks, pathways, and the multifactorial trigger factors and to delineate new drug targets in the cellular networks or pathways. Drug discovery processes should delineate new drugs targeting the intracellular networks and immune-related pathways.

The Hippo Pathway Regulates Homeostatic Growth of Stem Cell Niche Precursors in the Drosophila Ovary

PubMed Central

Sarikaya, Didem P.; Extavour, Cassandra G.

2015-01-01

The Hippo pathway regulates organ size, stem cell proliferation and tumorigenesis in adult organs. Whether the Hippo pathway influences establishment of stem cell niche size to accommodate changes in organ size, however, has received little attention. Here, we ask whether Hippo signaling influences the number of stem cell niches that are established during development of the Drosophila larval ovary, and whether it interacts with the same or different effector signaling pathways in different cell types. We demonstrate that canonical Hippo signaling regulates autonomous proliferation of the soma, while a novel hippo-independent activity of Yorkie regulates autonomous proliferation of the germ line. Moreover, we demonstrate that Hippo signaling mediates non-autonomous proliferation signals between germ cells and somatic cells, and contributes to maintaining the correct proportion of these niche precursors. Finally, we show that the Hippo pathway interacts with different growth pathways in distinct somatic cell types, and interacts with EGFR and JAK/STAT pathways to regulate non-autonomous proliferation of germ cells. We thus provide evidence for novel roles of the Hippo pathway in establishing the precise balance of soma and germ line, the appropriate number of stem cell niches, and ultimately regulating adult female reproductive capacity. PMID:25643260

Genomic and Transcriptomic Analyses of Indole-3-Acetic Acid Biosynthesis in Diatoms

NASA Astrophysics Data System (ADS)

Lim, R.; Armbrust, V.

2016-02-01

Indole-3-acetic acid (IAA) is a major plant growth hormone and a common mediator of plant-bacterial interactions. Recently, IAA has also been found to play a role in interactions between diatoms and bacteria, with IAA production by an associated Sulfitobacter leading to increased growth rates in the marine diatom Pseudo-nitzschia multiseries. It is unclear, however, if diatoms themselves are able to synthesize IAA and whether this capability is widespread throughout Bacillariophyta. Four major tryptophan-dependent IAA biosynthesis pathways have been identified in plants and bacteria, each denoted by the first intermediate downstream of tryptophan: the indole-3-pyruvate (IPyA), tryptamine (TAM), indole-3-acetaldoxime (IAOx) and indole-3-acetamide (IAM) pathways. To investigate the possibility of IAA biosynthesis in diatoms, we first analyzed publicly available genomes of raphid pennates P. multiseries, Phaeodactylum tricornutum, Fragilariopsis cylindrus and centric Thalassiosira pseudonana for potential homologs to plant and bacterial IAA biosynthesis genes. The P. multiseries, F. cylindrus and P. tricornutum genomes encode downstream enzymes for bacterial TAM and IAM and plant IPyA pathways. The more evolutionarily ancient T. pseudonana encodes one TAM enzyme in its genome. To investigate the potential distribution of these pathways more broadly, we surveyed the transcriptomes of 11 diatom species that include representatives from all four Bacillariophyta classes. Datasets used were sequenced as part of the Marine Microbial Eukaryote Transcriptome Sequencing Project (MMETSP) and obtained from cultures maintained axenically. Transcripts associated with the TAM pathway were most frequently detected, with potential homologs to required enzymes identified in 10 of the 11 species examined. Transcripts homologous to rate-limiting IPyA enzymes were detected in six species. Only two centric and araphid pennate species expressed transcripts associated with enzymes in the IAM and IAOx pathways. This pattern suggests multiple events of gene loss as the phylum expanded and diversified. Mass spectrometry analyses will be conducted to confirm the production of IAA in axenic cultures of P. pungens, P. multistriata, Skeletonema marinoi and F. cylindrus.

Predictive Genomic Analyses Inform the Basis for Vitamin Metabolism and Provisioning in Bacteria-Arthropod Endosymbioses

PubMed Central

Serbus, Laura R.; Rodriguez, Brian Garcia; Sharmin, Zinat; Momtaz, A. J. M. Zehadee; Christensen, Steen

2017-01-01

The requirement of vitamins for core metabolic processes creates a unique set of pressures for arthropods subsisting on nutrient-limited diets. While endosymbiotic bacteria carried by arthropods have been widely implicated in vitamin provisioning, the underlying molecular mechanisms are not well understood. To address this issue, standardized predictive assessment of vitamin metabolism was performed in 50 endosymbionts of insects and arachnids. The results predicted that arthropod endosymbionts overall have little capacity for complete de novo biosynthesis of conventional or active vitamin forms. Partial biosynthesis pathways were commonly predicted, suggesting a substantial role in vitamin provisioning. Neither taxonomic relationships between host and symbiont, nor the mode of host-symbiont interaction were clear predictors of endosymbiont vitamin pathway capacity. Endosymbiont genome size and the synthetic capacity of nonsymbiont taxonomic relatives were more reliable predictors. We developed a new software application that also predicted that last-step conversion of intermediates into active vitamin forms may contribute further to vitamin biosynthesis by endosymbionts. Most instances of predicted vitamin conversion were paralleled by predictions of vitamin use. This is consistent with achievement of provisioning in some cases through upregulation of pathways that were retained for endosymbiont benefit. The predicted absence of other enzyme classes further suggests a baseline of vitamin requirement by the majority of endosymbionts, as well as some instances of putative mutualism. Adaptation of this workflow to analysis of other organisms and metabolic pathways will provide new routes for considering the molecular basis for symbiosis on a comprehensive scale. PMID:28455417

Novel mechanism of JNK pathway activation by adenoviral E1A

PubMed Central

Morrison, Helen; Pospelova, Tatiana V.; Pospelov, Valery A.; Herrlich, Peter

2014-01-01

The adenoviral oncoprotein E1A influences cellular regulation by interacting with a number of cellular proteins. In collaboration with complementary oncogenes, E1A fully transforms primary cells. As part of this action, E1A inhibits transcription of c-Jun:Fos target genes while promoting that of c-Jun:ATF2-dependent genes including jun. Both c-Jun and ATF2 are hyperphosphorylated in response to E1A. In the current study, E1A was fused with the ligand binding domain of the estrogen receptor (E1A-ER) to monitor the immediate effect of E1A activation. With this approach we now show that E1A activates c-Jun N-terminal kinase (JNK), the upstream kinases MKK4 and MKK7, as well as the small GTPase Rac1. Activation of the JNK pathway requires the N-terminal domain of E1A, and, importantly, is independent of transcription. In addition, it requires the presence of ERM proteins. Downregulation of signaling components upstream of JNK inhibits E1A-dependent JNK/c-Jun activation. Taking these findings together, we show that E1A activates the JNK/c-Jun signaling pathway upstream of Rac1 in a transcription-independent manner, demonstrating a novel mechanism of E1A action. PMID:24742962

The structure of a ring-opened proliferating cell nuclear antigen-replication factor C complex revealed by fluorescence energy transfer.

PubMed

Zhuang, Zhihao; Yoder, Bonita L; Burgers, Peter M J; Benkovic, Stephen J

2006-02-21

Numerous proteins that function in DNA metabolic pathways are known to interact with the proliferating cell nuclear antigen (PCNA). The important function of PCNA in stimulating various cellular activities requires its topological linkage with DNA. Loading of the circular PCNA onto duplex DNA requires the activity of a clamp-loader [replication factor C (RFC)] complex and the energy derived from ATP hydrolysis. The mechanistic and structural details regarding PCNA loading by the RFC complex are still developing. In particular, the positive identification of a long-hypothesized structure of an open clamp-RFC complex as an intermediate in loading has remained elusive. In this study, we capture an open yeast PCNA clamp in a complex with RFC through fluorescence energy transfer experiments. We also follow the topological transitions of PCNA in the various steps of the clamp-loading pathway through both steady-state and stopped-flow fluorescence studies. We find that ATP effectively drives the clamp-loading process to completion with the formation of the closed PCNA bound to DNA, whereas ATPgammaS cannot. The information derived from this work complements that obtained from previous structural and mechanistic studies and provides a more complete picture of a eukaryotic clamp-loading pathway using yeast as a paradigm.

Interactions between late acting proteins required for peptidoglycan synthesis during sporulation

PubMed Central

Fay, Allison; Meyer, Pablo; Dworkin, Jonathan

2010-01-01

The requirement of peptidoglycan synthesis for growth complicates the analysis of interactions between proteins involved in this pathway. In particular, the later steps that involve membrane-linked substrates have proven largely recalcitrant to in vivo analysis. Here we have taken advantage of the peptidoglycan synthesis that occurs during sporulation in Bacillus subtilis to examine the interactions between SpoVE, a non-essential, sporulation-specific homolog of the well-conserved and essential SEDS proteins, and SpoVD, a non-essential class B penicillin binding protein (PBP). We found that localization of SpoVD is dependent on SpoVE and that SpoVD protects SpoVE from in vivo proteolysis. Co-immunoprecipitations and Fluorescence Resonance Energy Transfer experiments indicated that SpoVE and SpoVD interact and co-affinity purification in E. coli demonstrated that this interaction is direct. Finally, we generated a functional protein consisting of a SpoVE-SpoVD fusion and found that a loss-of-function point mutation in either part of the fusion resulted in a loss of function of the entire fusion that was not complemented by a wild type protein. Thus, SpoVE has a direct and functional interaction with SpoVD and this conclusion will facilitate understanding the essential function SpoVE and related SEDS proteins such as FtsW and RodA play in bacterial growth and division. PMID:20417640

E2 Proteins from High- and Low-Risk Human Papillomavirus Types Differ in Their Ability To Bind p53 and Induce Apoptotic Cell Death

PubMed Central

Parish, Joanna L.; Kowalczyk, Anna; Chen, Hsin-Tien; Roeder, Geraldine E.; Sessions, Richard; Buckle, Malcolm; Gaston, Kevin

2006-01-01

The E2 proteins from oncogenic (high-risk) human papillomaviruses (HPVs) can induce apoptotic cell death in both HPV-transformed and non-HPV-transformed cells. Here we show that the E2 proteins from HPV type 6 (HPV6) and HPV11, two nononcogenic (low-risk) HPV types, fail to induce apoptosis. Unlike the high-risk HPV16 E2 protein, these low-risk E2 proteins fail to bind p53 and fail to induce p53-dependent transcription activation. Interestingly, neither the ability of p53 to activate transcription nor the ability of p53 to bind DNA, are required for HPV16 E2-induced apoptosis in non-HPV-transformed cells. However, mutations that reduce the binding of the HPV16 E2 protein to p53 inhibit E2-induced apoptosis in non-HPV-transformed cells. In contrast, the interaction between HPV16 E2 and p53 is not required for this E2 protein to induce apoptosis in HPV-transformed cells. Thus, our data suggest that this high-risk HPV E2 protein induces apoptosis via two pathways. One pathway involves the binding of E2 to p53 and can operate in both HPV-transformed and non-HPV-transformed cells. The second pathway requires the binding of E2 to the viral genome and can only operate in HPV-transformed cells. PMID:16611918

Phosphatidic acid induces EHD3-containing membrane tubulation and is required for receptor recycling.

PubMed

Henmi, Yuji; Oe, Natsuko; Kono, Nozomu; Taguchi, Tomohiko; Takei, Kohji; Tanabe, Kenji

2016-03-01

EHD3 is localized on the tubular structures of early endosomes, and it regulates their trafficking pathway. However, the regulatory mechanism of EHD3-containing tubular structures remains poorly understood. An in vitro liposome co-sedimentation assay revealed that EHD3 interacted with phosphatidic acid through its helical domain and this interaction induced liposomal tubulations. Additionally, inhibiting phosphatidic acid synthesis with diacylglycerol kinase inhibitor or lysophosphatidic acid acyltransferase inhibitor significantly reduced the number of EHD3-containing tubules and impaired their trafficking from early endosomes. These results suggest that EHD3 and phosphatidic acid cooperatively regulate membrane deformation and trafficking from early endosomes. Copyright © 2016 Elsevier Inc. All rights reserved.

SPAK kinase is a substrate and target of PKCθ in T-cell receptor-induced AP-1 activation pathway

PubMed Central

Li, Yingqiu; Hu, Junru; Vita, Randi; Sun, Binggang; Tabata, Hiroki; Altman, Amnon

2004-01-01

Protein kinase C-θ (PKCθ) plays an important role in T-cell activation via stimulation of AP-1 and NF-κB. Here we report the isolation of SPAK, a Ste20-related upstream mitogen-activated protein kinase (MAPK), as a PKCθ-interacting kinase. SPAK interacted with PKCθ (but not with PKCα) via its 99 COOH-terminal residues. TCR/CD28 costimulation enhanced this association and stimulated the catalytic activity of SPAK. Recombinant SPAK was phosphorylated on Ser-311 in its kinase domain by PKCθ, but not by PKCα. The magnitude and duration of TCR/CD28-induced endogenous SPAK activation were markedly impaired in PKCθ-deficient T cells. Transfected SPAK synergized with constitutively active PKCθ to activate AP-1, but not NF-κB. This synergistic activity, as well as the receptor-induced SPAK activation, required the PKCθ-interacting region of SPAK, and Ser-311 mutation greatly reduced these activities of SPAK. Conversely, a SPAK-specific RNAi or a dominant-negative SPAK mutant inhibited PKCθ- and TCR/CD28-induced AP-1, but not NF-κB, activation. These results define SPAK as a substrate and target of PKCθ in a TCR/CD28-induced signaling pathway leading selectively to AP-1 (but not NF-κB) activation. PMID:14988727

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.

Hyaluronan, CD44, and Emmprin Regulate Lactate Efflux and Membrane Localization of Monocarboxylate Transporters in Human Breast Carcinoma Cells

PubMed Central

Slomiany, Mark G.; Grass, G. Daniel; Robertson, Angela D.; Yang, Xiao Y.; Maria, Bernard L.; Beeson, Craig; Toole, Bryan P.

2013-01-01

Interactions of hyaluronan with CD44 in tumor cells play important cooperative roles in various aspects of malignancy and drug resistance. Emmprin (CD147; basigin)is a cell surface glycoprotein of the immunoglobulin superfamily that is highly up-regulated in malignant cancer cells and stimulates hyaluronan production, as well as several downstream signaling pathways. Emmprin also interacts with various monocarboxylate transporters (MCT). Malignant cancer cells use the glycolytic pathway and require MCTs to efflux lactate that results from glycolysis. Glycolysis and lactate secretion contribute to malignant cell behaviors and drug resistance in tumor cells. In the present study, we find that perturbation of endogenous hyaluronan, using small hyaluronan oligosaccharides, rapidly inhibits lactate efflux from breast carcinoma cells; down-regulation of emmprin, using emmprin small interfering RNA, also results in decreased efflux. In addition, we find that CD44 coimmunoprecipitates with MCT1, MCT4, and emmprin and colocalizes with these proteins at the plasma membrane. Moreover, after treatment of the cells with hyaluronan oligosaccharides, CD44, MCT1, and MCT4 become localized intracellularly whereas emmprin remains at the cell membrane. Together, these data indicate that constitutive interactions among hyaluronan, CD44, and emmprin contribute to regulation of MCT localization and function in the plasma membrane of breast carcinoma cells. PMID:19176383

The CD63-Syntenin-1 Complex Controls Post-Endocytic Trafficking of Oncogenic Human Papillomaviruses.

PubMed

Gräßel, Linda; Fast, Laura Aline; Scheffer, Konstanze D; Boukhallouk, Fatima; Spoden, Gilles A; Tenzer, Stefan; Boller, Klaus; Bago, Ruzica; Rajesh, Sundaresan; Overduin, Michael; Berditchevski, Fedor; Florin, Luise

2016-08-31

Human papillomaviruses enter host cells via a clathrin-independent endocytic pathway involving tetraspanin proteins. However, post-endocytic trafficking required for virus capsid disassembly remains unclear. Here we demonstrate that the early trafficking pathway of internalised HPV particles involves tetraspanin CD63, syntenin-1 and ESCRT-associated adaptor protein ALIX. Following internalisation, viral particles are found in CD63-positive endosomes recruiting syntenin-1, a CD63-interacting adaptor protein. Electron microscopy and immunofluorescence experiments indicate that the CD63-syntenin-1 complex controls delivery of internalised viral particles to multivesicular endosomes. Accordingly, infectivity of high-risk HPV types 16, 18 and 31 as well as disassembly and post-uncoating processing of viral particles was markedly suppressed in CD63 or syntenin-1 depleted cells. Our analyses also present the syntenin-1 interacting protein ALIX as critical for HPV infection and CD63-syntenin-1-ALIX complex formation as a prerequisite for intracellular transport enabling viral capsid disassembly. Thus, our results identify the CD63-syntenin-1-ALIX complex as a key regulatory component in post-endocytic HPV trafficking.

Role of the PDZ-scaffold protein NHERF1/EBP50 in cancer biology: from signaling regulation to clinical relevance.

PubMed

Vaquero, J; Nguyen Ho-Bouldoires, T H; Clapéron, A; Fouassier, L

2017-06-01

The transmission of cellular information requires fine and subtle regulation of proteins that need to interact in a coordinated and specific way to form efficient signaling networks. The spatial and temporal coordination relies on scaffold proteins. Thanks to protein interaction domains such as PDZ domains, scaffold proteins organize multiprotein complexes enabling the proper transmission of cellular information through intracellular networks. NHERF1/EBP50 is a PDZ-scaffold protein that was initially identified as an organizer and regulator of transporters and channels at the apical side of epithelia through actin-binding ezrin-moesin-radixin proteins. Since, NHERF1/EBP50 has emerged as a major regulator of cancer signaling network by assembling cancer-related proteins. The PDZ-scaffold EBP50 carries either anti-tumor or pro-tumor functions, two antinomic functions dictated by EBP50 expression or subcellular localization. The dual function of NHERF1/EBP50 encompasses the regulation of several major signaling pathways engaged in cancer, including the receptor tyrosine kinases PDGFR and EGFR, PI3K/PTEN/AKT and Wnt-β-catenin pathways.

Expression of bone morphogenetic proteins and Msx genes during root formation.

PubMed

Yamashiro, T; Tummers, M; Thesleff, I

2003-03-01

Like crown development, root formation is also regulated by interactions between epithelial and mesenchymml tissues. Bone morphogenetic proteins (BMPs), together with the transcription factors Msx1 and Msx2, play important roles in these interactions during early tooth morphogenesis. To investigate the involvement of this signaling pathway in root development, we analyzed the expression patterns of Bmp2, Bmp3, Bmp4, and Bmp7 as well as Msx1 and Msx2 in the roots of mouse molars. Bmp4 was expressed in the apical mesenchyme and Msx2 in the root sheath. However, Bmps were not detected in the root sheath epithelium, and Msx transcripts were absent from the underlying mesenchyme. These findings indicate that this Bmp signaling pathway, required for tooth initiation, does not regulate root development, but we suggest that root shape may be regulated by a mechanism similar to that regulating crown shape in cap-stage tooth germs. Msx2 expression continued in the epithelial cell rests of Malassez, and the nearby cementoblasts intensely expressed Bmp3, which may regulate some functions of the fragmented epithelium.

Partially overlapping distribution of epsin1 and HIP1 at the synapse: analysis by immunoelectron microscopy.

PubMed

Yao, Pamela J; Bushlin, Ittai; Petralia, Ronald S

2006-01-10

Synapses of neurons use clathrin-mediated endocytic pathways for recycling of synaptic vesicles and trafficking of neurotransmitter receptors. Epsin 1 and huntingtin-interacting protein 1 (HIP1) are endocytic accessory proteins. Both proteins interact with clathrin and the AP2 adaptor complex and also bind to the phosphoinositide-containing plasma membrane via an epsin/AP180 N-terminal homology (ENTH/ANTH) domain. Epsin1 and HIP1 are found in neurons; however, their precise roles in synapses remain largely unknown. Using immunogold electron microscopy, we examine and compare the synaptic distribution of epsin1 and HIP1 in rat CA1 hippocampal synapse. We find that epsin1 is located across both sides of the synapse, whereas HIP1 displays a preference for the postsynaptic compartment. Within the synaptic compartments, espin1 is distributed similarly throughout, whereas postsynaptic HIP1 is concentrated near the plasma membrane. Our results suggest a dual role for epsin1 and HIP1 in the synapse: as broadly required factors for promoting clathrin assembly and as adaptors for specific endocytic pathways.

Human Sirtuin 2 Localization, Transient Interactions, and Impact on the Proteome Point to Its Role in Intracellular Trafficking.

PubMed

Budayeva, Hanna G; Cristea, Ileana M

2016-10-01

Human sirtuin 2 (SIRT2) is an NAD + -dependent deacetylase that primarily functions in the cytoplasm, where it can regulate α-tubulin acetylation levels. SIRT2 is linked to cancer progression, neurodegeneration, and infection with bacteria or viruses. However, the current knowledge about its interactions and the means through which it exerts its functions has remained limited. Here, we aimed to gain a better understanding of its cellular functions by characterizing SIRT2 subcellular localization, the identity and relative stability of its protein interactions, and its impact on the proteome of primary human fibroblasts. To assess the relative stability of SIRT2 interactions, we used immunoaffinity purification in conjunction with both label-free and metabolic labeling quantitative mass spectrometry. In addition to the expected associations with cytoskeleton proteins, including its known substrate TUBA1A, our results reveal that SIRT2 specifically interacts with proteins functioning in membrane trafficking, secretory processes, and transcriptional regulation. By quantifying their relative stability, we found most interactions to be transient, indicating a dynamic SIRT2 environment. We discover that SIRT2 localizes to the ER-Golgi intermediate compartment (ERGIC), and that this recruitment requires an intact ER-Golgi trafficking pathway. Further expanding these findings, we used microscopy and interaction assays to establish the interaction and coregulation of SIRT2 with liprin-β1 scaffolding protein (PPFiBP1), a protein with roles in focal adhesions disassembly. As SIRT2 functions may be accomplished via interactions, enzymatic activity, and transcriptional regulation, we next assessed the impact of SIRT2 levels on the cellular proteome. SIRT2 knockdown led to changes in the levels of proteins functioning in membrane trafficking, including some of its interaction partners. Altogether, our study expands the knowledge of SIRT2 cytoplasmic functions to define a previously unrecognized involvement in intracellular trafficking pathways, which may contribute to its roles in cellular homeostasis and human diseases. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Human Sirtuin 2 Localization, Transient Interactions, and Impact on the Proteome Point to Its Role in Intracellular Trafficking*

PubMed Central

Budayeva, Hanna G.; Cristea, Ileana M.

2016-01-01

Human sirtuin 2 (SIRT2) is an NAD+-dependent deacetylase that primarily functions in the cytoplasm, where it can regulate α-tubulin acetylation levels. SIRT2 is linked to cancer progression, neurodegeneration, and infection with bacteria or viruses. However, the current knowledge about its interactions and the means through which it exerts its functions has remained limited. Here, we aimed to gain a better understanding of its cellular functions by characterizing SIRT2 subcellular localization, the identity and relative stability of its protein interactions, and its impact on the proteome of primary human fibroblasts. To assess the relative stability of SIRT2 interactions, we used immunoaffinity purification in conjunction with both label-free and metabolic labeling quantitative mass spectrometry. In addition to the expected associations with cytoskeleton proteins, including its known substrate TUBA1A, our results reveal that SIRT2 specifically interacts with proteins functioning in membrane trafficking, secretory processes, and transcriptional regulation. By quantifying their relative stability, we found most interactions to be transient, indicating a dynamic SIRT2 environment. We discover that SIRT2 localizes to the ER-Golgi intermediate compartment (ERGIC), and that this recruitment requires an intact ER-Golgi trafficking pathway. Further expanding these findings, we used microscopy and interaction assays to establish the interaction and coregulation of SIRT2 with liprin-β1 scaffolding protein (PPFiBP1), a protein with roles in focal adhesions disassembly. As SIRT2 functions may be accomplished via interactions, enzymatic activity, and transcriptional regulation, we next assessed the impact of SIRT2 levels on the cellular proteome. SIRT2 knockdown led to changes in the levels of proteins functioning in membrane trafficking, including some of its interaction partners. Altogether, our study expands the knowledge of SIRT2 cytoplasmic functions to define a previously unrecognized involvement in intracellular trafficking pathways, which may contribute to its roles in cellular homeostasis and human diseases. PMID:27503897

The Hog1 Mitogen-Activated Protein Kinase Mediates a Hypoxic Response in Saccharomyces cerevisiae

PubMed Central

Hickman, Mark J.; Spatt, Dan; Winston, Fred

2011-01-01

We have studied hypoxic induction of transcription by studying the seripauperin (PAU) genes of Saccharomyces cerevisiae. Previous studies showed that PAU induction requires the depletion of heme and is dependent upon the transcription factor Upc2. We have now identified additional factors required for PAU induction during hypoxia, including Hog1, a mitogen-activated protein kinase (MAPK) whose signaling pathway originates at the membrane. Our results have led to a model in which heme and ergosterol depletion alters membrane fluidity, thereby activating Hog1 for hypoxic induction. Hypoxic activation of Hog1 is distinct from its previously characterized response to osmotic stress, as the two conditions cause different transcriptional consequences. Furthermore, Hog1-dependent hypoxic activation is independent of the S. cerevisiae general stress response. In addition to Hog1, specific components of the SAGA coactivator complex, including Spt20 and Sgf73, are also required for PAU induction. Interestingly, the mammalian ortholog of Spt20, p38IP, has been previously shown to interact with the mammalian ortholog of Hog1, p38. Taken together, our results have uncovered a previously unknown hypoxic-response pathway that may be conserved throughout eukaryotes. PMID:21467572

Chromatoid Body Protein TDRD6 Supports Long 3’ UTR Triggered Nonsense Mediated mRNA Decay

PubMed Central

Fanourgakis, Grigorios; Akpinar, Müge; Dahl, Andreas; Jessberger, Rolf

2016-01-01

Chromatoid bodies (CBs) are spermiogenesis-specific organelles of largely unknown function. CBs harbor various RNA species, RNA-associated proteins and proteins of the tudor domain family like TDRD6, which is required for a proper CB architecture. Proteome analysis of purified CBs revealed components of the nonsense-mediated mRNA decay (NMD) machinery including UPF1. TDRD6 is essential for UPF1 localization to CBs, for UPF1-UPF2 and UPF1-MVH interactions. Upon removal of TDRD6, the association of several mRNAs with UPF1 and UPF2 is disturbed, and the long 3’ UTR-stimulated but not the downstream exon-exon junction triggered pathway of NMD is impaired. Reduced association of the long 3’ UTR mRNAs with UPF1 and UPF2 correlates with increased stability and enhanced translational activity. Thus, we identified TDRD6 within CBs as required for mRNA degradation, specifically the extended 3’ UTR-triggered NMD pathway, and provide evidence for the requirement of NMD in spermiogenesis. This function depends on TDRD6-promoted assembly of mRNA and decay enzymes in CBs. PMID:27149095

Neurofibromin interacts with CRMP-2 and CRMP-4 in rat brain

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

Lin, Y.-L.; Hsueh, Y.-P., E-mail: yph@gate.sinica.edu.tw

Neurofibromin, encoded by the neurofibromatosis type 1 (NF1) gene, regulates the Ras and cAMP pathways and plays a role in proliferation and neuronal morphogenesis. The details of the molecular mechanism of neurofibromin action in these processes are still unclear. In this study, immunoprecipitation and proteomics were used to identify novel proteins from rat brain that interact with neurofibromin. Mass spectrometry analysis showed that two proteins, the collapsin response mediator protein-2 (CRMP-2) and propionyl-CoA carboxylase alpha chain (PCCA), associated with neurofibromin. Immunoprecipitation-immunoblotting analysis confirmed the interactions between neurofibromin and CRMP-2 and CRMP-4, but not CRMP-1, in rat brain. CDK5, a kinasemore » that regulates CRMP-2 in axonal outgrowth, was required for the interaction between neurofibromin and CRMP-2. Since both neurofibromin and CRMP proteins are involved in proliferation and axonal morphogenesis, these results suggest that the interaction with CRMPs contributes to the function of neurofibromin in tumorigenesis and neuronal morphogenesis.« less

Inferring molecular interactions pathways from eQTL data

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

Rashid, Imran; McDermott, Jason E.; Samudrala, Ram

Analysis of expression quantitative trait loci (eQTL) helps elucidate the connection between genotype, gene expression levels, and phenotype. However, standard statistical genetics can only attribute changes in expression levels to loci on the genome, not specific genes. Each locus can contain many genes, making it very difficult to discover which gene is controlling the expression levels of other genes. Furthermore, it is even more difficult to find a pathway of molecular interactions responsible for controlling the expression levels. Here we describe a series of techniques for finding explanatory pathways by exploring graphs of molecular interactions. We show several simple methodsmore » can find complete pathways the explain the mechanism of differential expression in eQTL data.« less

RIOK3 Is an Adaptor Protein Required for IRF3-Mediated Antiviral Type I Interferon Production

PubMed Central

Feng, Jun; De Jesus, Paul D.; Su, Victoria; Han, Stephanie; Gong, Danyang; Wu, Nicholas C.; Tian, Yuan; Li, Xudong; Wu, Ting-Ting; Chanda, Sumit K.

2014-01-01

ABSTRACT Detection of cytosolic nucleic acids by pattern recognition receptors leads to the induction of type I interferons (IFNs) and elicits the innate immune response. We report here the identification of RIOK3 as a novel adaptor protein that is essential for the cytosolic nucleic acid-induced type I IFN production and for the antiviral response to gammaherpesvirus through two independent kinome-wide RNA interference screens. RIOK3 knockdown blocks both cytosolic double-stranded B-form DNA and double-stranded RNA-induced IRF3 activation and IFN-β production. In contrast, the overexpression of RIOK3 activates IRF3 and induces IFN-β. RIOK3 functions downstream of TBK1 and upstream of IRF3 activation. Furthermore, RIOK3 physically interacts with both IRF3 and TBK1 and is necessary for the interaction between TBK1 and IRF3. In addition, global transcriptome analysis shows that the expression of many gene involved antiviral responses is dependent on RIOK3. Thus, knockdown of RIOK3 inhibits cellular antiviral responses against both DNA and RNA viruses (herpesvirus and influenza A virus). Our data suggest that RIOK3 plays a critical role in the antiviral type I IFN pathway by bridging TBK1 and IRF3. IMPORTANCE The innate immune response, such as the production of type I interferons, acts as the first line of defense, limiting infectious pathogens directly and shaping the adaptive immune response. In this study, we identified RIOK3 as a novel regulator of the antiviral type I interferon pathway. Specifically, we found that RIOK3 physically interacts with TBK1 and IRF3 and bridges the functions between TBK1 and IRF3 in the activation of type I interferon pathway. The identification of a cellular kinase that plays a role the type I interferon pathway adds another level of complexity in the regulation of innate immunity and will have implications for developing novel strategies to combat viral infection. PMID:24807708

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.

Examination of association to autism of common genetic variationin genes related to dopamine.

PubMed

Anderson, B M; Schnetz-Boutaud, N; Bartlett, J; Wright, H H; Abramson, R K; Cuccaro, M L; Gilbert, J R; Pericak-Vance, M A; Haines, J L

2008-12-01

Autism is a severe neurodevelopmental disorder characterized by a triad of complications. Autistic individuals display significant disturbances in language and reciprocal social interactions, combined with repetitive and stereotypic behaviors. Prevalence studies suggest that autism is more common than originally believed, with recent estimates citing a rate of one in 150. Although multiple genetic linkage and association studies have yielded multiple suggestive genes or chromosomal regions, a specific risk locus has yet to be identified and widely confirmed. Because many etiologies have been suggested for this complex syndrome, we hypothesize that one of the difficulties in identifying autism genes is that multiple genetic variants may be required to significantly increase the risk of developing autism. Thus, we took the alternative approach of examining 14 prominent dopamine pathway candidate genes for detailed study by genotyping 28 single nucleotide polymorphisms. Although we did observe a nominally significant association for rs2239535 (P=0.008) on chromosome 20, single-locus analysis did not reveal any results as significant after correction for multiple comparisons. No significant interaction was identified when Multifactor Dimensionality Reduction was employed to test specifically for multilocus effects. Although genome-wide linkage scans in autism have provided support for linkage to various loci along the dopamine pathway, our study does not provide strong evidence of linkage or association to any specific gene or combination of genes within the pathway. These results demonstrate that common genetic variation within the tested genes located within this pathway at most play a minor to moderate role in overall autism pathogenesis.

Disruption of Axonal Transport Perturbs Bone Morphogenetic Protein (BMP) - Signaling and Contributes to Synaptic Abnormalities in Two Neurodegenerative Diseases

PubMed Central

Kang, Min Jung; Hansen, Timothy J.; Mickiewicz, Monique; Kaczynski, Tadeusz J.; Fye, Samantha; Gunawardena, Shermali

2014-01-01

Formation of new synapses or maintenance of existing synapses requires the delivery of synaptic components from the soma to the nerve termini via axonal transport. One pathway that is important in synapse formation, maintenance and function of the Drosophila neuromuscular junction (NMJ) is the bone morphogenetic protein (BMP)-signaling pathway. Here we show that perturbations in axonal transport directly disrupt BMP signaling, as measured by its downstream signal, phospho Mad (p-Mad). We found that components of the BMP pathway genetically interact with both kinesin-1 and dynein motor proteins. Thick vein (TKV) vesicle motility was also perturbed by reductions in kinesin-1 or dynein motors. Interestingly, dynein mutations severely disrupted p-Mad signaling while kinesin-1 mutants showed a mild reduction in p-Mad signal intensity. Similar to mutants in components of the BMP pathway, both kinesin-1 and dynein motor protein mutants also showed synaptic morphological defects. Strikingly TKV motility and p-Mad signaling were disrupted in larvae expressing two human disease proteins; expansions of glutamine repeats (polyQ77) and human amyloid precursor protein (APP) with a familial Alzheimer's disease (AD) mutation (APPswe). Consistent with axonal transport defects, larvae expressing these disease proteins showed accumulations of synaptic proteins along axons and synaptic abnormalities. Taken together our results suggest that similar to the NGF-TrkA signaling endosome, a BMP signaling endosome that directly interacts with molecular motors likely exist. Thus problems in axonal transport occurs early, perturbs BMP signaling, and likely contributes to the synaptic abnormalities observed in these two diseases. PMID:25127478

MEK-1 Activates C-Raf Through a Ras-Independent Mechanism

PubMed Central

Leicht, Deborah T.; Balan, Vitaly; Zhu, Jun; Kaplun, Alexander; Bronisz, Agnieszka; Rana, Ajay; Tzivion, Guri

2013-01-01

C-Raf is a member of the Ras-Raf-MEK-ERK mitogen-activated protein kinase (MAPK) signaling pathway that plays key roles in diverse physiological processes and is upregulated in many human cancers. C-Raf activation involves binding to Ras, increased phosphorylation and interactions with co-factors. Here, we describe a Ras-independent in vivo pathway for C-Raf activation by its downstream target MEK. Using 32P-metabolic labeling and 2D-phosphopeptide mapping experiments, we show that MEK increases C-Raf phosphorylation by up-to 10-fold. This increase was associated with C-Raf kinase activation, matching the activity seen with growth factor stimulation. Consequently, coexpression of wildtype C-Raf and MEK was sufficient for full and constitutive activation of ERK. Notably, the ability of MEK to activate C-Raf was completely Ras independent, since mutants impaired in Ras binding that are irresponsive to growth factors or Ras were fully activated by MEK. The ability of MEK to activate C-Raf was only partially dependent on MEK kinase activity but required MEK binding to C-Raf, suggesting that the binding results in a conformational change that increases C-Raf susceptibility to phosphorylation and activation or in the stabilization of the phosphorylated-active form. These findings propose a novel Ras-independent mechanism for activating C-Raf and the MAPK pathway without the need for mutations in the pathway. This mechanism could be of significance in pathological conditions or cancers overexpressing C-Raf and MEK or in conditions where C-Raf-MEK interaction is enhanced due to the downregulation of RKIP and MST2. PMID:23360980

Evolutionary Proteomics Uncovers Ancient Associations of Cilia with Signaling Pathways.

PubMed

Sigg, Monika Abedin; Menchen, Tabea; Lee, Chanjae; Johnson, Jeffery; Jungnickel, Melissa K; Choksi, Semil P; Garcia, Galo; Busengdal, Henriette; Dougherty, Gerard W; Pennekamp, Petra; Werner, Claudius; Rentzsch, Fabian; Florman, Harvey M; Krogan, Nevan; Wallingford, John B; Omran, Heymut; Reiter, Jeremy F

2017-12-18

Cilia are organelles specialized for movement and signaling. To infer when during evolution signaling pathways became associated with cilia, we characterized the proteomes of cilia from sea urchins, sea anemones, and choanoflagellates. We identified 437 high-confidence ciliary candidate proteins conserved in mammals and discovered that Hedgehog and G-protein-coupled receptor pathways were linked to cilia before the origin of bilateria and transient receptor potential (TRP) channels before the origin of animals. We demonstrated that candidates not previously implicated in ciliary biology localized to cilia and further investigated ENKUR, a TRP channel-interacting protein identified in the cilia of all three organisms. ENKUR localizes to motile cilia and is required for patterning the left-right axis in vertebrates. Moreover, mutation of ENKUR causes situs inversus in humans. Thus, proteomic profiling of cilia from diverse eukaryotes defines a conserved ciliary proteome, reveals ancient connections to signaling, and uncovers a ciliary protein that underlies development and human disease. Copyright © 2017 Elsevier Inc. All rights reserved.

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

Language Learning Variability within the Dorsal and Ventral Streams as a Cue for Compensatory Mechanisms in Aphasia Recovery

PubMed Central

López-Barroso, Diana; de Diego-Balaguer, Ruth

2017-01-01

Dorsal and ventral pathways connecting perisylvian language areas have been shown to be functionally and anatomically segregated. Whereas the dorsal pathway integrates the sensory-motor information required for verbal repetition, the ventral pathway has classically been associated with semantic processes. The great individual differences characterizing language learning through life partly correlate with brain structure and function within these dorsal and ventral language networks. Variability and plasticity within these networks also underlie inter-individual differences in the recovery of linguistic abilities in aphasia. Despite the division of labor of the dorsal and ventral streams, studies in healthy individuals have shown how the interaction of them and the redundancy in the areas they connect allow for compensatory strategies in functions that are usually segregated. In this mini-review we highlight the need to examine compensatory mechanisms between streams in healthy individuals as a helpful guide to choosing the most appropriate rehabilitation strategies, using spared functions and targeting preserved compensatory networks for brain plasticity. PMID:29021751

African swine fever virus infects macrophages, the natural host cells, via clathrin- and cholesterol-dependent endocytosis.

PubMed

Galindo, Inmaculada; Cuesta-Geijo, Miguel Angel; Hlavova, Karolina; Muñoz-Moreno, Raquel; Barrado-Gil, Lucía; Dominguez, Javier; Alonso, Covadonga

2015-03-16

The main cellular target for African swine fever virus (ASFV) is the porcine macrophage. However, existing data about the early phases of infection were previously characterized in non-leukocyte cells such as Vero cells. Here, we report that ASFV enters the natural host cell using dynamin-dependent and clathrin-mediated endocytosis. This pathway is strongly pH-dependent during the first steps of infection in porcine macrophages. We investigated the effect of drugs inhibiting several endocytic pathways in macrophages and compared ASFV with vaccinia virus (VV), which apparently involves different entry pathways. The presence of cholesterol in cellular membranes was found to be essential for a productive ASFV infection while actin-dependent endocytosis and the participation of phosphoinositide-3-kinase (PI3K) activity were other cellular factors required in the process of viral entry. These findings improved our understanding of the ASFV interactions with macrophages that allow for successful viral replication. Copyright © 2015 Elsevier B.V. All rights reserved.

A System-Level Pathway-Phenotype Association Analysis Using Synthetic Feature Random Forest

PubMed Central

Pan, Qinxin; Hu, Ting; Malley, James D.; Andrew, Angeline S.; Karagas, Margaret R.; Moore, Jason H.

2015-01-01

As the cost of genome-wide genotyping decreases, the number of genome-wide association studies (GWAS) has increased considerably. However, the transition from GWAS findings to the underlying biology of various phenotypes remains challenging. As a result, due to its system-level interpretability, pathway analysis has become a popular tool for gaining insights on the underlying biology from high-throughput genetic association data. In pathway analyses, gene sets representing particular biological processes are tested for significant associations with a given phenotype. Most existing pathway analysis approaches rely on single-marker statistics and assume that pathways are independent of each other. As biological systems are driven by complex biomolecular interactions, embracing the complex relationships between single-nucleotide polymorphisms (SNPs) and pathways needs to be addressed. To incorporate the complexity of gene-gene interactions and pathway-pathway relationships, we propose a system-level pathway analysis approach, synthetic feature random forest (SF-RF), which is designed to detect pathway-phenotype associations without making assumptions about the relationships among SNPs or pathways. In our approach, the genotypes of SNPs in a particular pathway are aggregated into a synthetic feature representing that pathway via Random Forest (RF). Multiple synthetic features are analyzed using RF simultaneously and the significance of a synthetic feature indicates the significance of the corresponding pathway. We further complement SF-RF with pathway-based Statistical Epistasis Network (SEN) analysis that evaluates interactions among pathways. By investigating the pathway SEN, we hope to gain additional insights into the genetic mechanisms contributing to the pathway-phenotype association. We apply SF-RF to a population-based genetic study of bladder cancer and further investigate the mechanisms that help explain the pathway-phenotype associations using SEN. The bladder cancer associated pathways we found are both consistent with existing biological knowledge and reveal novel and plausible hypotheses for future biological validations. PMID:24535726

Relation extraction for biological pathway construction using node2vec.

PubMed

Kim, Munui; Baek, Seung Han; Song, Min

2018-06-13

Systems biology is an important field for understanding whole biological mechanisms composed of interactions between biological components. One approach for understanding complex and diverse mechanisms is to analyze biological pathways. However, because these pathways consist of important interactions and information on these interactions is disseminated in a large number of biomedical reports, text-mining techniques are essential for extracting these relationships automatically. In this study, we applied node2vec, an algorithmic framework for feature learning in networks, for relationship extraction. To this end, we extracted genes from paper abstracts using pkde4j, a text-mining tool for detecting entities and relationships. Using the extracted genes, a co-occurrence network was constructed and node2vec was used with the network to generate a latent representation. To demonstrate the efficacy of node2vec in extracting relationships between genes, performance was evaluated for gene-gene interactions involved in a type 2 diabetes pathway. Moreover, we compared the results of node2vec to those of baseline methods such as co-occurrence and DeepWalk. Node2vec outperformed existing methods in detecting relationships in the type 2 diabetes pathway, demonstrating that this method is appropriate for capturing the relatedness between pairs of biological entities involved in biological pathways. The results demonstrated that node2vec is useful for automatic pathway construction.

Modeling of the hypothalamic-pituitary-adrenal axis-mediated interaction between the serotonin regulation pathway and the stress response using a Boolean approximation: a novel study of depression

PubMed Central

2013-01-01

Major depressive disorder (MDD) is a multifactorial disorder known to be influenced by both genetic and environmental factors. MDD presents a heritability of 37%, and a genetic contribution has also been observed in studies of family members of individuals with MDD that imply that the probability of suffering the disorder is approximately three times higher if a first-degree family member is affected. Childhood maltreatment and stressful life events (SLEs) have been established as critical environmental factors that profoundly influence the onset of MDD. The serotonin pathway has been a strong candidate for genetic studies, but it only explains a small proportion of the heritability of the disorder, which implies the involvement of other pathways. The serotonin (5-HT) pathway interacts with the stress response pathway in a manner mediated by the hypothalamic-pituitary-adrenal (HPA) axis. To analyze the interaction between the pathways, we propose the use of a synchronous Boolean network (SBN) approximation. The principal aim of this work was to model the interaction between these pathways, taking into consideration the presence of selective serotonin reuptake inhibitors (SSRIs), in order to observe how the pathways interact and to examine if the system is stable. Additionally, we wanted to study which genes or metabolites have the greatest impact on model stability when knocked out in silico. We observed that the biological model generated predicts steady states (attractors) for each of the different runs performed, thereby proving that the system is stable. These attractors changed in shape, especially when anti-depressive drugs were also included in the simulation. This work also predicted that the genes with the greatest impact on model stability were those involved in the neurotrophin pathway, such as CREB, BDNF (which has been associated with major depressive disorder in a variety of studies) and TRkB, followed by genes and metabolites related to 5-HT synthesis. PMID:24093582

Disentangling the multigenic and pleiotropic nature of molecular function

PubMed Central

2015-01-01

Background Biological processes at the molecular level are usually represented by molecular interaction networks. Function is organised and modularity identified based on network topology, however, this approach often fails to account for the dynamic and multifunctional nature of molecular components. For example, a molecule engaging in spatially or temporally independent functions may be inappropriately clustered into a single functional module. To capture biologically meaningful sets of interacting molecules, we use experimentally defined pathways as spatial/temporal units of molecular activity. Results We defined functional profiles of Saccharomyces cerevisiae based on a minimal set of Gene Ontology terms sufficient to represent each pathway's genes. The Gene Ontology terms were used to annotate 271 pathways, accounting for pathway multi-functionality and gene pleiotropy. Pathways were then arranged into a network, linked by shared functionality. Of the genes in our data set, 44% appeared in multiple pathways performing a diverse set of functions. Linking pathways by overlapping functionality revealed a modular network with energy metabolism forming a sparse centre, surrounded by several denser clusters comprised of regulatory and metabolic pathways. Signalling pathways formed a relatively discrete cluster connected to the centre of the network. Genetic interactions were enriched within the clusters of pathways by a factor of 5.5, confirming the organisation of our pathway network is biologically significant. Conclusions Our representation of molecular function according to pathway relationships enables analysis of gene/protein activity in the context of specific functional roles, as an alternative to typical molecule-centric graph-based methods. The pathway network demonstrates the cooperation of multiple pathways to perform biological processes and organises pathways into functionally related clusters with interdependent outcomes. PMID:26678917

Pathways of iron acquisition and utilization in Leishmania

PubMed Central

Flannery, Andrew R.; Renberg, Rebecca L.; Andrews, Norma W.

2013-01-01

Iron is essential for many metabolic pathways, but is toxic in excess. Recent identification of the ferric iron reductase LFR1, the ferrous iron transporter LIT1, and the heme transporter LHR1 greatly advanced our understanding of how Leishmania parasites acquire iron and regulate its uptake. LFR1 and LIT1 have close orthologs in plants, and are required for Leishmania virulence. Consistent with the lack of heme biosynthesis in trypanosomatids, LHR1 and LABCG5, a protein involved in heme salvage from hemoglobin, seem essential for Leishmania survival. LFR1, LIT1 and LHR1 are upregulated under low iron availability, in agreement with the need to prevent excessive iron uptake. Future studies should clarify how Leishmania interacts with the iron homeostasis machinery of its host cell, the macrophage. PMID:23962817

BioPAX – A community standard for pathway data sharing

PubMed Central

Demir, Emek; Cary, Michael P.; Paley, Suzanne; Fukuda, Ken; Lemer, Christian; Vastrik, Imre; Wu, Guanming; D’Eustachio, Peter; Schaefer, Carl; Luciano, Joanne; Schacherer, Frank; Martinez-Flores, Irma; Hu, Zhenjun; Jimenez-Jacinto, Veronica; Joshi-Tope, Geeta; Kandasamy, Kumaran; Lopez-Fuentes, Alejandra C.; Mi, Huaiyu; Pichler, Elgar; Rodchenkov, Igor; Splendiani, Andrea; Tkachev, Sasha; Zucker, Jeremy; Gopinath, Gopal; Rajasimha, Harsha; Ramakrishnan, Ranjani; Shah, Imran; Syed, Mustafa; Anwar, Nadia; Babur, Ozgun; Blinov, Michael; Brauner, Erik; Corwin, Dan; Donaldson, Sylva; Gibbons, Frank; Goldberg, Robert; Hornbeck, Peter; Luna, Augustin; Murray-Rust, Peter; Neumann, Eric; Reubenacker, Oliver; Samwald, Matthias; van Iersel, Martijn; Wimalaratne, Sarala; Allen, Keith; Braun, Burk; Whirl-Carrillo, Michelle; Dahlquist, Kam; Finney, Andrew; Gillespie, Marc; Glass, Elizabeth; Gong, Li; Haw, Robin; Honig, Michael; Hubaut, Olivier; Kane, David; Krupa, Shiva; Kutmon, Martina; Leonard, Julie; Marks, Debbie; Merberg, David; Petri, Victoria; Pico, Alex; Ravenscroft, Dean; Ren, Liya; Shah, Nigam; Sunshine, Margot; Tang, Rebecca; Whaley, Ryan; Letovksy, Stan; Buetow, Kenneth H.; Rzhetsky, Andrey; Schachter, Vincent; Sobral, Bruno S.; Dogrusoz, Ugur; McWeeney, Shannon; Aladjem, Mirit; Birney, Ewan; Collado-Vides, Julio; Goto, Susumu; Hucka, Michael; Le Novère, Nicolas; Maltsev, Natalia; Pandey, Akhilesh; Thomas, Paul; Wingender, Edgar; Karp, Peter D.; Sander, Chris; Bader, Gary D.

2010-01-01

BioPAX (Biological Pathway Exchange) is a standard language to represent biological pathways at the molecular and cellular level. Its major use is to facilitate the exchange of pathway data (http://www.biopax.org). Pathway data captures our understanding of biological processes, but its rapid growth necessitates development of databases and computational tools to aid interpretation. However, the current fragmentation of pathway information across many databases with incompatible formats presents barriers to its effective use. BioPAX solves this problem by making pathway data substantially easier to collect, index, interpret and share. BioPAX can represent metabolic and signaling pathways, molecular and genetic interactions and gene regulation networks. BioPAX was created through a community process. Through BioPAX, millions of interactions organized into thousands of pathways across many organisms, from a growing number of sources, are available. Thus, large amounts of pathway data are available in a computable form to support visualization, analysis and biological discovery. PMID:20829833

Knowledge-driven genomic interactions: an application in ovarian cancer.

PubMed

Kim, Dokyoon; Li, Ruowang; Dudek, Scott M; Frase, Alex T; Pendergrass, Sarah A; Ritchie, Marylyn D

2014-01-01

Effective cancer clinical outcome prediction for understanding of the mechanism of various types of cancer has been pursued using molecular-based data such as gene expression profiles, an approach that has promise for providing better diagnostics and supporting further therapies. However, clinical outcome prediction based on gene expression profiles varies between independent data sets. Further, single-gene expression outcome prediction is limited for cancer evaluation since genes do not act in isolation, but rather interact with other genes in complex signaling or regulatory networks. In addition, since pathways are more likely to co-operate together, it would be desirable to incorporate expert knowledge to combine pathways in a useful and informative manner. Thus, we propose a novel approach for identifying knowledge-driven genomic interactions and applying it to discover models associated with cancer clinical phenotypes using grammatical evolution neural networks (GENN). In order to demonstrate the utility of the proposed approach, an ovarian cancer data from the Cancer Genome Atlas (TCGA) was used for predicting clinical stage as a pilot project. We identified knowledge-driven genomic interactions associated with cancer stage from single knowledge bases such as sources of pathway-pathway interaction, but also knowledge-driven genomic interactions across different sets of knowledge bases such as pathway-protein family interactions by integrating different types of information. Notably, an integration model from different sources of biological knowledge achieved 78.82% balanced accuracy and outperformed the top models with gene expression or single knowledge-based data types alone. Furthermore, the results from the models are more interpretable because they are framed in the context of specific biological pathways or other expert knowledge. The success of the pilot study we have presented herein will allow us to pursue further identification of models predictive of clinical cancer survival and recurrence. Understanding the underlying tumorigenesis and progression in ovarian cancer through the global view of interactions within/between different biological knowledge sources has the potential for providing more effective screening strategies and therapeutic targets for many types of cancer.

Prediction of GCRV virus-host protein interactome based on structural motif-domain interactions.

PubMed

Zhang, Aidi; He, Libo; Wang, Yaping

2017-03-02

Grass carp hemorrhagic disease, caused by grass carp reovirus (GCRV), is the most fatal causative agent in grass carp aquaculture. Protein-protein interactions between virus and host are one avenue through which GCRV can trigger infection and induce disease. Experimental approaches for the detection of host-virus interactome have many inherent limitations, and studies on protein-protein interactions between GCRV and its host remain rare. In this study, based on known motif-domain interaction information, we systematically predicted the GCRV virus-host protein interactome by using motif-domain interaction pair searching strategy. These proteins derived from different domain families and were predicted to interact with different motif patterns in GCRV. JAM-A protein was successfully predicted to interact with motifs of GCRV Sigma1-like protein, and shared the similar binding mode compared with orthoreovirus. Differentially expressed genes during GCRV infection process were extracted and mapped to our predicted interactome, the overlapped genes displayed different tissue expression distributions on the whole, the overall expression level in intestinal is higher than that of other three tissues, which may suggest that the functions of these genes are more active in intestinal. Function annotation and pathway enrichment analysis revealed that the host targets were largely involved in signaling pathway and immune pathway, such as interferon-gamma signaling pathway, VEGF signaling pathway, EGF receptor signaling pathway, B cell activation, and T cell activation. Although the predicted PPIs may contain some false positives due to limited data resource and poor research background in non-model species, the computational method still provide reasonable amount of interactions, which can be further validated by high throughput experiments. The findings of this work will contribute to the development of system biology for GCRV infectious diseases, and help guide the identification of novel receptors of GCRV in its host.

Co-introduction vs ecological fitting as pathways to the establishment of effective mutualisms during biological invasions.

PubMed

Le Roux, Johannes J; Hui, Cang; Keet, Jan-Hendrik; Ellis, Allan G

2017-09-01

Contents 1354 I. 1354 II. 1355 III. 1357 IV. 1357 V. 1359 1359 References 1359 SUMMARY: Interactions between non-native plants and their mutualists are often disrupted upon introduction to new environments. Using legume-rhizobium mutualistic interactions as an example, we discuss two pathways that can influence symbiotic associations in such situations: co-introduction of coevolved rhizobia; and utilization of, and adaptation to, resident rhizobia, hereafter referred to as 'ecological fitting'. Co-introduction and ecological fitting have distinct implications for successful legume invasions and their impacts. Under ecological fitting, initial impacts may be less severe and will accrue over longer periods as novel symbiotic associations and/or adaptations may require fine-tuning over time. Co-introduction will have more profound impacts that will accrue more rapidly as a result of positive feedbacks between densities of non-native rhizobia and their coevolved host plants, in turn enhancing competition between native and non-native rhizobia. Co-introduction can further impact invasion outcomes by the exchange of genetic material between native and non-native rhizobia, potentially resulting in decreased fitness of native legumes. A better understanding of the roles of these two pathways in the invasion dynamics of non-native legumes is much needed, and we highlight some of the exciting research avenues it presents. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Elucidation of the glucose transport pathway in glucose transporter 4 via steered molecular dynamics simulations.

PubMed

Sheena, Aswathy; Mohan, Suma S; Haridas, Nidhina Pachakkil A; Anilkumar, Gopalakrishnapillai

2011-01-01

GLUT4 is a predominant insulin regulated glucose transporter expressed in major glucose disposal tissues such as adipocytes and muscles. Under the unstimulated state, GLUT4 resides within intracellular vesicles. Various stimuli such as insulin translocate this protein to the plasma membrane for glucose transport. In the absence of a crystal structure for GLUT4, very little is known about the mechanism of glucose transport by this protein. Earlier we proposed a homology model for GLUT4 and performed a conventional molecular dynamics study revealing the conformational rearrangements during glucose and ATP binding. However, this study could not explain the transport of glucose through the permeation tunnel. To elucidate the molecular mechanism of glucose transport and its energetic, a steered molecular dynamics study (SMD) was used. Glucose was pulled from the extracellular end of GLUT4 to the cytoplasm along the pathway using constant velocity pulling method. We identified several key residues within the tunnel that interact directly with either the backbone ring or the hydroxyl groups of glucose. A rotation of glucose molecule was seen near the sugar binding site facilitating the sugar recognition process at the QLS binding site. This study proposes a possible glucose transport pathway and aids the identification of several residues that make direct interactions with glucose during glucose transport. Mutational studies are required to further validate the observation made in this study.

Applied neuroanatomy elective to reinforce and promote engagement with neurosensory pathways using interactive and artistic activities.

PubMed

Dao, Vinh; Yeh, Pon-Hsiu; Vogel, Kristine S; Moore, Charleen M

2015-01-01

One in six Americans is currently affected by neurologic disease. As the United States population ages, the number of neurologic complaints is expected to increase. Thus, there is a pressing need for more neurologists as well as more neurology training in other specialties. Often interest in neurology begins during medical school, so improving education in medical neural courses is a critical step toward producing more neurologists and better neurology training in other specialists. To this end, a novel applied neuroanatomy elective was designed at the University of Texas Health Science Center at San Antonio (UTHSCSA) to complement the traditional first-year medical neuroscience course and promote engagement and deep learning of the material with a focus on neurosensory pathways. The elective covered four neurosensory modalities (proprioception/balance, vision, auditory, and taste/olfaction) over four sessions, each with a short classroom component and a much longer activity component. At each session, students reviewed the neurosensory pathways through structured presentations and then applied them to preplanned interactive activities, many of which allowed students to utilize their artistic talents. Students were required to complete subjective pre-course and post-course surveys and reflections. The survey results and positive student comments suggest that the elective was a valuable tool when used in parallel with the traditional medical neuroscience course in promoting engagement and reinforcement of the neurosensory material. © 2014 American Association of Anatomists.

DNA repair targeted therapy: the past or future of cancer treatment?

PubMed Central

Gavande, Navnath S.; VanderVere-Carozza, Pamela S.; Hinshaw, Hilary D.; Jalal, Shadia I.; Sears, Catherine R.; Pawelczak, Katherine S.; Turchi, John J.

2016-01-01

The repair of DNA damage is a complex process that relies on particular pathways to remedy specific types of damage to DNA. The range of insults to DNA includes small, modest changes in structure including mismatched bases and simple methylation events to oxidized bases, intra- and interstrand DNA crosslinks, DNA double strand breaks and protein-DNA adducts. Pathways required for the repair of these lesions include mismatch repair, base excision repair, nucleotide excision repair, and the homology directed repair/Fanconi anemia pathway. Each of these pathways contributes to genetic stability, and mutations in genes encoding proteins involved in these pathways have been demonstrated to promote genetic instability and cancer. In fact, it has been suggested all cancers display defects in DNA repair. It has also been demonstrated that the ability of cancer cells to repair therapeutically induced DNA damage impacts therapeutic efficacy. This has led to targeting DNA repair pathways and proteins to develop anti-cancer agents that will increase sensitivity to traditional chemotherapeutics. While initial studies languished and were plagued by a lack of specificity and a defined mechanism of action, more recent approaches to exploit synthetic lethal interaction and develop high affinity chemical inhibitors have proven considerably more effective. In this review we will highlight recent advances and discuss previous failures in targeting DNA repair to pave the way for future DNA repair targeted agents and their use in cancer therapy. PMID:26896565

An overview of bioinformatics methods for modeling biological pathways in yeast

PubMed Central

Hou, Jie; Acharya, Lipi; Zhu, Dongxiao

2016-01-01

The advent of high-throughput genomics techniques, along with the completion of genome sequencing projects, identification of protein–protein interactions and reconstruction of genome-scale pathways, has accelerated the development of systems biology research in the yeast organism Saccharomyces cerevisiae. In particular, discovery of biological pathways in yeast has become an important forefront in systems biology, which aims to understand the interactions among molecules within a cell leading to certain cellular processes in response to a specific environment. While the existing theoretical and experimental approaches enable the investigation of well-known pathways involved in metabolism, gene regulation and signal transduction, bioinformatics methods offer new insights into computational modeling of biological pathways. A wide range of computational approaches has been proposed in the past for reconstructing biological pathways from high-throughput datasets. Here we review selected bioinformatics approaches for modeling biological pathways in S. cerevisiae, including metabolic pathways, gene-regulatory pathways and signaling pathways. We start with reviewing the research on biological pathways followed by discussing key biological databases. In addition, several representative computational approaches for modeling biological pathways in yeast are discussed. PMID:26476430

Biosynthesis and Metabolic Fate of Phenylalanine in Conifers

PubMed Central

Pascual, María B.; El-Azaz, Jorge; de la Torre, Fernando N.; Cañas, Rafael A.; Avila, Concepción; Cánovas, Francisco M.

2016-01-01

The amino acid phenylalanine (Phe) is a critical metabolic node that plays an essential role in the interconnection between primary and secondary metabolism in plants. Phe is used as a protein building block but it is also as a precursor for numerous plant compounds that are crucial for plant reproduction, growth, development, and defense against different types of stresses. The metabolism of Phe plays a central role in the channeling of carbon from photosynthesis to the biosynthesis of phenylpropanoids. The study of this metabolic pathway is particularly relevant in trees, which divert large amounts of carbon into the biosynthesis of Phe-derived compounds, particularly lignin, an important constituent of wood. The trunks of trees are metabolic sinks that consume a considerable percentage of carbon and energy from photosynthesis, and carbon is finally immobilized in wood. This paper reviews recent advances in the biosynthesis and metabolic utilization of Phe in conifer trees. Two alternative routes have been identified: the ancient phenylpyruvate pathway that is present in microorganisms, and the arogenate pathway that possibly evolved later during plant evolution. Additionally, an efficient nitrogen recycling mechanism is required to maintain sustained growth during xylem formation. The relevance of phenylalanine metabolic pathways in wood formation, the biotic interactions, and ultraviolet protection is discussed. The genetic manipulation and transcriptional regulation of the pathways are also outlined. PMID:27468292

Biosynthesis and Metabolic Fate of Phenylalanine in Conifers.

PubMed

Pascual, María B; El-Azaz, Jorge; de la Torre, Fernando N; Cañas, Rafael A; Avila, Concepción; Cánovas, Francisco M

2016-01-01

The amino acid phenylalanine (Phe) is a critical metabolic node that plays an essential role in the interconnection between primary and secondary metabolism in plants. Phe is used as a protein building block but it is also as a precursor for numerous plant compounds that are crucial for plant reproduction, growth, development, and defense against different types of stresses. The metabolism of Phe plays a central role in the channeling of carbon from photosynthesis to the biosynthesis of phenylpropanoids. The study of this metabolic pathway is particularly relevant in trees, which divert large amounts of carbon into the biosynthesis of Phe-derived compounds, particularly lignin, an important constituent of wood. The trunks of trees are metabolic sinks that consume a considerable percentage of carbon and energy from photosynthesis, and carbon is finally immobilized in wood. This paper reviews recent advances in the biosynthesis and metabolic utilization of Phe in conifer trees. Two alternative routes have been identified: the ancient phenylpyruvate pathway that is present in microorganisms, and the arogenate pathway that possibly evolved later during plant evolution. Additionally, an efficient nitrogen recycling mechanism is required to maintain sustained growth during xylem formation. The relevance of phenylalanine metabolic pathways in wood formation, the biotic interactions, and ultraviolet protection is discussed. The genetic manipulation and transcriptional regulation of the pathways are also outlined.

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.

Binding of Substrates to the Central Pore of the Vps4 ATPase Is Autoinhibited by the Microtubule Interacting and Trafficking (MIT) Domain and Activated by MIT Interacting Motifs (MIMs).

PubMed

Han, Han; Monroe, Nicole; Votteler, Jörg; Shakya, Binita; Sundquist, Wesley I; Hill, Christopher P

2015-05-22

The endosomal sorting complexes required for transport (ESCRT) pathway drives reverse topology membrane fission events within multiple cellular pathways, including cytokinesis, multivesicular body biogenesis, repair of the plasma membrane, nuclear membrane vesicle formation, and HIV budding. The AAA ATPase Vps4 is recruited to membrane necks shortly before fission, where it catalyzes disassembly of the ESCRT-III lattice. The N-terminal Vps4 microtubule-interacting and trafficking (MIT) domains initially bind the C-terminal MIT-interacting motifs (MIMs) of ESCRT-III subunits, but it is unclear how the enzyme then remodels these substrates in response to ATP hydrolysis. Here, we report quantitative binding studies that demonstrate that residues from helix 5 of the Vps2p subunit of ESCRT-III bind to the central pore of an asymmetric Vps4p hexamer in a manner that is dependent upon the presence of flexible nucleotide analogs that can mimic multiple states in the ATP hydrolysis cycle. We also find that substrate engagement is autoinhibited by the Vps4p MIT domain and that this inhibition is relieved by binding of either Type 1 or Type 2 MIM elements, which bind the Vps4p MIT domain through different interfaces. These observations support the model that Vps4 substrates are initially recruited by an MIM-MIT interaction that activates the Vps4 central pore to engage substrates and generate force, thereby triggering ESCRT-III disassembly. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Binding of Substrates to the Central Pore of the Vps4 ATPase Is Autoinhibited by the Microtubule Interacting and Trafficking (MIT) Domain and Activated by MIT Interacting Motifs (MIMs)*

PubMed Central

Han, Han; Monroe, Nicole; Votteler, Jörg; Shakya, Binita; Sundquist, Wesley I.; Hill, Christopher P.

2015-01-01

The endosomal sorting complexes required for transport (ESCRT) pathway drives reverse topology membrane fission events within multiple cellular pathways, including cytokinesis, multivesicular body biogenesis, repair of the plasma membrane, nuclear membrane vesicle formation, and HIV budding. The AAA ATPase Vps4 is recruited to membrane necks shortly before fission, where it catalyzes disassembly of the ESCRT-III lattice. The N-terminal Vps4 microtubule-interacting and trafficking (MIT) domains initially bind the C-terminal MIT-interacting motifs (MIMs) of ESCRT-III subunits, but it is unclear how the enzyme then remodels these substrates in response to ATP hydrolysis. Here, we report quantitative binding studies that demonstrate that residues from helix 5 of the Vps2p subunit of ESCRT-III bind to the central pore of an asymmetric Vps4p hexamer in a manner that is dependent upon the presence of flexible nucleotide analogs that can mimic multiple states in the ATP hydrolysis cycle. We also find that substrate engagement is autoinhibited by the Vps4p MIT domain and that this inhibition is relieved by binding of either Type 1 or Type 2 MIM elements, which bind the Vps4p MIT domain through different interfaces. These observations support the model that Vps4 substrates are initially recruited by an MIM-MIT interaction that activates the Vps4 central pore to engage substrates and generate force, thereby triggering ESCRT-III disassembly. PMID:25833946

A Web Tool for Generating High Quality Machine-readable Biological Pathways.

PubMed

Ramirez-Gaona, Miguel; Marcu, Ana; Pon, Allison; Grant, Jason; Wu, Anthony; Wishart, David S

2017-02-08

PathWhiz is a web server built to facilitate the creation of colorful, interactive, visually pleasing pathway diagrams that are rich in biological information. The pathways generated by this online application are machine-readable and fully compatible with essentially all web-browsers and computer operating systems. It uses a specially developed, web-enabled pathway drawing interface that permits the selection and placement of different combinations of pre-drawn biological or biochemical entities to depict reactions, interactions, transport processes and binding events. This palette of entities consists of chemical compounds, proteins, nucleic acids, cellular membranes, subcellular structures, tissues, and organs. All of the visual elements in it can be interactively adjusted and customized. Furthermore, because this tool is a web server, all pathways and pathway elements are publicly accessible. This kind of pathway "crowd sourcing" means that PathWhiz already contains a large and rapidly growing collection of previously drawn pathways and pathway elements. Here we describe a protocol for the quick and easy creation of new pathways and the alteration of existing pathways. To further facilitate pathway editing and creation, the tool contains replication and propagation functions. The replication function allows existing pathways to be used as templates to create or edit new pathways. The propagation function allows one to take an existing pathway and automatically propagate it across different species. Pathways created with this tool can be "re-styled" into different formats (KEGG-like or text-book like), colored with different backgrounds, exported to BioPAX, SBGN-ML, SBML, or PWML data exchange formats, and downloaded as PNG or SVG images. The pathways can easily be incorporated into online databases, integrated into presentations, posters or publications, or used exclusively for online visualization and exploration. This protocol has been successfully applied to generate over 2,000 pathway diagrams, which are now found in many online databases including HMDB, DrugBank, SMPDB, and ECMDB.

Determinants of Swe1p Degradation in Saccharomyces cerevisiae

PubMed Central

McMillan, John N.; Theesfeld, Chandra L.; Harrison, Jacob C.; Bardes, Elaine S. G.; Lew, Daniel J.

2002-01-01

Swe1p, the sole Wee1-family kinase in Saccharomyces cerevisiae, is synthesized during late G1 and is then degraded as cells proceed through the cell cycle. However, Swe1p degradation is halted by the morphogenesis checkpoint, which responds to insults that perturb bud formation. The Swe1p stabilization promotes cell cycle arrest through Swe1p-mediated inhibitory phosphorylation of Cdc28p until the cells can recover from the perturbation and resume bud formation. Swe1p degradation involves the relocalization of Swe1p from the nucleus to the mother-bud neck, and neck targeting requires the Swe1p-interacting protein Hsl7p. In addition, Swe1p degradation is stimulated by its substrate, cyclin/Cdc28p, and Swe1p is thought to be a target of the ubiquitin ligase SCFMet30 acting with the ubiquitin-conjugating enzyme Cdc34p. The basis for regulation of Swe1p degradation by the morphogenesis checkpoint remains unclear, and in order to elucidate that regulation we have dissected the Swe1p degradation pathway in more detail, yielding several novel findings. First, we show here that Met30p (and by implication SCFMet30) is not, in fact, required for Swe1p degradation. Second, cyclin/Cdc28p does not influence Swe1p neck targeting, but can directly phosphorylate Swe1p, suggesting that it acts downstream of neck targeting in the Swe1p degradation pathway. Third, a screen for functional but nondegradable mutants of SWE1 identified two small regions of Swe1p that are key to its degradation. One of these regions mediates interaction of Swe1p with Hsl7p, showing that the Swe1p-Hsl7p interaction is critical for Swe1p neck targeting and degradation. The other region did not appear to affect interactions with known Swe1p regulators, suggesting that other as-yet-unknown regulators exist. PMID:12388757

Dissecting Seed Mucilage Adherence Mediated by FEI2 and SOS5

PubMed Central

Griffiths, Jonathan S.; Crepeau, Marie-Jeanne; Ralet, Marie-Christine; Seifert, Georg J.; North, Helen M.

2016-01-01

The plant cell wall is held together by the interactions between four major components: cellulose, pectin, hemicellulose, and proteins. Mucilage is a powerful model system to study the interactions between these components as it is formed of polysaccharides that are deposited in the apoplast of seed coat epidermal cells during seed development. When seeds are hydrated, these polysaccharides expand rapidly out of the apoplastic pocket, and form an adherent halo of mucilage around the seed. In Arabidopsis, mutations in multiple genes have similar loss of mucilage adherence phenotypes including CELLULOSE SYNTHASE 5 (CESA5)/MUCILAGE-MODIFIED 3 (MUM3), MUM5/MUCI21, SALT-OVERLY SENSITIVE 5 (SOS5), and FEI2. Here, we examine the interactions between these factors to better understand how they participate to control mucilage adherence. Double mutant phenotypes indicated that MUM5 and CESA5 function in a common mechanism that adheres pectin to the seed through the biosynthesis of cellulose and xylan, whereas SOS5 and FEI2, encoding a fasciclin-like arabinogalactan protein or a receptor-like kinase, respectively, function through an independent pathway. Cytological analyses of mucilage indicates that heteromannans are associated with cellulose, and not in the pathway involving SOS5 or FEI2. A SOS5 fluorescent protein fusion (SOS5-mCITRINE) was localized throughout the mucilage pocket, consistent with a structural role in pectin adhesion. The relationship between SOS5 and FEI2 mediated mucilage adherence was examined in more detail and while sos5 and fei2 mutants show similar phenotypes, key differences in the macromolecular characteristics and amounts of mucilage polymers were observed. FEI2 thus appears to have additional, as well as overlapping functions, with SOS5. Given that FEI2 is required for SOS5 function, we propose that FEI2 serves to localize SOS5 at the plasma membrane where it establishes interactions with mucilage polysaccharides, notably pectins, required for mucilage adherence prior to SOS5 being released into the apoplast. PMID:27524986

Bovine Foamy Virus Transactivator BTas Interacts with Cellular RelB To Enhance Viral Transcription▿

PubMed Central

Wang, Jian; Tan, Juan; Guo, Hongyan; Zhang, Qicheng; Jia, Rui; Xu, Xuan; Geng, Yunqi; Qiao, Wentao

2010-01-01

Viruses are obligate intracellular parasites that depend on cellular machinery for their efficient transcription and replication. In a previous study we reported that bovine foamy virus (BFV) is able to activate the nuclear factor κB (NF-κB) pathway through the action of its transactivator BTas to enhance viral transcription. However, the mechanism used by NF-κB to enhance BFV transcription remains elusive. To address this question, we employed a yeast two-hybrid assay to screen for BTas-interacting proteins. We found that RelB, a member of NF-κB protein family, interacts with BTas. We confirmed the putative RelB-BTas interaction in vitro and in vivo and identified the protein regions responsible for the RelB-BTas interaction. Using a luciferase reporter assay, we next showed that RelB enhances BFV transcription (BTas-induced long terminal repeat [LTR] transactivation) and that this process requires both the localization of the RelB-BTas interaction in the nucleus and the Rel homology domain of RelB. The knockdown of the cellular endogenous RelB protein using small interfering RNA (siRNA) significantly attenuated BTas-induced LTR transcription. The results of chromatin immunoprecipitation (ChIP) analysis showed that endogenous RelB binds to the viral LTR in BFV-infected cells. Together, these results suggest that BFV engages the RelB protein as a cotransactivator of BTas to enhance viral transcription. In addition, our findings indicate that BFV infection upregulates cellular RelB expression through BTas-induced NF-κB activation. Thus, this study demonstrates the existence of a positive-feedback circuit in which BFV utilizes the host's NF-κB pathway through the RelB protein for efficient viral transcription. PMID:20844054

Bovine foamy virus transactivator BTas interacts with cellular RelB to enhance viral transcription.

PubMed

Wang, Jian; Tan, Juan; Guo, Hongyan; Zhang, Qicheng; Jia, Rui; Xu, Xuan; Geng, Yunqi; Qiao, Wentao

2010-11-01

Viruses are obligate intracellular parasites that depend on cellular machinery for their efficient transcription and replication. In a previous study we reported that bovine foamy virus (BFV) is able to activate the nuclear factor κB (NF-κB) pathway through the action of its transactivator BTas to enhance viral transcription. However, the mechanism used by NF-κB to enhance BFV transcription remains elusive. To address this question, we employed a yeast two-hybrid assay to screen for BTas-interacting proteins. We found that RelB, a member of NF-κB protein family, interacts with BTas. We confirmed the putative RelB-BTas interaction in vitro and in vivo and identified the protein regions responsible for the RelB-BTas interaction. Using a luciferase reporter assay, we next showed that RelB enhances BFV transcription (BTas-induced long terminal repeat [LTR] transactivation) and that this process requires both the localization of the RelB-BTas interaction in the nucleus and the Rel homology domain of RelB. The knockdown of the cellular endogenous RelB protein using small interfering RNA (siRNA) significantly attenuated BTas-induced LTR transcription. The results of chromatin immunoprecipitation (ChIP) analysis showed that endogenous RelB binds to the viral LTR in BFV-infected cells. Together, these results suggest that BFV engages the RelB protein as a cotransactivator of BTas to enhance viral transcription. In addition, our findings indicate that BFV infection upregulates cellular RelB expression through BTas-induced NF-κB activation. Thus, this study demonstrates the existence of a positive-feedback circuit in which BFV utilizes the host's NF-κB pathway through the RelB protein for efficient viral transcription.

Redox Regulation and the Autistic Spectrum: Role of Tryptophan Catabolites, Immuno-inflammation, Autoimmunity and the Amygdala

PubMed Central

Anderson, George; Maes, Michael

2014-01-01

The autistic spectrum disorders (ASD) form a set of multi-faceted disorders with significant genetic, epigenetic and environmental determinants. Oxidative and nitrosative stress (O&NS), immuno-inflammatory pathways, mitochondrial dysfunction and dysregulation of the tryptophan catabolite (TRYCATs) pathway play significant interactive roles in driving the early developmental etiology and course of ASD. O&NS interactions with immuno-inflammatory pathways mediate their effects centrally via the regulation of astrocyte and microglia responses, including regional variations in TRYCATs produced. Here we review the nature of these interactions and propose an early developmental model whereby different ASD genetic susceptibilities interact with environmental and epigenetic processes, resulting in glia biasing the patterning of central interarea interactions. A role for decreased local melatonin and N-acetylserotonin production by immune and glia cells may be a significant treatment target. PMID:24669209

Differential reconstructed gene interaction networks for deriving toxicity threshold in chemical risk assessment.

PubMed

Yang, Yi; Maxwell, Andrew; Zhang, Xiaowei; Wang, Nan; Perkins, Edward J; Zhang, Chaoyang; Gong, Ping

2013-01-01

Pathway alterations reflected as changes in gene expression regulation and gene interaction can result from cellular exposure to toxicants. Such information is often used to elucidate toxicological modes of action. From a risk assessment perspective, alterations in biological pathways are a rich resource for setting toxicant thresholds, which may be more sensitive and mechanism-informed than traditional toxicity endpoints. Here we developed a novel differential networks (DNs) approach to connect pathway perturbation with toxicity threshold setting. Our DNs approach consists of 6 steps: time-series gene expression data collection, identification of altered genes, gene interaction network reconstruction, differential edge inference, mapping of genes with differential edges to pathways, and establishment of causal relationships between chemical concentration and perturbed pathways. A one-sample Gaussian process model and a linear regression model were used to identify genes that exhibited significant profile changes across an entire time course and between treatments, respectively. Interaction networks of differentially expressed (DE) genes were reconstructed for different treatments using a state space model and then compared to infer differential edges/interactions. DE genes possessing differential edges were mapped to biological pathways in databases such as KEGG pathways. Using the DNs approach, we analyzed a time-series Escherichia coli live cell gene expression dataset consisting of 4 treatments (control, 10, 100, 1000 mg/L naphthenic acids, NAs) and 18 time points. Through comparison of reconstructed networks and construction of differential networks, 80 genes were identified as DE genes with a significant number of differential edges, and 22 KEGG pathways were altered in a concentration-dependent manner. Some of these pathways were perturbed to a degree as high as 70% even at the lowest exposure concentration, implying a high sensitivity of our DNs approach. Findings from this proof-of-concept study suggest that our approach has a great potential in providing a novel and sensitive tool for threshold setting in chemical risk assessment. In future work, we plan to analyze more time-series datasets with a full spectrum of concentrations and sufficient replications per treatment. The pathway alteration-derived thresholds will also be compared with those derived from apical endpoints such as cell growth rate.

Tetratricopeptide-motif-mediated interaction of FANCG with recombination proteins XRCC3 and BRCA2.

PubMed

Hussain, Shobbir; Wilson, James B; Blom, Eric; Thompson, Larry H; Sung, Patrick; Gordon, Susan M; Kupfer, Gary M; Joenje, Hans; Mathew, Christopher G; Jones, Nigel J

2006-05-10

Fanconi anaemia is an inherited chromosomal instability disorder characterised by cellular sensitivity to DNA interstrand crosslinkers, bone-marrow failure and a high risk of cancer. Eleven FA genes have been identified, one of which, FANCD1, is the breast cancer susceptibility gene BRCA2. At least eight FA proteins form a nuclear core complex required for monoubiquitination of FANCD2. The BRCA2/FANCD1 protein is connected to the FA pathway by interactions with the FANCG and FANCD2 proteins, both of which co-localise with the RAD51 recombinase, which is regulated by BRCA2. These connections raise the question of whether any of the FANC proteins of the core complex might also participate in other complexes involved in homologous recombination repair. We therefore tested known FA proteins for direct interaction with RAD51 and its paralogs XRCC2 and XRCC3. FANCG was found to interact with XRCC3, and this interaction was disrupted by the FA-G patient derived mutation L71P. FANCG was co-immunoprecipitated with both XRCC3 and BRCA2 from extracts of human and hamster cells. The FANCG-XRCC3 and FANCG-BRCA2 interactions did not require the presence of other FA proteins from the core complex, suggesting that FANCG also participates in a DNA repair complex that is downstream and independent of FANCD2 monoubiquitination. Additionally, XRCC3 and BRCA2 proteins co-precipitate in both human and hamster cells and this interaction requires FANCG. The FANCG protein contains multiple tetratricopeptide repeat motifs (TPRs), which function as scaffolds to mediate protein-protein interactions. Mutation of one or more of these motifs disrupted all of the known interactions of FANCG. We propose that FANCG, in addition to stabilising the FA core complex, may have a role in building multiprotein complexes that facilitate homologous recombination repair.

Mitochondrial thiol oxidase Erv1: both shuttle cysteine residues are required for its function with distinct roles

PubMed Central

Ang, Swee Kim; Zhang, Mengqi; Lodi, Tiziana; Lu, Hui

2014-01-01

Erv1 (essential for respiration and viability 1), is an essential component of the MIA (mitochondrial import and assembly) pathway, playing an important role in the oxidative folding of mitochondrial intermembrane space proteins. In the MIA pathway, Mia40, a thiol oxidoreductase with a CPC motif at its active site, oxidizes newly imported substrate proteins. Erv1 a FAD-dependent thiol oxidase, in turn reoxidizes Mia40 via its N-terminal Cys30–Cys33 shuttle disulfide. However, it is unclear how the two shuttle cysteine residues of Erv1 relay electrons from the Mia40 CPC motif to the Erv1 active-site Cys130–Cys133 disulfide. In the present study, using yeast genetic approaches we showed that both shuttle cysteine residues of Erv1 are required for cell growth. In organelle and in vitro studies confirmed that both shuttle cysteine residues were indeed required for import of MIA pathway substrates and Erv1 enzyme function to oxidize Mia40. Furthermore, our results revealed that the two shuttle cysteine residues of Erv1 are functionally distinct. Although Cys33 is essential for forming the intermediate disulfide Cys33–Cys130′ and transferring electrons to the redox active-site directly, Cys30 plays two important roles: (i) dominantly interacts and receives electrons from the Mia40 CPC motif; and (ii) resolves the Erv1 Cys33–Cys130 intermediate disulfide. Taken together, we conclude that both shuttle cysteine residues are required for Erv1 function, and play complementary, but distinct, roles to ensure rapid turnover of active Erv1. PMID:24625320

Effective connectivity of ascending and descending frontal-thalamic pathways during sustained attention: Complex brain network interactions in adolescence

PubMed Central

Jagtap, Pranav; Diwadkar, Vaibhav A.

2016-01-01

Frontal-thalamic interactions are crucial for bottom-up gating and top-down control, yet have not been well studied from brain network perspectives. We applied network modeling of fMRI signals (Dynamic Causal Modeling; DCM) to investigate frontal-thalamic interactions during an attention task with parametrically varying levels of demand. fMRI was collected while subjects participated in a sustained continuous performance task with low and high attention demands. 162 competing model architectures were employed in DCM to evaluate hypotheses on bilateral frontal-thalamic connections and their modulation by attention demand, selected at a second level using Bayesian Model Selection. The model architecture evinced significant contextual modulation by attention of ascending (thalamus → dPFC) and descending (dPFC → thalamus) pathways. However, modulation of these pathways was asymmetric: While positive modulation of the ascending pathway was comparable across attention demand, modulation of the descending pathway was significantly greater when attention demands were increased. Increased modulation of the (dPFC → thalamus) pathway in response to increased attention demand constitutes novel evidence of attention-related gain in the connectivity of the descending attention pathway. By comparison demand-independent modulation of the ascending (thalamus → dPFC) pathway suggests unbiased thalamic inputs to the cortex in the context of the paradigm. PMID:27145923

A Network-Based Kernel Machine Test for the Identification of Risk Pathways in Genome-Wide Association Studies

PubMed Central

Freytag, Saskia; Manitz, Juliane; Schlather, Martin; Kneib, Thomas; Amos, Christopher I.; Risch, Angela; Chang-Claude, Jenny; Heinrich, Joachim; Bickeböller, Heike

2014-01-01

Biological pathways provide rich information and biological context on the genetic causes of complex diseases. The logistic kernel machine test integrates prior knowledge on pathways in order to analyze data from genome-wide association studies (GWAS). Here, the kernel converts genomic information of two individuals to a quantitative value reflecting their genetic similarity. With the selection of the kernel one implicitly chooses a genetic effect model. Like many other pathway methods, none of the available kernels accounts for topological structure of the pathway or gene-gene interaction types. However, evidence indicates that connectivity and neighborhood of genes are crucial in the context of GWAS, because genes associated with a disease often interact. Thus, we propose a novel kernel that incorporates the topology of pathways and information on interactions. Using simulation studies, we demonstrate that the proposed method maintains the type I error correctly and can be more effective in the identification of pathways associated with a disease than non-network-based methods. We apply our approach to genome-wide association case control data on lung cancer and rheumatoid arthritis. We identify some promising new pathways associated with these diseases, which may improve our current understanding of the genetic mechanisms. PMID:24434848

minepath.org: a free interactive pathway analysis web server.

PubMed

Koumakis, Lefteris; Roussos, Panos; Potamias, George

2017-07-03

( www.minepath.org ) is a web-based platform that elaborates on, and radically extends the identification of differentially expressed sub-paths in molecular pathways. Besides the network topology, the underlying MinePath algorithmic processes exploit exact gene-gene molecular relationships (e.g. activation, inhibition) and are able to identify differentially expressed pathway parts. Each pathway is decomposed into all its constituent sub-paths, which in turn are matched with corresponding gene expression profiles. The highly ranked, and phenotype inclined sub-paths are kept. Apart from the pathway analysis algorithm, the fundamental innovation of the MinePath web-server concerns its advanced visualization and interactive capabilities. To our knowledge, this is the first pathway analysis server that introduces and offers visualization of the underlying and active pathway regulatory mechanisms instead of genes. Other features include live interaction, immediate visualization of functional sub-paths per phenotype and dynamic linked annotations for the engaged genes and molecular relations. The user can download not only the results but also the corresponding web viewer framework of the performed analysis. This feature provides the flexibility to immediately publish results without publishing source/expression data, and get all the functionality of a web based pathway analysis viewer. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Genetic interactions within inositol-related pathways are associated with longitudinal changes in ventricle size

PubMed Central

Koran, Mary Ellen I.; Hohman, Timothy J.; Meda, Shashwath A.; Thornton-Wells, Tricia A.

2013-01-01

The genetic etiology of late onset Alzheimer disease (LOAD) has proven complex, involving clinical and genetic heterogeneity and gene-gene interactions. Recent genome wide association studies (GWAS) in LOAD have led to the discovery of novel genetic risk factors; however, the investigation of gene-gene interactions has been limited. Conventional genetic studies often use binary disease status as the primary phenotype, but for complex brain-based diseases, neuroimaging data can serve as quantitative endophenotypes that correlate with disease status and closely reflect pathological changes. In the Alzheimer's Disease Neuroimaging Initiative (ADNI) cohort, we tested for association of genetic interactions with longitudinal MRI measurements of the inferior lateral ventricles (ILVs), which have repeatedly shown a relationship to LOAD status and progression. We performed linear regression to evaluate the ability of pathway-derived SNP-SNP pairs to predict the slope of change in volume of the ILVs. After Bonferroni correction, we identified four significant interactions in the right ILV (RILV) corresponding to gene-gene pairs SYNJ2-PI4KA, PARD3-MYH2, PDE3A-ABHD12B and OR2L13-PRKG1 and one significant interaction in the left ILV (LILV) corresponding to SYNJ2-PI4KA. The SNP-SNP interaction corresponding to SYNJ2-PI4KA was identical in the RILV and LILV and was the most significant interaction in each (RILV: p=9.10×10−12; LILV: p=8.20×10−13). Both genes belong to the inositol phosphate signaling pathway which has been previously associated with neurodegeneration in AD and we discuss the possibility that perturbation of this pathway results in a down-regulation of the Akt cell survival pathway and, thereby, decreased neuronal survival, as reflected by increased volume of the ventricles. PMID:24077433

DNA requirements for interaction of the C-terminal region of Ku80 with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs).

PubMed

Radhakrishnan, Sarvan Kumar; Lees-Miller, Susan P

2017-09-01

Non-homologous end joining (NHEJ) is the major pathway for the repair of ionizing radiation induced DNA double strand breaks (DSBs) in human cells. Critical to NHEJ is the DNA-dependent interaction of the Ku70/80 heterodimer with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to form the DNA-PK holoenzyme. However, precisely how Ku recruits DNA-PKcs to DSBs ends to enhance its kinase activity has remained enigmatic, with contradictory findings reported in the literature. Here we address the role of the Ku80 C-terminal region (CTR) in the DNA-dependent interaction of Ku70/80 with DNA-PKcs using purified components and defined DNA structures. Our results show that the Ku80 CTR is required for interaction with DNA-PKcs on short segments of blunt ended 25bp dsDNA or 25bp dsDNA with a 15-base poly dA single stranded (ss) DNA extension, but this requirement is less stringent on longer dsDNA molecules (35bp blunt ended dsDNA) or 25bp duplex DNA with either a 15-base poly dT or poly dC ssDNA extension. Moreover, the DNA-PKcs-Ku complex preferentially forms on 25 bp DNA with a poly-pyrimidine ssDNA extension.Our work clarifies the role of the Ku80 CTR and dsDNA ends on the interaction of DNA-PKcs with Ku and provides key information to guide assembly and biology of NHEJ complexes. Copyright © 2017 Elsevier B.V. All rights reserved.

Electrostatic Interactions in the Binding Pathway of a Transient Protein Complex Studied by NMR and Isothermal Titration Calorimetry*

PubMed Central

Meneses, Erick; Mittermaier, Anthony

2014-01-01

Much of our knowledge of protein binding pathways is derived from extremely stable complexes that interact very tightly, with lifetimes of hours to days. Much less is known about weaker interactions and transient complexes because these are challenging to characterize experimentally. Nevertheless, these types of interactions are ubiquitous in living systems. The combination of NMR relaxation dispersion Carr–Purcell–Meiboom–Gill (CPMG) experiments and isothermal titration calorimetry allows the quantification of rapid binding kinetics for complexes with submillisecond lifetimes that are difficult to study using conventional techniques. We have used this approach to investigate the binding pathway of the Src homology 3 (SH3) domain from the Fyn tyrosine kinase, which forms complexes with peptide targets whose lifetimes are on the order of about a millisecond. Long range electrostatic interactions have been shown to play a critical role in the binding pathways of tightly binding complexes. The role of electrostatics in the binding pathways of transient complexes is less well understood. Similarly to previously studied tight complexes, we find that SH3 domain association rates are enhanced by long range electrostatics, whereas short range interactions are formed late in the docking process. However, the extent of electrostatic association rate enhancement is several orders of magnitudes less, whereas the electrostatic-free basal association rate is significantly greater. Thus, the SH3 domain is far less reliant on electrostatic enhancement to achieve rapid association kinetics than are previously studied systems. This suggests that there may be overall differences in the role played by electrostatics in the binding pathways of extremely stable versus transient complexes. PMID:25122758

Identification of core pathways based on attractor and crosstalk in ischemic stroke.

PubMed

Diao, Xiufang; Liu, Aijuan

2018-02-01

Ischemic stroke is a leading cause of mortality and disability around the world. It is an important task to identify dysregulated pathways which infer molecular and functional insights existing in high-throughput experimental data. Gene expression profile of E-GEOD-16561 was collected. Pathways were obtained from the database of Kyoto Encyclopedia of Genes and Genomes and Retrieval of Interacting Genes was used to download protein-protein interaction sets. Attractor and crosstalk approaches were applied to screen dysregulated pathways. A total of 20 differentially expressed genes were identified in ischemic stroke. Thirty-nine significant differential pathways were identified according to P<0.01 and 28 pathways were identified with RP<0.01 and 17 pathways were identified with impact factor >250. On the basis of the three criteria, 11 significant dysfunctional pathways were identified. Among them, Epstein-Barr virus infection was the most significant differential pathway. In conclusion, with the method based on attractor and crosstalk, significantly dysfunctional pathways were identified. These pathways are expected to provide molecular mechanism of ischemic stroke and represents a novel potential therapeutic target for ischemic stroke treatment.

H3K9me2/3 Binding of the MBT Domain Protein LIN-61 Is Essential for Caenorhabditis elegans Vulva Development

PubMed Central

Koester-Eiserfunke, Nora; Fischle, Wolfgang

2011-01-01

MBT domain proteins are involved in developmental processes and tumorigenesis. In vitro binding and mutagenesis studies have shown that individual MBT domains within clustered MBT repeat regions bind mono- and dimethylated histone lysine residues with little to no sequence specificity but discriminate against the tri- and unmethylated states. However, the exact function of promiscuous histone methyl-lysine binding in the biology of MBT domain proteins has not been elucidated. Here, we show that the Caenorhabditis elegans four MBT domain protein LIN-61, in contrast to other MBT repeat factors, specifically interacts with histone H3 when methylated on lysine 9, displaying a strong preference for di- and trimethylated states (H3K9me2/3). Although the fourth MBT repeat is implicated in this interaction, H3K9me2/3 binding minimally requires MBT repeats two to four. Further, mutagenesis of residues conserved with other methyl-lysine binding MBT regions in the fourth MBT repeat does not abolish interaction, implicating a distinct binding mode. In vivo, H3K9me2/3 interaction of LIN-61 is required for C. elegans vulva development within the synMuvB pathway. Mutant LIN-61 proteins deficient in H3K9me2/3 binding fail to rescue lin-61 synMuvB function. Also, previously identified point mutant synMuvB alleles are deficient in H3K9me2/3 interaction although these target residues that are outside of the fourth MBT repeat. Interestingly, lin-61 genetically interacts with two other synMuvB genes, hpl-2, an HP1 homologous H3K9me2/3 binding factor, and met-2, a SETDB1 homologous H3K9 methyl transferase (H3K9MT), in determining C. elegans vulva development and fertility. Besides identifying the first sequence specific and di-/trimethylation binding MBT domain protein, our studies imply complex multi-domain regulation of ligand interaction of MBT domains. Our results also introduce a mechanistic link between LIN-61 function and biology, and they establish interplay of the H3K9me2/3 binding proteins, LIN-61 and HPL-2, as well as the H3K9MT MET-2 in distinct developmental pathways. PMID:21437264

Pollen Killer Gene S35 Function Requires Interaction with an Activator That Maps Close to S24, Another Pollen Killer Gene in Rice.

PubMed

Kubo, Takahiko; Yoshimura, Atsushi; Kurata, Nori

2016-05-03

Pollen killer genes disable noncarrier pollens, and are responsible for male sterility and segregation distortion in hybrid populations of distantly related plant species. The genetic networks and the molecular mechanisms underlying the pollen killer system remain largely unknown. Two pollen killer genes, S24 and S35, have been found in an intersubspecific cross of Oryza sativa ssp. indica and japonica The effect of S24 is counteracted by an unlinked locus EFS Additionally, S35 has been proposed to interact with S24 to induce pollen sterility. These genetic interactions are suggestive of a single S24-centric genetic pathway (EFS-S24-S35) for the pollen killer system. To examine this hypothetical genetic pathway, the S35 and the S24 regions were further characterized and genetically dissected in this study. Our results indicated that S35 causes pollen sterility independently of both the EFS and S24 genes, but is dependent on a novel gene close to the S24 locus, named incentive for killing pollen (INK). We confirmed the phenotypic effect of the INK gene separately from the S24 gene, and identified the INK locus within an interval of less than 0.6 Mb on rice chromosome 5. This study characterized the genetic effect of the two independent genetic pathways of INK-S35 and EFS-S24 in indica-japonica hybrid progeny. Our results provide clear evidence that hybrid male sterility in rice is caused by several pollen killer networks with multiple factors positively and negatively regulating pollen killer genes. Copyright © 2016 Kubo et al.

Pollen Killer Gene S35 Function Requires Interaction with an Activator That Maps Close to S24, Another Pollen Killer Gene in Rice

PubMed Central

Kubo, Takahiko; Yoshimura, Atsushi; Kurata, Nori

2016-01-01

Pollen killer genes disable noncarrier pollens, and are responsible for male sterility and segregation distortion in hybrid populations of distantly related plant species. The genetic networks and the molecular mechanisms underlying the pollen killer system remain largely unknown. Two pollen killer genes, S24 and S35, have been found in an intersubspecific cross of Oryza sativa ssp. indica and japonica. The effect of S24 is counteracted by an unlinked locus EFS. Additionally, S35 has been proposed to interact with S24 to induce pollen sterility. These genetic interactions are suggestive of a single S24-centric genetic pathway (EFS–S24–S35) for the pollen killer system. To examine this hypothetical genetic pathway, the S35 and the S24 regions were further characterized and genetically dissected in this study. Our results indicated that S35 causes pollen sterility independently of both the EFS and S24 genes, but is dependent on a novel gene close to the S24 locus, named incentive for killing pollen (INK). We confirmed the phenotypic effect of the INK gene separately from the S24 gene, and identified the INK locus within an interval of less than 0.6 Mb on rice chromosome 5. This study characterized the genetic effect of the two independent genetic pathways of INK–S35 and EFS–S24 in indica–japonica hybrid progeny. Our results provide clear evidence that hybrid male sterility in rice is caused by several pollen killer networks with multiple factors positively and negatively regulating pollen killer genes. PMID:27172610

Growth of chronic myeloid leukemia cells is inhibited by infection with Ad-SH2-HA adenovirus that disrupts Grb2-Bcr-Abl complexes.

PubMed

Peng, Zhi; Luo, Hong-Wei; Yuan, Ying; Shi, Jing; Huang, Shi-Feng; Li, Chun-Li; Cao, Wei-Xi; Huang, Zong-Gan; Feng, Wen-Li

2011-05-01

The persistence of Bcr-Abl-positive cells in patients on imatinib therapy indicates that inhibition of the Bcr-Abl kinase activity alone might not be sufficient to eradicate the leukemia cells. Many downstream effectors of Bcr-Abl have been described, including activation of both the Grb2-SoS-Ras-MAPK and Grb2-Gab2-PI3K-Akt pathways. The Bcr-Abl-Grb2 interaction, which is mediated by the direct interaction of the Grb2 SH2 domain with the phospho-Bcr-Abl Y177, is required for activation of these signaling pathways. Therefore, disrupting their interaction represents a potential therapeutic strategy for inhibiting the oncogenic downstream signals of Bcr-Abl. Adenovirus Ad-SH2-HA expressing the Grb2 SH2 domain was constructed and applied in this study. As expected, Ad-SH2-HA efficiently infected CML cells and functioned by binding to the phospho-Bcr-Abl Y177 site, competitively disrupting the Grb2 SH2-phospho-Bcr-Abl Y177 complex. They induced potent anti-proliferation and apoptosis-inducing effects in CML cell lines. Moreover, the Ras, MAPK and Akt activities were significantly reduced in the Ad-SH2-HA treated cells. These were not observed with the point-mutated control adenovirus Ad-Sm-HA with abolished phospho-Bcr-Abl Y177 binding sites. These data indicate that, in addition to the direct targeting of Bcr-Abl, selective inhibition of its downstream signaling pathways may be a therapeutic option for CML, and the Ad-SH2-HA-mediated killing strategy could be explored as a promising anti-leukemia agent in CML.

Network-based study reveals potential infection pathways of hepatitis-C leading to various diseases.

PubMed

Mukhopadhyay, Anirban; Maulik, Ujjwal

2014-01-01

Protein-protein interaction network-based study of viral pathogenesis has been gaining popularity among computational biologists in recent days. In the present study we attempt to investigate the possible pathways of hepatitis-C virus (HCV) infection by integrating the HCV-human interaction network, human protein interactome and human genetic disease association network. We have proposed quasi-biclique and quasi-clique mining algorithms to integrate these three networks to identify infection gateway host proteins and possible pathways of HCV pathogenesis leading to various diseases. Integrated study of three networks, namely HCV-human interaction network, human protein interaction network, and human proteins-disease association network reveals potential pathways of infection by the HCV that lead to various diseases including cancers. The gateway proteins have been found to be biologically coherent and have high degrees in human interactome compared to the other virus-targeted proteins. The analyses done in this study provide possible targets for more effective anti-hepatitis-C therapeutic involvement.

Network-Based Study Reveals Potential Infection Pathways of Hepatitis-C Leading to Various Diseases

PubMed Central

Mukhopadhyay, Anirban; Maulik, Ujjwal

2014-01-01

Protein-protein interaction network-based study of viral pathogenesis has been gaining popularity among computational biologists in recent days. In the present study we attempt to investigate the possible pathways of hepatitis-C virus (HCV) infection by integrating the HCV-human interaction network, human protein interactome and human genetic disease association network. We have proposed quasi-biclique and quasi-clique mining algorithms to integrate these three networks to identify infection gateway host proteins and possible pathways of HCV pathogenesis leading to various diseases. Integrated study of three networks, namely HCV-human interaction network, human protein interaction network, and human proteins-disease association network reveals potential pathways of infection by the HCV that lead to various diseases including cancers. The gateway proteins have been found to be biologically coherent and have high degrees in human interactome compared to the other virus-targeted proteins. The analyses done in this study provide possible targets for more effective anti-hepatitis-C therapeutic involvement. PMID:24743187

Hybrid drug combination: Combination of ferulic acid and metformin as anti-diabetic therapy.

PubMed

Nankar, Rakesh; Prabhakar, P K; Doble, Mukesh

2017-12-15

Ferulic acid, an anti-oxidant phytochemical present in several dietary components, is known to produce wide range of pharmacological effects. It is approved for use in food industry as a preservative and in sports food. Previous reports from our lab have shown synergistic interaction of ferulic acid with metformin in cell lines and diabetic rats. The purpose of this review is to compile information about anti-diabetic activity of ferulic acid in in vitro and in vivo models with special emphasis on activity of ferulic acid when combined with metformin. The mechanism of synergistic interaction between ferulic acid and metformin is also proposed after carefully studying effects of these compounds on molecules involved in glucose metabolism. Scientific literature for the purpose of this review was collected using online search engines and databases such as ScienceDirect, Scopus, PubMed and Google scholar. Ferulic acid forms resonance stabilized phenoxyl radical which scavenges free radicals and reduce oxidative stress. It improves glucose and lipid profile in diabetic rats by enhancing activities of antioxidant enzymes, superoxide dismutase and catalase in the pancreatic tissue. Combining ferulic acid with metformin improves both, in vitro glucose uptake activity and in vivo hypoglycemic activity of the latter. It is possible to reduce the dose of metformin by four folds (from 50 to 12.5 mg/kg body weight) by combining it with 10 mg of ferulic acid/kg body weight in diabetic rats. Ferulic acid improves glucose uptake through PI3-K pathway whereas metformin activates AMPK pathway to improve glucose uptake. The synergistic interaction of ferulic acid and metformin is due their action on parallel pathways which are involved in glucose uptake. Due to synergistic nature of their interaction, it is possible to reduce the dose of metformin (by combining with ferulic acid) required to achieve normoglycemia. Since the dose of metformin is reduced, the dose associated side effects of metformin therapy can be reduced. Copyright © 2017 Elsevier GmbH. All rights reserved.

Drosophila VAMP7 regulates Wingless intracellular trafficking.

PubMed

Gao, Han; He, Fang; Lin, Xinhua; Wu, Yihui

2017-01-01

Drosophila Wingless (Wg) is a morphogen that determines cell fate during development. Previous studies have shown that endocytic pathways regulate Wg trafficking and signaling. Here, we showed that loss of vamp7, a gene required for vesicle fusion, dramatically increased Wg levels and decreased Wg signaling. Interestingly, we found that levels of Dally-like (Dlp), a glypican that can interact with Wg to suppress Wg signaling at the dorsoventral boundary of the Drosophila wing, were also increased in vamp7 mutant cells. Moreover, Wg puncta in Rab4-dependent recycling endosomes were Dlp positive. We hypothesize that VAMP7 is required for Wg intracellular trafficking and the accumulation of Wg in Rab4-dependent recycling endosomes might affect Wg signaling.

The Caenorhabditis elegans RDE-10/RDE-11 complex regulates RNAi by promoting secondary siRNA amplification.

PubMed

Zhang, Chi; Montgomery, Taiowa A; Fischer, Sylvia E J; Garcia, Susana M D A; Riedel, Christian G; Fahlgren, Noah; Sullivan, Christopher M; Carrington, James C; Ruvkun, Gary

2012-05-22

In nematodes, plants, and fungi, RNAi is remarkably potent and persistent due to the amplification of initial silencing signals by RNA-dependent RNA polymerases (RdRPs). In Caenorhabditis elegans (C. elegans), the interaction between the RNA-induced silencing complex (RISC) loaded with primary small interfering RNAs (siRNAs) and the target messenger RNA (mRNA) leads to the recruitment of RdRPs and synthesis of secondary siRNAs using the target mRNA as the template. The mechanism and genetic requirements for secondary siRNA accumulation are not well understood. From a forward genetic screen for C. elegans genes required for RNAi, we identified rde-10, and through proteomic analysis of RDE-10-interacting proteins, we identified a protein complex containing the new RNAi factor RDE-11, the known RNAi factors RSD-2 and ERGO-1, and other candidate RNAi factors. The RNAi defective genes rde-10 and rde-11 encode a novel protein and a RING-type zinc finger domain protein, respectively. Mutations in rde-10 and rde-11 genes cause dosage-sensitive RNAi deficiencies: these mutants are resistant to low dosage but sensitive to high dosage of double-stranded RNAs. We assessed the roles of rde-10, rde-11, and other dosage-sensitive RNAi-defective genes rsd-2, rsd-6, and haf-6 in both exogenous and endogenous small RNA pathways using high-throughput sequencing and qRT-PCR. These genes are required for the accumulation of secondary siRNAs in both exogenous and endogenous RNAi pathways. The RDE-10/RDE-11 complex is essential for the amplification of RNAi in C. elegans by promoting secondary siRNA accumulation. Copyright © 2012 Elsevier Ltd. All rights reserved.

The Caenorhabditis elegans RDE-10/RDE-11 complex regulates RNAi by promoting secondary siRNA amplification

PubMed Central

Zhang, Chi; Montgomery, Taiowa A.; Fischer, Sylvia E. J.; Garcia, Susana M. D. A.; Riedel, Christian G.; Fahlgren, Noah; Sullivan, Christopher M.; Carrington, James C.; Ruvkun, Gary

2012-01-01

SUMMARY Background In nematodes, plants and fungi, RNAi is remarkably potent and persistent due to the amplification of initial silencing signals by RNA-dependent RNA polymerases (RdRPs). In Caenorhabditis elegans (C. elegans), the interaction between the RNA-induced silencing complex (RISC) loaded with primary siRNAs and the target mRNA leads to the recruitment of RdRPs and synthesis of secondary siRNAs using the target mRNA as the template. The mechanism and genetic requirements for secondary siRNA accumulation are not well understood. Results From a forward genetic screen for C. elegans genes required for RNAi, we identified rde-10 and through proteomic analysis of RDE-10-interacting proteins, we identified a protein complex containing the new RNAi factor RDE-11, the known RNAi factors RSD-2 and ERGO-1, as well as other candidate RNAi factors. The RNAi defective genes rde-10 and rde-11 encode a novel protein and a RING-type zinc finger domain protein, respectively. Mutations in rde-10 and rde-11 genes cause dosage-sensitive RNAi deficiencies: these mutants are resistant to low dosage, but sensitive to high dosage of double-stranded RNAs (dsRNAs). We assessed the roles of rde-10, rde-11, and other dosage-sensitive RNAi-defective genes rsd-2, rsd-6 and haf-6 in both exogenous and endogenous small RNA pathways using high-throughput sequencing and qRT-PCR. These genes are required for the accumulation of secondary siRNAs in both exogenous and endogenous RNAi pathways. Conclusions The RDE-10/RDE-11 complex is essential for the amplification of RNAi in C. elegans by promoting secondary siRNA accumulation. PMID:22542102

PUB1 Interacts with the Receptor Kinase DMI2 and Negatively Regulates Rhizobial and Arbuscular Mycorrhizal Symbioses through Its Ubiquitination Activity in Medicago truncatula.

PubMed

Vernié, Tatiana; Camut, Sylvie; Camps, Céline; Rembliere, Céline; de Carvalho-Niebel, Fernanda; Mbengue, Malick; Timmers, Ton; Gasciolli, Virginie; Thompson, Richard; le Signor, Christine; Lefebvre, Benoit; Cullimore, Julie; Hervé, Christine

2016-04-01

PUB1, an E3 ubiquitin ligase, which interacts with and is phosphorylated by the LYK3 symbiotic receptor kinase, negatively regulates rhizobial infection and nodulation during the nitrogen-fixing root nodule symbiosis in Medicago truncatula In this study, we show that PUB1 also interacts with and is phosphorylated by DOES NOT MAKE INFECTIONS 2, the key symbiotic receptor kinase of the common symbiosis signaling pathway, required for both the rhizobial and the arbuscular mycorrhizal (AM) endosymbioses. We also show here that PUB1 expression is activated during successive stages of root colonization by Rhizophagus irregularis that is compatible with its interaction with DOES NOT MAKE INFECTIONS 2. Through characterization of a mutant, pub1-1, affected by the E3 ubiquitin ligase activity of PUB1, we have shown that the ubiquitination activity of PUB1 is required to negatively modulate successive stages of infection and development of rhizobial and AM symbioses. In conclusion, PUB1 represents, to our knowledge, a novel common component of symbiotic signaling integrating signal perception through interaction with and phosphorylation by two key symbiotic receptor kinases, and downstream signaling via its ubiquitination activity to fine-tune both rhizobial and AM root endosymbioses. © 2016 American Society of Plant Biologists. All Rights Reserved.

DNA packaging and the pathway of bacteriophage T4 head assembly.

PubMed Central

Hsiao, C L; Black, L W

1977-01-01

A cold-sensitive mutation in the structural gene for a minor phage T4 capsid protein (p20) leads to formation of heads containing p20 and cleaved head proteins and empty of DNA. Such heads can be filled with DNA and converted to active phages in vivo uponshift to high temperature. It appears that p20 has two distinct roles in head assembly: first, in construction of the prehead shell (blocked by ts and am mutation) and, second,in DNA packaging (blocked by cs mutation). The latter function is closely associated with gene 17 product, previously known to be required for DNA packagaing. Temperature shift studies of cs-ts double mutants and other observations allow determination of phage function required for DNA packaging. Contrary to previous proposals, we find that T4 DNA packaging is not directly coupled to and can follow DNA synthesis, protein cleavage, prehead core removal, and gene 21-mediated cleavage-induced increase in head volume. Our evidence suggests that an altered head assembly pathway exists and that DNA packaging is probably initiated by DNA-capsid (p20) interaction. Images PMID:269421

Brassinosteroids

PubMed Central

Clouse, Steven D.

2011-01-01

Brassinosteroids (BRs) are endogenous plant hormones essential for the proper regulation of multiple physiological processes required for normal plant growth and development. Since their discovery more than 30 years ago, extensive research on the mechanisms of BR action using biochemistry, mutant studies, proteomics and genome-wide transcriptome analyses, has helped refine the BR biosynthetic pathway, identify the basic molecular components required to relay the BR signal from perception to gene regulation, and expand the known physiological responses influenced by BRs. These mechanistic advances have helped answer the intriguing question of how BRs can have such dramatic pleiotropic effects on a broad range of diverse developmental pathways and have further pointed to BR interactions with other plant hormones and environmental cues. This chapter briefly reviews historical aspects of BR research and then summarizes the current state of knowledge on BR biosynthesis, metabolism and signal transduction. Recent studies uncovering novel phosphorelays and gene regulatory networks through which BR influences both vegetative and reproductive development are examined and placed in the context of known BR physiological responses including cell elongation and division, vascular differentiation, flowering, pollen development and photomorphogenesis. PMID:22303275

An Interactive Platform to Visualize Data-Driven Clinical Pathways for the Management of Multiple Chronic Conditions.

PubMed

Zhang, Yiye; Padman, Rema

2017-01-01

Patients with multiple chronic conditions (MCC) pose an increasingly complex health management challenge worldwide, particularly due to the significant gap in our understanding of how to provide coordinated care. Drawing on our prior research on learning data-driven clinical pathways from actual practice data, this paper describes a prototype, interactive platform for visualizing the pathways of MCC to support shared decision making. Created using Python web framework, JavaScript library and our clinical pathway learning algorithm, the visualization platform allows clinicians and patients to learn the dominant patterns of co-progression of multiple clinical events from their own data, and interactively explore and interpret the pathways. We demonstrate functionalities of the platform using a cluster of 36 patients, identified from a dataset of 1,084 patients, who are diagnosed with at least chronic kidney disease, hypertension, and diabetes. Future evaluation studies will explore the use of this platform to better understand and manage MCC.

Tel2 mediates activation and localization of ATM/Tel1 kinase to a double-strand break.

PubMed

Anderson, Carol M; Korkin, Dmitry; Smith, Dana L; Makovets, Svetlana; Seidel, Jeffrey J; Sali, Andrej; Blackburn, Elizabeth H

2008-04-01

The kinases ATM and ATR (Tel1 and Mec1 in the yeast Saccharomyces cerevisiae) control the response to DNA damage. We report that S. cerevisiae Tel2 acts at an early step of the TEL1/ATM pathway of DNA damage signaling. We show that Tel1 and Tel2 interact, and that even when Tel1 protein levels are high, this interaction is specifically required for Tel1 localization to a DNA break and its activation of downstream targets. Computational analysis revealed structural homology between Tel2 and Ddc2 (ATRIP in vertebrates), a partner of Mec1, suggesting a common structural principle used by partners of phoshoinositide 3-kinase-like kinases.

Characterization of a unique motif in LIM mineralization protein-1 that interacts with jun activation-domain-binding protein 1.

PubMed

Sangadala, Sreedhara; Yoshioka, Katsuhito; Enyo, Yoshio; Liu, Yunshan; Titus, Louisa; Boden, Scott D

2014-01-01

Development and repair of the skeletal system and other organs are highly dependent on precise regulation of the bone morphogenetic protein (BMP) pathway. The use of BMPs clinically to induce bone formation has been limited in part by the requirement of much higher doses of recombinant proteins in primates than were needed in cell culture or rodents. Therefore, increasing cellular responsiveness to BMPs has become our focus. We determined that an osteogenic LIM mineralization protein, LMP-1 interacts with Smurf1 (Smad ubiquitin regulatory factor 1) and prevents ubiquitination of Smads resulting in potentiation of BMP activity. In the region of LMP-1 responsible for bone formation, there is a motif that directly interacts with the Smurf1 WW2 domain and thus effectively competes for binding with Smad1 and Smad5, key signaling proteins in the BMP pathway. Here we show that the same region also contains a motif that interacts with Jun activation-domain-binding protein 1 (Jab1) which targets a common Smad, Smad4, shared by both the BMP and transforming growth factor-β (TGF-β) pathways, for proteasomal degradation. Jab1 was first identified as a coactivator of the transcription factor c-Jun. Jab1 binds to Smad4, Smad5, and Smad7, key intracellular signaling molecules of the TGF-β superfamily, and causes ubiquitination and/or degradation of these Smads. We confirmed a direct interaction of Jab1 with LMP-1 using recombinantly expressed wild-type and mutant proteins in slot-blot-binding assays. We hypothesized that LMP-1 binding to Jab1 prevents the binding and subsequent degradation of these Smads causing increased accumulation of osteogenic Smads in cells. We identified a sequence motif in LMP-1 that was predicted to interact with Jab1 based on the MAME/MAST sequence analysis of several cellular signaling molecules that are known to interact with Jab-1. We further mutated the potential key interacting residues in LMP-1 and showed loss of binding to Jab1 in binding assays in vitro. The activities of various wild-type and mutant LMP-1 proteins were evaluated using a BMP-responsive luciferase reporter and alkaline phosphatase assay in mouse myoblastic cells that were differentiated toward the osteoblastic phenotype. Finally, to strengthen physiological relevance of LMP-1 and Jab1 interaction, we showed that overexpression of LMP-1 caused nuclear accumulation of Smad4 upon BMP treatment which is reflective of increased Smad signaling in cells.

A Model of an Integrated Immune System Pathway in Homo sapiens and Its Interaction with Superantigen Producing Expression Regulatory Pathway in Staphylococcus aureus: Comparing Behavior of Pathogen Perturbed and Unperturbed Pathway

PubMed Central

Tomar, Namrata; De, Rajat K.

2013-01-01

Response of an immune system to a pathogen attack depends on the balance between the host immune defense and the virulence of the pathogen. Investigation of molecular interactions between the proteins of a host and a pathogen helps in identifying the pathogenic proteins. It is necessary to understand the dynamics of a normally behaved host system to evaluate the capacity of its immune system upon pathogen attack. In this study, we have compared the behavior of an unperturbed and pathogen perturbed host system. Moreover, we have developed a formalism under Flux Balance Analysis (FBA) for the optimization of conflicting objective functions. We have constructed an integrated pathway system, which includes Staphylococcal Superantigen (SAg) expression regulatory pathway and TCR signaling pathway of Homo sapiens. We have implemented the method on this pathway system and observed the behavior of host signaling molecules upon pathogen attack. The entire study has been divided into six different cases, based on the perturbed/unperturbed conditions. In other words, we have investigated unperturbed and pathogen perturbed human TCR signaling pathway, with different combinations of optimization of concentrations of regulatory and signaling molecules. One of these cases has aimed at finding out whether minimization of the toxin production in a pathogen leads to the change in the concentration levels of the proteins coded by TCR signaling pathway genes in the infected host. Based on the computed results, we have hypothesized that the balance between TCR signaling inhibitory and stimulatory molecules can keep TCR signaling system into resting/stimulating state, depending upon the perturbation. The proposed integrated host-pathogen interaction pathway model has accurately reflected the experimental evidences, which we have used for validation purpose. The significance of this kind of investigation lies in revealing the susceptible interaction points that can take back the Staphylococcal Enterotoxin (SE)-challenged system within the range of normal behavior. PMID:24324645

Predictive Genomic Analyses Inform the Basis for Vitamin Metabolism and Provisioning in Bacteria-Arthropod Endosymbioses.

PubMed

Serbus, Laura R; Rodriguez, Brian Garcia; Sharmin, Zinat; Momtaz, A J M Zehadee; Christensen, Steen

2017-06-07

The requirement of vitamins for core metabolic processes creates a unique set of pressures for arthropods subsisting on nutrient-limited diets. While endosymbiotic bacteria carried by arthropods have been widely implicated in vitamin provisioning, the underlying molecular mechanisms are not well understood. To address this issue, standardized predictive assessment of vitamin metabolism was performed in 50 endosymbionts of insects and arachnids. The results predicted that arthropod endosymbionts overall have little capacity for complete de novo biosynthesis of conventional or active vitamin forms. Partial biosynthesis pathways were commonly predicted, suggesting a substantial role in vitamin provisioning. Neither taxonomic relationships between host and symbiont, nor the mode of host-symbiont interaction were clear predictors of endosymbiont vitamin pathway capacity. Endosymbiont genome size and the synthetic capacity of nonsymbiont taxonomic relatives were more reliable predictors. We developed a new software application that also predicted that last-step conversion of intermediates into active vitamin forms may contribute further to vitamin biosynthesis by endosymbionts. Most instances of predicted vitamin conversion were paralleled by predictions of vitamin use. This is consistent with achievement of provisioning in some cases through upregulation of pathways that were retained for endosymbiont benefit. The predicted absence of other enzyme classes further suggests a baseline of vitamin requirement by the majority of endosymbionts, as well as some instances of putative mutualism. Adaptation of this workflow to analysis of other organisms and metabolic pathways will provide new routes for considering the molecular basis for symbiosis on a comprehensive scale. Copyright © 2017 Serbus et al.

Network-Based Identification of Adaptive Pathways in Evolved Ethanol-Tolerant Bacterial Populations

PubMed Central

Swings, Toon; Weytjens, Bram; Schalck, Thomas; Bonte, Camille; Verstraeten, Natalie; Michiels, Jan

2017-01-01

Abstract Efficient production of ethanol for use as a renewable fuel requires organisms with a high level of ethanol tolerance. However, this trait is complex and increased tolerance therefore requires mutations in multiple genes and pathways. Here, we use experimental evolution for a system-level analysis of adaptation of Escherichia coli to high ethanol stress. As adaptation to extreme stress often results in complex mutational data sets consisting of both causal and noncausal passenger mutations, identifying the true adaptive mutations in these settings is not trivial. Therefore, we developed a novel method named IAMBEE (Identification of Adaptive Mutations in Bacterial Evolution Experiments). IAMBEE exploits the temporal profile of the acquisition of mutations during evolution in combination with the functional implications of each mutation at the protein level. These data are mapped to a genome-wide interaction network to search for adaptive mutations at the level of pathways. The 16 evolved populations in our data set together harbored 2,286 mutated genes with 4,470 unique mutations. Analysis by IAMBEE significantly reduced this number and resulted in identification of 90 mutated genes and 345 unique mutations that are most likely to be adaptive. Moreover, IAMBEE not only enabled the identification of previously known pathways involved in ethanol tolerance, but also identified novel systems such as the AcrAB-TolC efflux pump and fatty acids biosynthesis and even allowed to gain insight into the temporal profile of adaptation to ethanol stress. Furthermore, this method offers a solid framework for identifying the molecular underpinnings of other complex traits as well. PMID:28961727

Thyroid Hormone Regulation of Metabolism

PubMed Central

Mullur, Rashmi; Liu, Yan-Yun

2014-01-01

Thyroid hormone (TH) is required for normal development as well as regulating metabolism in the adult. The thyroid hormone receptor (TR) isoforms, α and β, are differentially expressed in tissues and have distinct roles in TH signaling. Local activation of thyroxine (T4), to the active form, triiodothyronine (T3), by 5′-deiodinase type 2 (D2) is a key mechanism of TH regulation of metabolism. D2 is expressed in the hypothalamus, white fat, brown adipose tissue (BAT), and skeletal muscle and is required for adaptive thermogenesis. The thyroid gland is regulated by thyrotropin releasing hormone (TRH) and thyroid stimulating hormone (TSH). In addition to TRH/TSH regulation by TH feedback, there is central modulation by nutritional signals, such as leptin, as well as peptides regulating appetite. The nutrient status of the cell provides feedback on TH signaling pathways through epigentic modification of histones. Integration of TH signaling with the adrenergic nervous system occurs peripherally, in liver, white fat, and BAT, but also centrally, in the hypothalamus. TR regulates cholesterol and carbohydrate metabolism through direct actions on gene expression as well as cross-talk with other nuclear receptors, including peroxisome proliferator-activated receptor (PPAR), liver X receptor (LXR), and bile acid signaling pathways. TH modulates hepatic insulin sensitivity, especially important for the suppression of hepatic gluconeogenesis. The role of TH in regulating metabolic pathways has led to several new therapeutic targets for metabolic disorders. Understanding the mechanisms and interactions of the various TH signaling pathways in metabolism will improve our likelihood of identifying effective and selective targets. PMID:24692351

An overview of bioinformatics methods for modeling biological pathways in yeast.

PubMed

Hou, Jie; Acharya, Lipi; Zhu, Dongxiao; Cheng, Jianlin

2016-03-01

The advent of high-throughput genomics techniques, along with the completion of genome sequencing projects, identification of protein-protein interactions and reconstruction of genome-scale pathways, has accelerated the development of systems biology research in the yeast organism Saccharomyces cerevisiae In particular, discovery of biological pathways in yeast has become an important forefront in systems biology, which aims to understand the interactions among molecules within a cell leading to certain cellular processes in response to a specific environment. While the existing theoretical and experimental approaches enable the investigation of well-known pathways involved in metabolism, gene regulation and signal transduction, bioinformatics methods offer new insights into computational modeling of biological pathways. A wide range of computational approaches has been proposed in the past for reconstructing biological pathways from high-throughput datasets. Here we review selected bioinformatics approaches for modeling biological pathways inS. cerevisiae, including metabolic pathways, gene-regulatory pathways and signaling pathways. We start with reviewing the research on biological pathways followed by discussing key biological databases. In addition, several representative computational approaches for modeling biological pathways in yeast are discussed. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Interaction of Human Cytomegalovirus Tegument Proteins ppUL35 and ppUL35A with Sorting Nexin 5 Regulates Glycoprotein B (gpUL55) Localization.

PubMed

Maschkowitz, Gregor; Gärtner, Sabine; Hofmann-Winkler, Heike; Fickenscher, Helmut; Winkler, Michael

2018-05-01

Human cytomegalovirus (HCMV) is a widespread human pathogen that causes asymptomatic infection in healthy individuals but poses a serious threat to immunocompromised patients. During the late phase of HCMV infection, the viral capsid is transported to the cytoplasmic viral assembly center (cVAC), where it is enclosed by the tegument protein layer and the viral envelope. The cVAC consists of circularly arranged vesicles from the trans -Golgi and endosomal networks. The HCMV gene UL35 encodes ppUL35 and its shorter form, ppUL35A. We have previously shown that the UL35 gene is involved in HCMV assembly, but it is unknown how UL35 proteins regulate viral assembly. Here we show that sorting nexin 5 (SNX5), a component of the retromer and part of the retrograde transport pathway, interacts with UL35 proteins. Expression of wild-type proteins but not mutants defective in SNX5 binding resulted in the cellular redistribution of the cation-independent mannose-6-phosphate receptor (CI-M6PR), indicating that UL35 proteins bind and negatively regulate SNX5 to modulate cellular transport pathways. Furthermore, binding of UL35 proteins to SNX5 was required for efficient viral replication and for transport of the most abundant HCMV glycoprotein B (gB; gpUL55) to the cVAC. These results indicate that ppUL35 and ppUL35A control the localization of the essential gB through the regulation of a retrograde transport pathway. Thus, this work is the first to define a molecular interaction between a tegument protein and a vesicular transport factor to regulate glycoprotein localization. IMPORTANCE Human cytomegalovirus is ubiquitously present in the healthy population, but reactivation or reinfection can cause serious, life-threatening infections in immunocompromised patients. For completion of its lytic cycle, human cytomegalovirus induces formation of an assembly center where mature virus particles are formed from multiple viral proteins. Viral glycoproteins use separate vesicular pathways for transport to the assembly center, which are incompletely understood. Our research identified a viral structural protein which affects the localization of one of the major glycoproteins. We could link this change in glycoprotein localization to an interaction of the structural protein with a cellular protein involved in regulation of vesicle transport. This increases our understanding of how the virus intersects into cellular regulatory pathways to enhance its own replication. Copyright © 2018 American Society for Microbiology.

Interactions of Notch1 and TLR4 signaling pathways in DRG neurons of in vivo and in vitro models of diabetic neuropathy.

PubMed

Chen, Tianhua; Li, Hao; Yin, Yiting; Zhang, Yuanpin; Liu, Zhen; Liu, Huaxiang

2017-11-02

Understanding the interactions between Notch1 and toll-like receptor 4 (TLR4) signaling pathways in the development of diabetic peripheral neuropathy may lead to interpretation of the mechanisms and novel approaches for preventing diabetic neuropathic pain. In the present study, the interactions between Notch1 and TLR4 signaling pathways were investigated by using dorsal root ganglion (DRG) from diabetic neuropathic pain rats and cultured DRG neurons under high glucose challenge. The results showed that high glucose induced not only Notch1 mRNA, HES1 mRNA, and TLR4 mRNA expression, but also Notch1 intracellular domain (NICD1) and TLR4 protein expression in DRG neurons. The proportion of NICD1-immunoreactive (IR) and TLR4-IR neurons in DRG cultures was also increased after high glucose challenge. The above alterations could be partially reversed by inhibition of either Notch1 or TLR4 signaling pathway. Inhibition of either Notch1 or TLR4 signaling pathway could improve mechanical allodynia and thermal hyperalgesia thresholds. Inhibition of Notch1 or TLR4 signaling also decreased tumor necrosis factor-α (TNF-α) levels in DRG from diabetic neuropathic rats. These data imply that the interaction between Notch1 and TLR4 signaling pathways is one of the important mechanisms in the development or progression of diabetic neuropathy.

Autocrine IL-10 activation of the STAT3 pathway is required for pathological macrophage differentiation in polycystic kidney disease

PubMed Central

Peda, Jacqueline D.; Salah, Sally M.; Wallace, Darren P.; Fields, Patrick E.; Grantham, Connor J.; Fields, Timothy A.

2016-01-01

ABSTRACT Polycystic kidney disease (PKD) is characterized by slow expansion of fluid-filled cysts derived from tubules within the kidney. Cystic expansion results in injury to surrounding parenchyma and leads to inflammation, scarring and ultimately loss of renal function. Macrophages are a key element in this process, promoting cyst epithelial cell proliferation, cyst expansion and disease progression. Previously, we have shown that the microenvironment established by cystic epithelial cells can ‘program’ macrophages, inducing M2-like macrophage polarization that is characterized by expression of markers that include Arg1 and Il10. Here, we functionally characterize these macrophages, demonstrating that their differentiation enhances their ability to promote cyst cell proliferation. This observation indicates a model of reciprocal pathological interactions between cysts and the innate immune system: cyst epithelial cells promote macrophage polarization to a phenotype that, in turn, is especially efficient in promoting cyst cell proliferation and cyst growth. To better understand the genesis of this macrophage phenotype, we examined the role of IL-10, a regulatory cytokine shown to be important for macrophage-stimulated tissue repair in other settings. Herein, we show that the acquisition of the pathological macrophage phenotype requires IL-10 secretion by the macrophages. Further, we demonstrate a requirement for IL-10-dependent autocrine activation of the STAT3 pathway. These data suggest that the IL-10 pathway in macrophages plays an essential role in the pathological relationship between cysts and the innate immune system in PKD, and thus could be a potential therapeutic target. PMID:27491076

The Argonaute CSR-1 and its 22G-RNA cofactors are required for holocentric chromosome segregation.

PubMed

Claycomb, Julie M; Batista, Pedro J; Pang, Ka Ming; Gu, Weifeng; Vasale, Jessica J; van Wolfswinkel, Josien C; Chaves, Daniel A; Shirayama, Masaki; Mitani, Shohei; Ketting, René F; Conte, Darryl; Mello, Craig C

2009-10-02

RNAi-related pathways regulate diverse processes, from developmental timing to transposon silencing. Here, we show that in C. elegans the Argonaute CSR-1, the RNA-dependent RNA polymerase EGO-1, the Dicer-related helicase DRH-3, and the Tudor-domain protein EKL-1 localize to chromosomes and are required for proper chromosome segregation. In the absence of these factors chromosomes fail to align at the metaphase plate and kinetochores do not orient to opposing spindle poles. Surprisingly, the CSR-1-interacting small RNAs (22G-RNAs) are antisense to thousands of germline-expressed protein-coding genes. Nematodes assemble holocentric chromosomes in which continuous kinetochores must span the expressed domains of the genome. We show that CSR-1 interacts with chromatin at target loci but does not downregulate target mRNA or protein levels. Instead, our findings support a model in which CSR-1 complexes target protein-coding domains to promote their proper organization within the holocentric chromosomes of C. elegans.

New insights into siRNA amplification and RNAi.

PubMed

Zhang, Chi; Ruvkun, Gary

2012-08-01

In the nematode Caenorhabditis elegans (C. elegans), gene inactivation by RNA interference can achieve remarkable potency due to the amplification of initial silencing triggers by RNA-dependent RNA polymerases (RdRPs). RdRPs catalyze the biogenesis of an abundant species of secondary small interfering RNAs (siRNAs) using the target mRNA as template. The interaction between primary siRNAs derived from the exogenous double-stranded RNA (dsRNA) trigger and the target mRNA is required for the recruitment of RdRPs. Other genetic requirements for RdRP activities have not been characterized. Recent studies have identified the RDE-10/RDE-11 complex which interacts with the primary siRNA bound target mRNA and acts upstream of the RdRPs. rde-10 and rde-11 mutants show an RNAi defective phenotype because the biogenesis of secondary siRNAs is completely abolished. In addition, the RDE-10/RDE-11 complex plays a similar role in the endogenous RNAi pathway for the biogenesis of a subset of siRNAs targeting recently acquired, duplicated genes.

Nontranscriptional regulation of SYK by the coactivator OCA-B is required at multiple stages of B cell development.

PubMed

Siegel, Rachael; Kim, Unkyu; Patke, Alina; Yu, Xin; Ren, Xiaodi; Tarakhovsky, Alexander; Roeder, Robert G

2006-05-19

OCA-B was originally identified as a nuclear transcriptional coactivator that is essential for antigen-driven immune responses. The later identification of a membrane bound, myristoylated form of OCA-B suggested additional, unique functions in B cell signaling pathways. This study has shown that OCA-B also functions in the pre-B1-to-pre-B2 cell transition and, most surprisingly, that it directly interacts with SYK, a tyrosine kinase critical for pre-BCR and BCR signaling. This unprecedented type of interaction-a transcriptional coactivator with a signaling kinase-occurs in the cytoplasm and directly regulates SYK stability. This study indicates that OCA-B is required for pre-BCR and BCR signaling at multiple stages of B cell development through its nontranscriptional regulation of SYK. Combined with the deregulation of OCA-B target genes, this may help explain the multitude of defects observed in B cell development and immune responses of Oca-b-/- mice.

Modelling the structure of a ceRNA-theoretical, bipartite microRNA-mRNA interaction network regulating intestinal epithelial cellular pathways using R programming.

PubMed

Robinson, J M; Henderson, W A

2018-01-12

We report a method using functional-molecular databases and network modelling to identify hypothetical mRNA-miRNA interaction networks regulating intestinal epithelial barrier function. The model forms a data-analysis component of our cell culture experiments, which produce RNA expression data from Nanostring Technologies nCounter ® system. The epithelial tight-junction (TJ) and actin cytoskeleton interact as molecular components of the intestinal epithelial barrier. Upstream regulation of TJ-cytoskeleton interaction is effected by the Rac/Rock/Rho signaling pathway and other associated pathways which may be activated or suppressed by extracellular signaling from growth factors, hormones, and immune receptors. Pathway activations affect epithelial homeostasis, contributing to degradation of the epithelial barrier associated with osmotic dysregulation, inflammation, and tumor development. The complexity underlying miRNA-mRNA interaction networks represents a roadblock for prediction and validation of competing-endogenous RNA network function. We developed a network model to identify hypothetical co-regulatory motifs in a miRNA-mRNA interaction network related to epithelial function. A mRNA-miRNA interaction list was generated using KEGG and miRWalk2.0 databases. R-code was developed to quantify and visualize inherent network structures. We identified a sub-network with a high number of shared, targeting miRNAs, of genes associated with cellular proliferation and cancer, including c-MYC and Cyclin D.

Ligand Receptor-Mediated Regulation of Growth in Plants.

PubMed

Haruta, Miyoshi; Sussman, Michael R

2017-01-01

Growth and development of multicellular organisms are coordinately regulated by various signaling pathways involving the communication of inter- and intracellular components. To form the appropriate body patterns, cellular growth and development are modulated by either stimulating or inhibiting these pathways. Hormones and second messengers help to mediate the initiation and/or interaction of the various signaling pathways in all complex multicellular eukaryotes. In plants, hormones include small organic molecules, as well as larger peptides and small proteins, which, as in animals, act as ligands and interact with receptor proteins to trigger rapid biochemical changes and induce the intracellular transcriptional and long-term physiological responses. During the past two decades, the availability of genetic and genomic resources in the model plant species, Arabidopsis thaliana, has greatly helped in the discovery of plant hormone receptors and the components of signal transduction pathways and mechanisms used by these immobile but highly complex organisms. Recently, it has been shown that two of the most important plant hormones, auxin and abscisic acid (ABA), act through signaling pathways that have not yet been recognized in animals. For example, auxins stimulate cell elongation by bringing negatively acting transcriptional repressor proteins to the proteasome to be degraded, thus unleashing the gene expression program required for increasing cell size. The "dormancy" inducing hormone, ABA, binds to soluble receptor proteins and inhibits a specific class of protein phosphatases (PP2C), which activates phosphorylation signaling leading to transcriptional changes needed for the desiccation of the seeds prior to entering dormancy. While these two hormone receptors have no known animal counterparts, there are also many similarities between animal and plant signaling pathways. For example, in plants, the largest single gene family in the genome is the protein kinase family (approximately 5% of the protein coding genes), although the specific function for only a few dozen of these kinases is clearly established. Recent comparative genomics studies have revealed that parasitic nematodes and pathogenic microbes produce plant peptide hormone mimics that target specific plant plasma membrane receptor-like protein kinases, thus usurping endogenous signaling pathways for their own pathogenic purposes. With biochemical, genetic, and physiological analyses of the regulation of hormone receptor signal pathways, we are thus just now beginning to understand how plants optimize the development of their body shape and cope with constantly changing environmental conditions. © 2017 Elsevier Inc. All rights reserved.

Global regulation by the seven-component Pi signaling system.

PubMed

Hsieh, Yi-Ju; Wanner, Barry L

2010-04-01

This review concerns how Escherichia coli detects environmental inorganic orthophosphate (P(i)) to regulate genes of the phosphate (Pho) regulon by the PhoR/PhoB two-component system (TCS). P(i) control by the PhoR/PhoB TCS is a paradigm of a bacterial signal transduction pathway in which occupancy of a cell surface receptor(s) controls gene expression in the cytoplasm. The P(i) signaling pathway requires seven proteins, all of which probably interact in a membrane-associated signaling complex. Our latest studies show that P(i) signaling involves three distinct processes, which appear to correspond to different states of the sensory histidine kinase PhoR: an inhibition state, an activation state, and a deactivation state. We describe a revised model for P(i) signal transduction of the E. coli Pho regulon. Copyright 2010 Elsevier Ltd. All rights reserved.

Exploring the formation pathways of donor-acceptor catenanes in aqueous dynamic combinatorial libraries.

PubMed

Cougnon, Fabien B L; Au-Yeung, Ho Yu; Pantoş, G Dan; Sanders, Jeremy K M

2011-03-09

The discovery through dynamic combinatorial chemistry (DCC) of a new generation of donor-acceptor [2]catenanes highlights the power of DCC to access unprecedented structures. While conventional thinking has limited the scope of donor-acceptor catenanes to strictly alternating stacks of donor (D) and acceptor (A) aromatic units, DCC is demonstrated in this paper to give access to unusual DAAD, DADD, and ADAA stacks. Each of these catenanes has specific structural requirements, allowing control of their formation. On the basis of these results, and on the observation that the catenanes represent kinetic bottlenecks in the reaction pathway, we propose a mechanism that explains and predicts the structures formed. Furthermore, the spontaneous assembly of catenanes in aqueous dynamic systems gives a fundamental insight into the role played by hydrophobic effect and donor-acceptor interactions when building such complex architectures.

Protein prenylation: a new mode of host-pathogen interaction.

PubMed

Amaya, Moushimi; Baranova, Ancha; van Hoek, Monique L

2011-12-09

Post translational modifications are required for proteins to be fully functional. The three step process, prenylation, leads to farnesylation or geranylgeranylation, which increase the hydrophobicity of the prenylated protein for efficient anchoring into plasma membranes and/or organellar membranes. Prenylated proteins function in a number of signaling and regulatory pathways that are responsible for basic cell operations. Well characterized prenylated proteins include Ras, Rac and Rho. Recently, pathogenic prokaryotic proteins, such as SifA and AnkB, have been shown to be prenylated by eukaryotic host cell machinery, but their functions remain elusive. The identification of other bacterial proteins undergoing this type of host-directed post-translational modification shows promise in elucidating host-pathogen interactions to develop new therapeutics. This review incorporates new advances in the study of protein prenylation into a broader aspect of biology with a focus on host-pathogen interaction. Copyright © 2011 Elsevier Inc. All rights reserved.

Viral Disease Networks?

NASA Astrophysics Data System (ADS)

Gulbahce, Natali; Yan, Han; Vidal, Marc; Barabasi, Albert-Laszlo

2010-03-01

Viral infections induce multiple perturbations that spread along the links of the biological networks of the host cells. Understanding the impact of these cascading perturbations requires an exhaustive knowledge of the cellular machinery as well as a systems biology approach that reveals how individual components of the cellular system function together. Here we describe an integrative method that provides a new approach to studying virus-human interactions and its correlations with diseases. Our method involves the combined utilization of protein - protein interactions, protein -- DNA interactions, metabolomics and gene - disease associations to build a ``viraldiseasome''. By solely using high-throughput data, we map well-known viral associated diseases and predict new candidate viral diseases. We use microarray data of virus-infected tissues and patient medical history data to further test the implications of the viral diseasome. We apply this method to Epstein-Barr virus and Human Papillomavirus and shed light into molecular development of viral diseases and disease pathways.

Cell Adhesions: Actin-Based Modules that Mediate Cell-Extracellular Matrix and Cell-Cell Interactions

PubMed Central

Bachir, Alexia; Horwitz, Alan Rick; Nelson, W. James; Bianchini, Julie M.

2018-01-01

Cell adhesions link cells to the extracellular matrix (ECM) and to each other, and depend on interactions with the actin cytoskeleton. Both cell-ECM and cell-cell adhesion sites contain discrete, yet overlapping functional modules. These modules establish physical association with the actin cytoskeleton, locally modulate actin organization and dynamics, and trigger intracellular signaling pathways. Interplay between these modules generates distinct actin architectures that underlie different stages, types, and functions of cell-ECM and cell-cell adhesions. Actomyosin contractility is required to generate mature, stable adhesions, as well as sense and translate the mechanical properties of the cellular environment to changes in cell organization and behavior. In this chapter we discuss the organization and function of different adhesion modules and how they interact with the actin cytoskeleton. We highlight the molecular mechanisms of mechanotransduction in adhesions, and how adhesion molecules mediate crosstalk between cell-ECM and cell-cell adhesion sites. PMID:28679638

Predictive motor control of sensory dynamics in Auditory Active Sensing

PubMed Central

Morillon, Benjamin; Hackett, Troy A.; Kajikawa, Yoshinao; Schroeder, Charles E.

2016-01-01

Neuronal oscillations present potential physiological substrates for brain operations that require temporal prediction. We review this idea in the context of auditory perception. Using speech as an exemplar, we illustrate how hierarchically organized oscillations can be used to parse and encode complex input streams. We then consider the motor system as a major source of rhythms (temporal priors) in auditory processing, that act in concert with attention to sharpen sensory representations and link them across areas. We discuss the anatomo-functional pathways that could mediate this audio-motor interaction, and notably the potential role of the somatosensory cortex. Finally, we reposition temporal predictions in the context of internal models, discussing how they interact with feature-based or spatial predictions. We argue that complementary predictions interact synergistically according to the organizational principles of each sensory system, forming multidimensional filters crucial to perception. PMID:25594376

Modulators of 14-3-3 Protein–Protein Interactions

PubMed Central

2017-01-01

Direct interactions between proteins are essential for the regulation of their functions in biological pathways. Targeting the complex network of protein–protein interactions (PPIs) has now been widely recognized as an attractive means to therapeutically intervene in disease states. Even though this is a challenging endeavor and PPIs have long been regarded as “undruggable” targets, the last two decades have seen an increasing number of successful examples of PPI modulators, resulting in growing interest in this field. PPI modulation requires novel approaches and the integrated efforts of multiple disciplines to be a fruitful strategy. This perspective focuses on the hub-protein 14-3-3, which has several hundred identified protein interaction partners, and is therefore involved in a wide range of cellular processes and diseases. Here, we aim to provide an integrated overview of the approaches explored for the modulation of 14-3-3 PPIs and review the examples resulting from these efforts in both inhibiting and stabilizing specific 14-3-3 protein complexes by small molecules, peptide mimetics, and natural products. PMID:28968506

Rewiring MAP kinases in Saccharomyces cerevisiae to regulate novel targets through ubiquitination.

PubMed

Groves, Benjamin; Khakhar, Arjun; Nadel, Cory M; Gardner, Richard G; Seelig, Georg

2016-08-15

Evolution has often copied and repurposed the mitogen-activated protein kinase (MAPK) signaling module. Understanding how connections form during evolution, in disease and across individuals requires knowledge of the basic tenets that govern kinase-substrate interactions. We identify criteria sufficient for establishing regulatory links between a MAPK and a non-native substrate. The yeast MAPK Fus3 and human MAPK ERK2 can be functionally redirected if only two conditions are met: the kinase and substrate contain matching interaction domains and the substrate includes a phospho-motif that can be phosphorylated by the kinase and recruit a downstream effector. We used a panel of interaction domains and phosphorylation-activated degradation motifs to demonstrate modular and scalable retargeting. We applied our approach to reshape the signaling behavior of an existing kinase pathway. Together, our results demonstrate that a MAPK can be largely defined by its interaction domains and compatible phospho-motifs and provide insight into how MAPK-substrate connections form.

His86 from the N-terminus of frataxin coordinates iron and is required for Fe-S cluster synthesis.

PubMed

Gentry, Leslie E; Thacker, Matthew A; Doughty, Reece; Timkovich, Russell; Busenlehner, Laura S

2013-09-03

Human frataxin has a vital role in the biosynthesis of iron-sulfur (Fe-S) clusters in mitochondria, and its deficiency causes the neurodegenerative disease Friedreich's ataxia. Proposed functions for frataxin in the Fe-S pathway include iron donation to the Fe-S cluster machinery and regulation of cysteine desulfurase activity to control the rate of Fe-S production, although further molecular detail is required to distinguish these two possibilities. It is well established that frataxin can coordinate iron using glutamate and aspartate side chains on the protein surface; however, in this work we identify a new iron coordinating residue in the N-terminus of human frataxin using complementary spectroscopic and structural approaches. Further, we demonstrate that His86 in this N-terminal region is required for high affinity iron coordination and iron assembly of Fe-S clusters by ISCU as part of the Fe-S cluster biosynthetic complex. If a binding site that includes His86 is important for Fe-S cluster synthesis as part of its chaperone function, this raises the possibility that either iron binding at the acidic surface of frataxin may be spurious or that it is required for protein-protein interactions with the Fe-S biosynthetic quaternary complex. Our data suggest that iron coordination to frataxin may be significant to the Fe-S cluster biosynthesis pathway in mitochondria.

Identification of interleukin-1 beta as a key mediator in the upregulation of Cav3.2–USP5 interactions in the pain pathway

PubMed Central

Stemkowski, Patrick L; Garcia-Caballero, Agustin; Gadotti, Vinicius M; M’Dahoma, Said; Chen, Lina; Souza, Ivana A

2017-01-01

We recently reported that nerve injury or peripheral inflammation triggers an upregulation of the deubiquitinase, USP5 in mouse dorsal root ganglion and spinal dorsal horn. This leads to dysregulated ubiquitination of Cav3.2 T-type calcium channels, thus increasing Cav3.2 channel plasma membrane expression and nociceptive signaling in the primary afferent pain pathway. This phenomenon could be recapitulated by noninvasive, optogenetic activation of transient receptor potential vanilloid-1–expressing nociceptors, indicating that neuronal activity is a key player in this process. Given the relevance of the pro-inflammatory cytokine interleukin-1 beta in many forms of pathological pain, we hypothesized that interleukin-1 beta may be a critical cofactor required to drive upregulation of interactions between USP5 and Cav3.2 channels. Here, we report that gene expression, as well as protein levels for interleukin-1 beta and the endogenous interleukin-1 receptor-I antagonist, IL-1Ra are unaltered following conditioning stimulation of optogenetically targeted cutaneous nociceptors, indicating that neuronal activity is not a driver of interleukin-1 beta signaling. In contrast, co-immunoprecipitation experiments revealed that intrathecal administration of interleukin-1 beta in wild-type mice led to an increase in the interaction between USP5 and Cav3.2 in the spinal dorsal horn. Moreover, disruption of the interaction between USP5 and Cav3.2 with TAT peptides suppressed acute nocifensive responses produced by interleukin-1 beta, which was similar to that achieved by elimination of T-type channel activity with the channel blockers, mibefradil, or TTA-A2. Finally, this upregulation could be maintained in dorsal root ganglion neuron cultures exposed overnight to interleukin-1 beta, while the copresence of interleukin-1 receptor antagonist or the dampening of neuronal cell activity with tetrodotoxin attenuated this response. Altogether, our findings identify interleukin-1 beta as an upstream trigger for the upregulation of interactions between USP5 and Cav3.2 channels in the pain pathway, presumably by triggering increased firing activity in afferent fibers. PMID:28741432

Human DHEA sulfation requires direct interaction between PAPS synthase 2 and DHEA sulfotransferase SULT2A1.

PubMed

Mueller, Jonathan W; Idkowiak, Jan; Gesteira, Tarsis F; Vallet, Cecilia; Hardman, Rebecca; van den Boom, Johannes; Dhir, Vivek; Knauer, Shirley K; Rosta, Edina; Arlt, Wiebke

2018-06-22

The high-energy sulfate donor 3'-phosphoadenosine-5'-phosphosulfate (PAPS), generated by human PAPS synthase isoforms PAPSS1 and PAPSS2, is required for all human sulfation pathways. Sulfotransferase SULT2A1 uses PAPS for sulfation of the androgen precursor dehydroepiandrosterone (DHEA), thereby reducing downstream activation of DHEA to active androgens. Human PAPSS2 mutations manifest with undetectable DHEA sulfate, androgen excess, and metabolic disease, suggesting that ubiquitous PAPSS1 cannot compensate for deficient PAPSS2 in supporting DHEA sulfation. In knockdown studies in human adrenocortical NCI-H295R1 cells, we found that PAPSS2, but not PAPSS1, is required for efficient DHEA sulfation. Specific APS kinase activity, the rate-limiting step in PAPS biosynthesis, did not differ between PAPSS1 and PAPSS2. Co-expression of cytoplasmic SULT2A1 with a cytoplasmic PAPSS2 variant supported DHEA sulfation more efficiently than co-expression with nuclear PAPSS2 or nuclear/cytosolic PAPSS1. Proximity ligation assays revealed protein-protein interactions between SULT2A1 and PAPSS2 and, to a lesser extent, PAPSS1. Molecular docking studies showed a putative binding site for SULT2A1 within the PAPSS2 APS kinase domain. Energy-dependent scoring of docking solutions identified the interaction as specific for the PAPSS2 and SULT2A1 isoforms. These findings elucidate the mechanistic basis for the selective requirement for PAPSS2 in human DHEA sulfation. © 2018 Mueller et al.

A Ca(2)+ signaling pathway regulates a K(+) channel for low-K response in Arabidopsis.

PubMed

Li, Legong; Kim, Beom-Gi; Cheong, Yong Hwa; Pandey, Girdhar K; Luan, Sheng

2006-08-15

Nutrient sensing is critical for plant adaptation to the environment. Because of extensive farming and erosion, low content of mineral nutrients such as potassium (K(+)) in soils becomes a limiting factor for plant growth. In response to low-K conditions, plants enhance their capability of K(+) uptake through an unknown signaling mechanism. Here we report the identification of a Ca(2+)-dependent pathway for low-K response in Arabidopsis. We are not aware of any other example of a molecular pathway for a nutrient response in plants. Earlier genetic analyses revealed three genes encoding two Ca(2+) sensors (CBL1 and CBL9) and their target protein kinase (CIPK23) to be critical for plant growth on low-K media and for stomatal regulation, indicating that these calcium signaling components participate in the low-K response and turgor regulation. In this study, we show that the protein kinase CIPK23 interacted with, and phosphorylated, a voltage-gated inward K(+) channel (AKT1) required for K(+) acquisition in Arabidopsis. In the Xenopus oocyte system, our studies showed that interacting calcium sensors (CBL1 and CBL9) together with target kinase CIPK23, but not either component alone, activated the AKT1 channel in a Ca(2+)-dependent manner, connecting the Ca(2+) signal to enhanced K(+) uptake through activation of a K(+) channel. Disruption of both CBL1 and CBL9 or CIPK23 gene in Arabidopsis reduced the AKT1 activity in the mutant roots, confirming that the Ca(2+)-CBL-CIPK pathway functions to orchestrate transporting activities in planta according to external K(+) availability.

Transcriptomic Insights into Phenological Development and Cold Tolerance of Wheat Grown in the Field1[OPEN

PubMed Central

Li, Qiang; Byrns, Brook; Badawi, Mohamed A.; Diallo, Abdoulaye Banire; Danyluk, Jean; Sarhan, Fathey; Zou, Jitao

2018-01-01

Cold acclimation and winter survival in cereal species is determined by complicated environmentally regulated gene expression. However, studies investigating these complex cold responses are mostly conducted in controlled environments that only consider the responses to single environmental variables. In this study, we have comprehensively profiled global transcriptional responses in crowns of field-grown spring and winter wheat (Triticum aestivum) genotypes and their near-isogenic lines with the VRN-A1 alleles swapped. This in-depth analysis revealed multiple signaling, interactive pathways that influence cold tolerance and phenological development to optimize plant growth and development in preparation for a wide range of over-winter stresses. Investigation of genetic differences at the VRN-A1 locus revealed that a vernalization requirement maintained a higher level of cold response pathways while VRN-A1 genetically promoted floral development. Our results also demonstrated the influence of genetic background on the expression of cold and flowering pathways. The link between delayed shoot apex development and the induction of cold tolerance was reflected by the gradual up-regulation of abscisic acid-dependent and C-REPEAT-BINDING FACTOR pathways. This was accompanied by the down-regulation of key genes involved in meristem development as the autumn progressed. The chromosome location of differentially expressed genes between the winter and spring wheat genetic backgrounds showed a striking pattern of biased gene expression on chromosomes 6A and 6D, indicating a transcriptional regulation at the genome level. This finding adds to the complexity of the genetic cascades and gene interactions that determine the evolutionary patterns of both phenological development and cold tolerance traits in wheat. PMID:29259104

Regulators of nonsulfur purple phototrophic bacteria and the interactive control of CO2 assimilation, nitrogen fixation, hydrogen metabolism and energy generation.

PubMed

Dubbs, James M; Tabita, F Robert

2004-06-01

For the metabolically diverse nonsulfur purple phototrophic bacteria, maintaining redox homeostasis requires balancing the activities of energy supplying and energy-utilizing pathways, often in the face of drastic changes in environmental conditions. These organisms, members of the class Alphaproteobacteria, primarily use CO2 as an electron sink to achieve redox homeostasis. After noting the consequences of inactivating the capacity for CO2 reduction through the Calvin-Benson-Bassham (CBB) pathway, it was shown that the molecular control of many additional important biological processes catalyzed by nonsulfur purple bacteria is linked to expression of the CBB genes. Several regulator proteins are involved, with the two component Reg/Prr regulatory system playing a major role in maintaining redox poise in these organisms. Reg/Prr was shown to be a global regulator involved in the coordinate control of a number of metabolic processes including CO2 assimilation, nitrogen fixation, hydrogen metabolism and energy-generation pathways. Accumulating evidence suggests that the Reg/Prr system senses the oxidation/reduction state of the cell by monitoring a signal associated with electron transport. The response regulator RegA/PrrA activates or represses gene expression through direct interaction with target gene promoters where it often works in concert with other regulators that can be either global or specific. For the key CO2 reduction pathway, which clearly triggers whether other redox balancing mechanisms are employed, the ability to activate or inactivate the specific regulator CbbR is of paramount importance. From these studies, it is apparent that a detailed understanding of how diverse regulatory elements integrate and control metabolism will eventually be achieved.

Emory University: High-Throughput Protein-Protein Interaction Analysis for Hippo Pathway Profiling | Office of Cancer Genomics

Cancer.gov

The CTD2 Center at Emory University used high-throughput protein-protein interaction (PPI) mapping for Hippo signaling pathway profiling to rapidly unveil promising PPIs as potential therapeutic targets and advance functional understanding of signaling circuitry in cells. Read the abstract.

Role of Nitrogen and Carbon Transport, Regulation, and Metabolism Genes for Saccharomyces cerevisiae Survival In Vivo†

PubMed Central

Kingsbury, Joanne M.; Goldstein, Alan L.; McCusker, John H.

2006-01-01

Saccharomyces cerevisiae is both an emerging opportunistic pathogen and a close relative of pathogenic Candida species. To better understand the ecology of fungal infection, we investigated the importance of pathways involved in uptake, metabolism, and biosynthesis of nitrogen and carbon compounds for survival of a clinical S. cerevisiae strain in a murine host. Potential nitrogen sources in vivo include ammonium, urea, and amino acids, while potential carbon sources include glucose, lactate, pyruvate, and fatty acids. Using mutants unable to either transport or utilize these compounds, we demonstrated that no individual nitrogen source was essential, while glucose was the most significant primary carbon source for yeast survival in vivo. Hydrolysis of the storage carbohydrate glycogen made a slight contribution for in vivo survival compared with a substantial requirement for trehalose hydrolysis. The ability to sense and respond to low glucose concentrations was also important for survival. In contrast, there was little or no requirement in vivo in this assay for any of the nitrogen-sensing pathways, nitrogen catabolite repression, the ammonium- or amino acid-sensing pathways, or general control. By using auxotrophic mutants, we found that some nitrogenous compounds (polyamines, methionine, and lysine) can be acquired from the host, while others (threonine, aromatic amino acids, isoleucine, and valine) must be synthesized by the pathogen. Our studies provide insights into the yeast-host environment interaction and identify potential antifungal drug targets. PMID:16682459

An editor for pathway drawing and data visualization in the Biopathways Workbench.

PubMed

Byrnes, Robert W; Cotter, Dawn; Maer, Andreia; Li, Joshua; Nadeau, David; Subramaniam, Shankar

2009-10-02

Pathway models serve as the basis for much of systems biology. They are often built using programs designed for the purpose. Constructing new models generally requires simultaneous access to experimental data of diverse types, to databases of well-characterized biological compounds and molecular intermediates, and to reference model pathways. However, few if any software applications provide all such capabilities within a single user interface. The Pathway Editor is a program written in the Java programming language that allows de-novo pathway creation and downloading of LIPID MAPS (Lipid Metabolites and Pathways Strategy) and KEGG lipid metabolic pathways, and of measured time-dependent changes to lipid components of metabolism. Accessed through Java Web Start, the program downloads pathways from the LIPID MAPS Pathway database (Pathway) as well as from the LIPID MAPS web server http://www.lipidmaps.org. Data arises from metabolomic (lipidomic), microarray, and protein array experiments performed by the LIPID MAPS consortium of laboratories and is arranged by experiment. Facility is provided to create, connect, and annotate nodes and processes on a drawing panel with reference to database objects and time course data. Node and interaction layout as well as data display may be configured in pathway diagrams as desired. Users may extend diagrams, and may also read and write data and non-lipidomic KEGG pathways to and from files. Pathway diagrams in XML format, containing database identifiers referencing specific compounds and experiments, can be saved to a local file for subsequent use. The program is built upon a library of classes, referred to as the Biopathways Workbench, that convert between different file formats and database objects. An example of this feature is provided in the form of read/construct/write access to models in SBML (Systems Biology Markup Language) contained in the local file system. Inclusion of access to multiple experimental data types and of pathway diagrams within a single interface, automatic updating through connectivity to an online database, and a focus on annotation, including reference to standardized lipid nomenclature as well as common lipid names, supports the view that the Pathway Editor represents a significant, practicable contribution to current pathway modeling tools.

Exosites in the substrate specificity of blood coagulation reactions.

PubMed

Bock, P E; Panizzi, P; Verhamme, I M A

2007-07-01

The specificity of blood coagulation proteinases for substrate, inhibitor, and effector recognition is mediated by exosites on the surfaces of the catalytic domains, physically separated from the catalytic site. Some thrombin ligands bind specifically to either exosite I or II, while others engage both exosites. The involvement of different, overlapping constellations of exosite residues enables binding of structurally diverse ligands. The flexibility of the thrombin structure is central to the mechanism of complex formation and the specificity of exosite interactions. Encounter complex formation is driven by electrostatic ligand-exosite interactions, followed by conformational rearrangement to a stable complex. Exosites on some zymogens are in low affinity proexosite states and are expressed concomitant with catalytic site activation. The requirement for exosite expression controls the specificity of assembly of catalytic complexes on the coagulation pathway, such as the membrane-bound factor Xa*factor Va (prothrombinase) complex, and prevents premature assembly. Substrate recognition by prothrombinase involves a two-step mechanism with initial docking of prothrombin to exosites, followed by a conformational change to engage the FXa catalytic site. Prothrombin and its activation intermediates bind prothrombinase in two alternative conformations determined by the zymogen to proteinase transition that are hypothesized to involve prothrombin (pro)exosite I interactions with FVa, which underpin the sequential activation pathway. The role of exosites as the major source of substrate specificity has stimulated development of exosite-targeted anticoagulants for treatment of thrombosis.

Single-Molecule Microscopy and Force Spectroscopy of Membrane Proteins

NASA Astrophysics Data System (ADS)

Engel, Andreas; Janovjak, Harald; Fotiadis, Dimtrios; Kedrov, Alexej; Cisneros, David; Müller, Daniel J.

Single-molecule atomic force microscopy (AFM) provides novel ways to characterize the structure-function relationship of native membrane proteins. High-resolution AFM topographs allow observing the structure of single proteins at sub-nanometer resolution as well as their conformational changes, oligomeric state, molecular dynamics and assembly. We will review these feasibilities illustrating examples of membrane proteins in native and reconstituted membranes. Classification of individual topographs of single proteins allows understanding the principles of motions of their extrinsic domains, to learn about their local structural flexibilities and to find the entropy minima of certain conformations. Combined with the visualization of functionally related conformational changes these insights allow understanding why certain flexibilities are required for the protein to function and how structurally flexible regions allow certain conformational changes. Complementary to AFM imaging, single-molecule force spectroscopy (SMFS) experiments detect molecular interactions established within and between membrane proteins. The sensitivity of this method makes it possible to measure interactions that stabilize secondary structures such as transmembrane α-helices, polypeptide loops and segments within. Changes in temperature or protein-protein assembly do not change the locations of stable structural segments, but influence their stability established by collective molecular interactions. Such changes alter the probability of proteins to choose a certain unfolding pathway. Recent examples have elucidated unfolding and refolding pathways of membrane proteins as well as their energy landscapes.

Kidins220/ARMS as a functional mediator of multiple receptor signalling pathways.

PubMed

Neubrand, Veronika E; Cesca, Fabrizia; Benfenati, Fabio; Schiavo, Giampietro

2012-04-15

An increasing body of evidence suggests that several membrane receptors--in addition to activating distinct signalling cascades--also engage in substantial crosstalk with each other, thereby adjusting their signalling outcome as a function of specific input information. However, little is known about the molecular mechanisms that control their coordination and integration of downstream signalling. A protein that is likely to have a role in this process is kinase-D-interacting substrate of 220 kDa [Kidins220, also known as ankyrin repeat-rich membrane spanning (ARMS), hereafter referred to as Kidins220/ARMS]. Kidins220/ARMS is a conserved membrane protein that is preferentially expressed in the nervous system and interacts with the microtubule and actin cytoskeleton. It interacts with neurotrophin, ephrin, vascular endothelial growth factor (VEGF) and glutamate receptors, and is a common downstream target of several trophic stimuli. Kidins220/ARMS is required for neuronal differentiation and survival, and its expression levels modulate synaptic plasticity. Kidins220/ARMS knockout mice show developmental defects mainly in the nervous and cardiovascular systems, suggesting a crucial role for this protein in modulating the cross talk between different signalling pathways. In this Commentary, we summarise existing knowledge regarding the physiological functions of Kidins220/ARMS, and highlight some interesting directions for future studies on the role of this protein in health and disease.

A global interaction network maps a wiring diagram of cellular function

PubMed Central

Costanzo, Michael; VanderSluis, Benjamin; Koch, Elizabeth N.; Baryshnikova, Anastasia; Pons, Carles; Tan, Guihong; Wang, Wen; Usaj, Matej; Hanchard, Julia; Lee, Susan D.; Pelechano, Vicent; Styles, Erin B.; Billmann, Maximilian; van Leeuwen, Jolanda; van Dyk, Nydia; Lin, Zhen-Yuan; Kuzmin, Elena; Nelson, Justin; Piotrowski, Jeff S.; Srikumar, Tharan; Bahr, Sondra; Chen, Yiqun; Deshpande, Raamesh; Kurat, Christoph F.; Li, Sheena C.; Li, Zhijian; Usaj, Mojca Mattiazzi; Okada, Hiroki; Pascoe, Natasha; Luis, Bryan-Joseph San; Sharifpoor, Sara; Shuteriqi, Emira; Simpkins, Scott W.; Snider, Jamie; Suresh, Harsha Garadi; Tan, Yizhao; Zhu, Hongwei; Malod-Dognin, Noel; Janjic, Vuk; Przulj, Natasa; Troyanskaya, Olga G.; Stagljar, Igor; Xia, Tian; Ohya, Yoshikazu; Gingras, Anne-Claude; Raught, Brian; Boutros, Michael; Steinmetz, Lars M.; Moore, Claire L.; Rosebrock, Adam P.; Caudy, Amy A.; Myers, Chad L.; Andrews, Brenda; Boone, Charles

2017-01-01

We generated a global genetic interaction network for Saccharomyces cerevisiae, constructing over 23 million double mutants, identifying ~550,000 negative and ~350,000 positive genetic interactions. This comprehensive network maps genetic interactions for essential gene pairs, highlighting essential genes as densely connected hubs. Genetic interaction profiles enabled assembly of a hierarchical model of cell function, including modules corresponding to protein complexes and pathways, biological processes, and cellular compartments. Negative interactions connected functionally related genes, mapped core bioprocesses, and identified pleiotropic genes, whereas positive interactions often mapped general regulatory connections among gene pairs, rather than shared functionality. The global network illustrates how coherent sets of genetic interactions connect protein complex and pathway modules to map a functional wiring diagram of the cell. PMID:27708008

The Retinoblastoma pathway regulates stem cell proliferation in freshwater planarians.

PubMed

Zhu, Shu Jun; Pearson, Bret J

2013-01-15

Freshwater planarians are flatworms of the Lophotrochozoan superphylum and are well known for their regenerative abilities, which rely on a large population of pluripotent adult stem cells. However, the mechanisms by which planarians maintain a precise population of adult stem cells while balancing proliferation and cell death, remain to be elucidated. Here we have identified, characterized, and functionally tested the core Retinoblastoma (Rb) pathway components in planarian adult stem cell biology. The Rb pathway is an ancient and conserved mechanism of proliferation control from plants to animals and is composed of three core components: an Rb protein, and a transcription factor heterodimer of E2F and DP proteins. Although the planarian genome contains all components of the Rb pathway, we found that they have undergone gene loss from the ancestral state, similar to other species in their phylum. The single Rb homolog (Smed-Rb) was highly expressed in planarian stem cells and was required for stem cell maintenance, similar to the Rb-homologs p107 and p130 in vertebrates. We show that planarians and their phylum have undergone the most severe reduction in E2F genes observed thus far, and the single remaining E2F was predicted to be a repressive-type E2F (Smed-E2F4-1). Knockdown of either Smed-E2F4-1 or its dimerization partner Dp (Smed-Dp) by RNAi resulted in temporary hyper-proliferation. Finally, we showed that known Rb-interacting genes in other systems, histone deacetylase 1 and cyclinD (Smed-HDAC1; Smed-cycD), were similar to Rb in expression and phenotypes when knocked down by RNAi, suggesting that these established interactions with Rb may also be conserved in planarians. Together, these results showed that planarians use the conserved components of the Rb tumor suppressor pathway to control proliferation and cell survival. Copyright © 2012 Elsevier Inc. All rights reserved.

Srs2 overexpression reveals a helicase-independent role at replication forks that requires diverse cell functions

PubMed Central

León Ortiz, Ana María; Reid, Robert J. D.; Dittmar, John C.; Rothstein, Rodney; Nicolas, Alain

2011-01-01

Srs2 is a 3’ to 5’ DNA helicase that regulates many aspects of DNA metabolism in Saccharomyces cerevisiae. It is best known for its ability to counteract homologous recombination by dismantling Rad51 filaments, but is also involved in checkpoint activation, adaptation and recovery, and in resolution of late recombination intermediates. To further address its biological roles and uncover new genetic interactions, we examined the consequences of overexpressing SRS2 as well as two helicase-dead mutants, srs2-K41A and srs2-K41R, in the collection of 4827 yeast haploid deletion mutants. We identified 274 genes affecting a large variety of cellular functions that are required for cell growth when SRS2 or its mutants are overexpressed. Further analysis of these interactions reveals that Srs2 acts independently of its helicase function at replication forks likely through its recruitment by the sumoylated PCNA replication clamp. This helicase-independent function is responsible for the negative interactions with DNA metabolism genes and for the toxicity of SRS2 overexpression in many of the diverse cellular pathways revealed in our screens. PMID:21459050

A physiologically required G protein-coupled receptor (GPCR)-regulator of G protein signaling (RGS) interaction that compartmentalizes RGS activity.

PubMed

Croft, Wayne; Hill, Claire; McCann, Eilish; Bond, Michael; Esparza-Franco, Manuel; Bennett, Jeannette; Rand, David; Davey, John; Ladds, Graham

2013-09-20

G protein-coupled receptors (GPCRs) can interact with regulator of G protein signaling (RGS) proteins. However, the effects of such interactions on signal transduction and their physiological relevance have been largely undetermined. Ligand-bound GPCRs initiate by promoting exchange of GDP for GTP on the Gα subunit of heterotrimeric G proteins. Signaling is terminated by hydrolysis of GTP to GDP through intrinsic GTPase activity of the Gα subunit, a reaction catalyzed by RGS proteins. Using yeast as a tool to study GPCR signaling in isolation, we define an interaction between the cognate GPCR (Mam2) and RGS (Rgs1), mapping the interaction domains. This reaction tethers Rgs1 at the plasma membrane and is essential for physiological signaling response. In vivo quantitative data inform the development of a kinetic model of the GTPase cycle, which extends previous attempts by including GPCR-RGS interactions. In vivo and in silico data confirm that GPCR-RGS interactions can impose an additional layer of regulation through mediating RGS subcellular localization to compartmentalize RGS activity within a cell, thus highlighting their importance as potential targets to modulate GPCR signaling pathways.

Insights into significant pathways and gene interaction networks underlying breast cancer cell line MCF-7 treated with 17β-estradiol (E2).

PubMed

Huan, Jinliang; Wang, Lishan; Xing, Li; Qin, Xianju; Feng, Lingbin; Pan, Xiaofeng; Zhu, Ling

2014-01-01

Estrogens are known to regulate the proliferation of breast cancer cells and to alter their cytoarchitectural and phenotypic properties, but the gene networks and pathways by which estrogenic hormones regulate these events are only partially understood. We used global gene expression profiling by Affymetrix GeneChip microarray analysis, with KEGG pathway enrichment, PPI network construction, module analysis and text mining methods to identify patterns and time courses of genes that are either stimulated or inhibited by estradiol (E2) in estrogen receptor (ER)-positive MCF-7 human breast cancer cells. Of the genes queried on the Affymetrix Human Genome U133 plus 2.0 microarray, we identified 628 (12h), 852 (24h) and 880 (48 h) differentially expressed genes (DEGs) that showed a robust pattern of regulation by E2. From pathway enrichment analysis, we found out the changes of metabolic pathways of E2 treated samples at each time point. At 12h time point, the changes of metabolic pathways were mainly focused on pathways in cancer, focal adhesion, and chemokine signaling pathway. At 24h time point, the changes were mainly enriched in neuroactive ligand-receptor interaction, cytokine-cytokine receptor interaction and calcium signaling pathway. At 48 h time point, the significant pathways were pathways in cancer, regulation of actin cytoskeleton, cell adhesion molecules (CAMs), axon guidance and ErbB signaling pathway. Of interest, our PPI network analysis and module analysis found that E2 treatment induced enhancement of PRSS23 at the three time points and PRSS23 was in the central position of each module. Text mining results showed that the important genes of DEGs have relationship with signal pathways, such as ERbB pathway (AREG), Wnt pathway (NDP), MAPK pathway (NTRK3, TH), IP3 pathway (TRA@) and some transcript factors (TCF4, MAF). Our studies highlight the diverse gene networks and metabolic and cell regulatory pathways through which E2 operates to achieve its widespread effects on breast cancer cells. © 2013 Elsevier B.V. All rights reserved.

Innate immune humoral factors, C1q and factor H, with differential pattern recognition properties, alter macrophage response to carbon nanotubes.

PubMed

Pondman, Kirsten M; Pednekar, Lina; Paudyal, Basudev; Tsolaki, Anthony G; Kouser, Lubna; Khan, Haseeb A; Shamji, Mohamed H; Ten Haken, Bennie; Stenbeck, Gudrun; Sim, Robert B; Kishore, Uday

2015-11-01

Interaction between the complement system and carbon nanotubes (CNTs) can modify their intended biomedical applications. Pristine and derivatised CNTs can activate complement primarily via the classical pathway which enhances uptake of CNTs and suppresses pro-inflammatory response by immune cells. Here, we report that the interaction of C1q, the classical pathway recognition molecule, with CNTs involves charge pattern and classical pathway activation that is partly inhibited by factor H, a complement regulator. C1q and its globular modules, but not factor H, enhanced uptake of CNTs by macrophages and modulated the pro-inflammatory immune response. Thus, soluble complement factors can interact differentially with CNTs and alter the immune response even without complement activation. Coating CNTs with recombinant C1q globular heads offers a novel way of controlling classical pathway activation in nanotherapeutics. Surprisingly, the globular heads also enhance clearance by phagocytes and down-regulate inflammation, suggesting unexpected complexity in receptor interaction. Carbon nanotubes (CNTs) maybe useful in the clinical setting as targeting drug carriers. However, it is also well known that they can interact and activate the complement system, which may have a negative impact on the applicability of CNTs. In this study, the authors functionalized multi-walled CNT (MWNT), and investigated the interaction with the complement pathway. These studies are important so as to gain further understanding of the underlying mechanism in preparation for future use of CNTs in the clinical setting. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

A human functional protein interaction network and its application to cancer data analysis

PubMed Central

2010-01-01

Background One challenge facing biologists is to tease out useful information from massive data sets for further analysis. A pathway-based analysis may shed light by projecting candidate genes onto protein functional relationship networks. We are building such a pathway-based analysis system. Results We have constructed a protein functional interaction network by extending curated pathways with non-curated sources of information, including protein-protein interactions, gene coexpression, protein domain interaction, Gene Ontology (GO) annotations and text-mined protein interactions, which cover close to 50% of the human proteome. By applying this network to two glioblastoma multiforme (GBM) data sets and projecting cancer candidate genes onto the network, we found that the majority of GBM candidate genes form a cluster and are closer than expected by chance, and the majority of GBM samples have sequence-altered genes in two network modules, one mainly comprising genes whose products are localized in the cytoplasm and plasma membrane, and another comprising gene products in the nucleus. Both modules are highly enriched in known oncogenes, tumor suppressors and genes involved in signal transduction. Similar network patterns were also found in breast, colorectal and pancreatic cancers. Conclusions We have built a highly reliable functional interaction network upon expert-curated pathways and applied this network to the analysis of two genome-wide GBM and several other cancer data sets. The network patterns revealed from our results suggest common mechanisms in the cancer biology. Our system should provide a foundation for a network or pathway-based analysis platform for cancer and other diseases. PMID:20482850

A role for the mevalonate pathway in early plant symbiotic signaling

PubMed Central

Venkateshwaran, Muthusubramanian; Jayaraman, Dhileepkumar; Chabaud, Mireille; Genre, Andrea; Balloon, Allison J.; Maeda, Junko; Forshey, Kari; den Os, Désirée; Kwiecien, Nicholas W.; Coon, Joshua J.; Barker, David G.; Ané, Jean-Michel

2015-01-01

Rhizobia and arbuscular mycorrhizal fungi produce signals that are perceived by host legume receptors at the plasma membrane and trigger sustained oscillations of the nuclear and perinuclear Ca2+ concentration (Ca2+ spiking), which in turn leads to gene expression and downstream symbiotic responses. The activation of Ca2+ spiking requires the plasma membrane-localized receptor-like kinase Does not Make Infections 2 (DMI2) as well as the nuclear cation channel DMI1. A key enzyme regulating the mevalonate (MVA) pathway, 3-Hydroxy-3-Methylglutaryl CoA Reductase 1 (HMGR1), interacts with DMI2 and is required for the legume–rhizobium symbiosis. Here, we show that HMGR1 is required to initiate Ca2+ spiking and symbiotic gene expression in Medicago truncatula roots in response to rhizobial and arbuscular mycorrhizal fungal signals. Furthermore, MVA, the direct product of HMGR1 activity, is sufficient to induce nuclear-associated Ca2+ spiking and symbiotic gene expression in both wild-type plants and dmi2 mutants, but interestingly not in dmi1 mutants. Finally, MVA induced Ca2+ spiking in Human Embryonic Kidney 293 cells expressing DMI1. This demonstrates that the nuclear cation channel DMI1 is sufficient to support MVA-induced Ca2+ spiking in this heterologous system. PMID:26199419

A role for the mevalonate pathway in early plant symbiotic signaling.

PubMed

Venkateshwaran, Muthusubramanian; Jayaraman, Dhileepkumar; Chabaud, Mireille; Genre, Andrea; Balloon, Allison J; Maeda, Junko; Forshey, Kari; den Os, Désirée; Kwiecien, Nicholas W; Coon, Joshua J; Barker, David G; Ané, Jean-Michel

2015-08-04

Rhizobia and arbuscular mycorrhizal fungi produce signals that are perceived by host legume receptors at the plasma membrane and trigger sustained oscillations of the nuclear and perinuclear Ca(2+) concentration (Ca(2+) spiking), which in turn leads to gene expression and downstream symbiotic responses. The activation of Ca(2+) spiking requires the plasma membrane-localized receptor-like kinase Does not Make Infections 2 (DMI2) as well as the nuclear cation channel DMI1. A key enzyme regulating the mevalonate (MVA) pathway, 3-Hydroxy-3-Methylglutaryl CoA Reductase 1 (HMGR1), interacts with DMI2 and is required for the legume-rhizobium symbiosis. Here, we show that HMGR1 is required to initiate Ca(2+) spiking and symbiotic gene expression in Medicago truncatula roots in response to rhizobial and arbuscular mycorrhizal fungal signals. Furthermore, MVA, the direct product of HMGR1 activity, is sufficient to induce nuclear-associated Ca(2+) spiking and symbiotic gene expression in both wild-type plants and dmi2 mutants, but interestingly not in dmi1 mutants. Finally, MVA induced Ca(2+) spiking in Human Embryonic Kidney 293 cells expressing DMI1. This demonstrates that the nuclear cation channel DMI1 is sufficient to support MVA-induced Ca(2+) spiking in this heterologous system.

Role of RGM coreceptors in bone morphogenetic protein signaling

PubMed Central

Halbrooks, Peter J; Ding, Ru; Wozney, John M; Bain, Gerard

2007-01-01

Background The repulsive guidance molecule (RGM) proteins, originally discovered for their roles in neuronal development, have been recently identified as co-receptors in the bone morphogenetic protein (BMP) signaling pathway. BMPs are members of the TGFβ superfamily of signaling cytokines, and serve to regulate many aspects of cellular growth and differentiation. Results Here, we investigate whether RGMa, RGMb, and RGMc play required roles in BMP and TGFβ signaling in the mouse myoblast C2C12 cell line. These cells are responsive to BMPs and are frequently used to study BMP/TGFβ signaling pathways. Using siRNA reagents to specifically knock down each RGM protein, we show that the RGM co-receptors are required for significant BMP signaling as reported by two cell-based BMP activity assays: endogenous alkaline phosphatase activity and a luciferase-based BMP reporter assay. Similar cell-based assays using a TGFβ-induced luciferase reporter show that the RGM co-receptors are not required for TGFβ signaling. The binding interaction of each RGM co-receptor to each of BMP2 and BMP12 is observed and quantified, and equilibrium dissociation constants in the low nanomolar range are reported. Conclusion Our results demonstrate that the RGMs play a significant role in BMP signaling and reveal that these molecules cannot functionally compensate for one another. PMID:17615080

Yorkie regulates epidermal wound healing in Drosophila larvae independently of cell proliferation and apoptosis.

PubMed

Tsai, Chang-Ru; Anderson, Aimee E; Burra, Sirisha; Jo, Juyeon; Galko, Michael J

2017-07-01

Yorkie (Yki), the transcriptional co-activator of the Hippo signaling pathway, has well-characterized roles in balancing apoptosis and cell division during organ growth control. Yki is also required in diverse tissue regenerative contexts. In most cases this requirement reflects its well-characterized roles in balancing apoptosis and cell division. Whether Yki has repair functions outside of the control of cell proliferation, death, and growth is not clear. Here we show that Yki and Scalloped (Sd) are required for epidermal wound closure in the Drosophila larval epidermis. Using a GFP-tagged Yki transgene we show that Yki transiently translocates to some epidermal nuclei upon wounding. Genetic analysis strongly suggests that Yki interacts with the known wound healing pathway, Jun N-terminal kinase (JNK), but not with Platelet Derived Growth Factor/Vascular-Endothelial Growth Factor receptor (Pvr). Yki likely acts downstream of or parallel to JNK signaling and does not appear to regulate either proliferation or apoptosis in the larval epidermis during wound repair. Analysis of actin structures after wounding suggests that Yki and Sd promote wound closure through actin regulation. In sum, we found that Yki regulates an epithelial tissue repair process independently of its previously documented roles in balancing proliferation and apoptosis. Copyright © 2017 Elsevier Inc. All rights reserved.

Vectorial signalling mechanism required for cell-cell communication during sporulation in Bacillus subtilis.

PubMed

Diez, Veronica; Schujman, Gustavo E; Gueiros-Filho, Frederico J; de Mendoza, Diego

2012-01-01

Spore formation in Bacillus subtilis takes place in a sporangium consisting of two chambers, the forespore and the mother cell, which are linked by pathways of cell-cell communication. One pathway, which couples the proteolytic activation of the mother cell transcription factor σ(E) to the action of a forespore synthesized signal molecule, SpoIIR, has remained enigmatic. Signalling by SpoIIR requires the protein to be exported to the intermembrane space between forespore and mother cell, where it will interact with and activate the integral membrane protease SpoIIGA. Here we show that SpoIIR signal activity as well as the cleavage of its N-terminal extension is strictly dependent on the prespore fatty acid biosynthetic machinery. We also report that a conserved threonine residue (T27) in SpoIIR is required for processing, suggesting that signalling of SpoIIR is dependent on fatty acid synthesis probably because of acylation of T27. In addition, SpoIIR localization in the forespore septal membrane depends on the presence of SpoIIGA. The orchestration of σ(E) activation in the intercellular space by an acylated signal protein provides a new paradigm to ensure local transmission of a weak signal across the bilayer to control cell-cell communication during development. © 2011 Blackwell Publishing Ltd.

Genome scale transcriptomics of baculovirus-insect interactions.

PubMed

Nguyen, Quan; Nielsen, Lars K; Reid, Steven

2013-11-12

Baculovirus-insect cell technologies are applied in the production of complex proteins, veterinary and human vaccines, gene delivery vectors' and biopesticides. Better understanding of how baculoviruses and insect cells interact would facilitate baculovirus-based production. While complete genomic sequences are available for over 58 baculovirus species, little insect genomic information is known. The release of the Bombyx mori and Plutella xylostella genomes, the accumulation of EST sequences for several Lepidopteran species, and especially the availability of two genome-scale analysis tools, namely oligonucleotide microarrays and next generation sequencing (NGS), have facilitated expression studies to generate a rich picture of insect gene responses to baculovirus infections. This review presents current knowledge on the interaction dynamics of the baculovirus-insect system' which is relatively well studied in relation to nucleocapsid transportation, apoptosis, and heat shock responses, but is still poorly understood regarding responses involved in pro-survival pathways, DNA damage pathways, protein degradation, translation, signaling pathways, RNAi pathways, and importantly metabolic pathways for energy, nucleotide and amino acid production. We discuss how the two genome-scale transcriptomic tools can be applied for studying such pathways and suggest that proteomics and metabolomics can produce complementary findings to transcriptomic studies.

EphrinA2 Receptor (EphA2) Is an Invasion and Intracellular Signaling Receptor for Chlamydia trachomatis

PubMed Central

Subbarayal, Prema; Karunakaran, Karthika; Winkler, Ann-Cathrin; Rother, Marion; Gonzalez, Erik; Meyer, Thomas F.; Rudel, Thomas

2015-01-01

The obligate intracellular bacterium Chlamydia trachomatis invades into host cells to replicate inside a membrane-bound vacuole called inclusion. Multiple different host proteins are recruited to the inclusion and are functionally modulated to support chlamydial development. Invaded and replicating Chlamydia induces a long-lasting activation of the PI3 kinase signaling pathway that is required for efficient replication. We identified the cell surface tyrosine kinase EphrinA2 receptor (EphA2) as a chlamydial adherence and invasion receptor that induces PI3 kinase (PI3K) activation, promoting chlamydial replication. Interfering with binding of C. trachomatis serovar L2 (Ctr) to EphA2, downregulation of EphA2 expression or inhibition of EphA2 activity significantly reduced Ctr infection. Ctr interacts with and activates EphA2 on the cell surface resulting in Ctr and receptor internalization. During chlamydial replication, EphA2 remains active accumulating around the inclusion and interacts with the p85 regulatory subunit of PI3K to support the activation of the PI3K/Akt signaling pathway that is required for normal chlamydial development. Overexpression of full length EphA2, but not the mutant form lacking the intracellular cytoplasmic domain, enhanced PI3K activation and Ctr infection. Despite the depletion of EphA2 from the cell surface, Ctr infection induces upregulation of EphA2 through the activation of the ERK pathway, which keeps the infected cell in an apoptosis-resistant state. The significance of EphA2 as an entry and intracellular signaling receptor was also observed with the urogenital C. trachomatis-serovar D. Our findings provide the first evidence for a host cell surface receptor that is exploited for invasion as well as for receptor-mediated intracellular signaling to facilitate chlamydial replication. In addition, the engagement of a cell surface receptor at the inclusion membrane is a new mechanism by which Chlamydia subverts the host cell and induces apoptosis resistance. PMID:25906164

EphrinA2 receptor (EphA2) is an invasion and intracellular signaling receptor for Chlamydia trachomatis.

PubMed

Subbarayal, Prema; Karunakaran, Karthika; Winkler, Ann-Cathrin; Rother, Marion; Gonzalez, Erik; Meyer, Thomas F; Rudel, Thomas

2015-04-01

The obligate intracellular bacterium Chlamydia trachomatis invades into host cells to replicate inside a membrane-bound vacuole called inclusion. Multiple different host proteins are recruited to the inclusion and are functionally modulated to support chlamydial development. Invaded and replicating Chlamydia induces a long-lasting activation of the PI3 kinase signaling pathway that is required for efficient replication. We identified the cell surface tyrosine kinase EphrinA2 receptor (EphA2) as a chlamydial adherence and invasion receptor that induces PI3 kinase (PI3K) activation, promoting chlamydial replication. Interfering with binding of C. trachomatis serovar L2 (Ctr) to EphA2, downregulation of EphA2 expression or inhibition of EphA2 activity significantly reduced Ctr infection. Ctr interacts with and activates EphA2 on the cell surface resulting in Ctr and receptor internalization. During chlamydial replication, EphA2 remains active accumulating around the inclusion and interacts with the p85 regulatory subunit of PI3K to support the activation of the PI3K/Akt signaling pathway that is required for normal chlamydial development. Overexpression of full length EphA2, but not the mutant form lacking the intracellular cytoplasmic domain, enhanced PI3K activation and Ctr infection. Despite the depletion of EphA2 from the cell surface, Ctr infection induces upregulation of EphA2 through the activation of the ERK pathway, which keeps the infected cell in an apoptosis-resistant state. The significance of EphA2 as an entry and intracellular signaling receptor was also observed with the urogenital C. trachomatis-serovar D. Our findings provide the first evidence for a host cell surface receptor that is exploited for invasion as well as for receptor-mediated intracellular signaling to facilitate chlamydial replication. In addition, the engagement of a cell surface receptor at the inclusion membrane is a new mechanism by which Chlamydia subverts the host cell and induces apoptosis resistance.

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.

DNA hybridization kinetics: zippering, internal displacement and sequence dependence.

PubMed

Ouldridge, Thomas E; Sulc, Petr; Romano, Flavio; Doye, Jonathan P K; Louis, Ard A

2013-10-01

Although the thermodynamics of DNA hybridization is generally well established, the kinetics of this classic transition is less well understood. Providing such understanding has new urgency because DNA nanotechnology often depends critically on binding rates. Here, we explore DNA oligomer hybridization kinetics using a coarse-grained model. Strand association proceeds through a complex set of intermediate states, with successful binding events initiated by a few metastable base-pairing interactions, followed by zippering of the remaining bonds. But despite reasonably strong interstrand interactions, initial contacts frequently dissociate because typical configurations in which they form differ from typical states of similar enthalpy in the double-stranded equilibrium ensemble. Initial contacts must be stabilized by two or three base pairs before full zippering is likely, resulting in negative effective activation enthalpies. Non-Arrhenius behavior arises because the number of base pairs required for nucleation increases with temperature. In addition, we observe two alternative pathways-pseudoknot and inchworm internal displacement-through which misaligned duplexes can rearrange to form duplexes. These pathways accelerate hybridization. Our results explain why experimentally observed association rates of GC-rich oligomers are higher than rates of AT- rich equivalents, and more generally demonstrate how association rates can be modulated by sequence choice.

Sensitivity of projected long-term CO2 emissions across the Shared Socioeconomic Pathways

NASA Astrophysics Data System (ADS)

Marangoni, G.; Tavoni, M.; Bosetti, V.; Borgonovo, E.; Capros, P.; Fricko, O.; Gernaat, D. E. H. J.; Guivarch, C.; Havlik, P.; Huppmann, D.; Johnson, N.; Karkatsoulis, P.; Keppo, I.; Krey, V.; Ó Broin, E.; Price, J.; van Vuuren, D. P.

2017-01-01

Scenarios showing future greenhouse gas emissions are needed to estimate climate impacts and the mitigation efforts required for climate stabilization. Recently, the Shared Socioeconomic Pathways (SSPs) have been introduced to describe alternative social, economic and technical narratives, spanning a wide range of plausible futures in terms of challenges to mitigation and adaptation. Thus far the key drivers of the uncertainty in emissions projections have not been robustly disentangled. Here we assess the sensitivities of future CO2 emissions to key drivers characterizing the SSPs. We use six state-of-the-art integrated assessment models with different structural characteristics, and study the impact of five families of parameters, related to population, income, energy efficiency, fossil fuel availability, and low-carbon energy technology development. A recently developed sensitivity analysis algorithm allows us to parsimoniously compute both the direct and interaction effects of each of these drivers on cumulative emissions. The study reveals that the SSP assumptions about energy intensity and economic growth are the most important determinants of future CO2 emissions from energy combustion, both with and without a climate policy. Interaction terms between parameters are shown to be important determinants of the total sensitivities.

A reproducible approach to high-throughput biological data acquisition and integration

PubMed Central

Rahnavard, Gholamali; Waldron, Levi; McIver, Lauren; Shafquat, Afrah; Franzosa, Eric A.; Miropolsky, Larissa; Sweeney, Christopher

2015-01-01

Modern biological research requires rapid, complex, and reproducible integration of multiple experimental results generated both internally and externally (e.g., from public repositories). Although large systematic meta-analyses are among the most effective approaches both for clinical biomarker discovery and for computational inference of biomolecular mechanisms, identifying, acquiring, and integrating relevant experimental results from multiple sources for a given study can be time-consuming and error-prone. To enable efficient and reproducible integration of diverse experimental results, we developed a novel approach for standardized acquisition and analysis of high-throughput and heterogeneous biological data. This allowed, first, novel biomolecular network reconstruction in human prostate cancer, which correctly recovered and extended the NFκB signaling pathway. Next, we investigated host-microbiome interactions. In less than an hour of analysis time, the system retrieved data and integrated six germ-free murine intestinal gene expression datasets to identify the genes most influenced by the gut microbiota, which comprised a set of immune-response and carbohydrate metabolism processes. Finally, we constructed integrated functional interaction networks to compare connectivity of peptide secretion pathways in the model organisms Escherichia coli, Bacillus subtilis, and Pseudomonas aeruginosa. PMID:26157642

Constructing phylogenetic trees using interacting pathways.

PubMed

Wan, Peng; Che, Dongsheng

2013-01-01

Phylogenetic trees are used to represent evolutionary relationships among biological species or organisms. The construction of phylogenetic trees is based on the similarities or differences of their physical or genetic features. Traditional approaches of constructing phylogenetic trees mainly focus on physical features. The recent advancement of high-throughput technologies has led to accumulation of huge amounts of biological data, which in turn changed the way of biological studies in various aspects. In this paper, we report our approach of building phylogenetic trees using the information of interacting pathways. We have applied hierarchical clustering on two domains of organisms-eukaryotes and prokaryotes. Our preliminary results have shown the effectiveness of using the interacting pathways in revealing evolutionary relationships.

Inositol Pentakisphosphate Isomers Bind PH Domains with Varying Specificity and Inhibit Phosphoinositide Interactions

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

S Jackson; S Al-Saigh; C Schultz

2011-12-31

PH domains represent one of the most common domains in the human proteome. These domains are recognized as important mediators of protein-phosphoinositide and protein-protein interactions. Phosphoinositides are lipid components of the membrane that function as signaling molecules by targeting proteins to their sites of action. Phosphoinositide based signaling pathways govern a diverse range of important cellular processes including membrane remodeling, differentiation, proliferation and survival. Myo-Inositol phosphates are soluble signaling molecules that are structurally similar to the head groups of phosphoinositides. These molecules have been proposed to function, at least in part, by regulating PH domain-phosphoinositide interactions. Given the structural similaritymore » of inositol phosphates we were interested in examining the specificity of PH domains towards the family of myo-inositol pentakisphosphate isomers. In work reported here we demonstrate that the C-terminal PH domain of pleckstrin possesses the specificity required to discriminate between different myo-inositol pentakisphosphate isomers. The structural basis for this specificity was determined using high-resolution crystal structures. Moreover, we show that while the PH domain of Grp1 does not possess this high degree of specificity, the PH domain of protein kinase B does. These results demonstrate that some PH domains possess enough specificity to discriminate between myo-inositol pentakisphosphate isomers allowing for these molecules to differentially regulate interactions with phosphoinositides. Furthermore, this work contributes to the growing body of evidence supporting myo-inositol phosphates as regulators of important PH domain-phosphoinositide interactions. Finally, in addition to expanding our knowledge of cellular signaling, these results provide a basis for developing tools to probe biological pathway.« less

EspL is a bacterial cysteine protease effector that cleaves RHIM proteins to block necroptosis and inflammation.

PubMed

Pearson, Jaclyn S; Giogha, Cristina; Mühlen, Sabrina; Nachbur, Ueli; Pham, Chi L L; Zhang, Ying; Hildebrand, Joanne M; Oates, Clare V; Lung, Tania Wong Fok; Ingle, Danielle; Dagley, Laura F; Bankovacki, Aleksandra; Petrie, Emma J; Schroeder, Gunnar N; Crepin, Valerie F; Frankel, Gad; Masters, Seth L; Vince, James; Murphy, James M; Sunde, Margaret; Webb, Andrew I; Silke, John; Hartland, Elizabeth L

2017-01-13

Cell death signalling pathways contribute to tissue homeostasis and provide innate protection from infection. Adaptor proteins such as receptor-interacting serine/threonine-protein kinase 1 (RIPK1), receptor-interacting serine/threonine-protein kinase 3 (RIPK3), TIR-domain-containing adapter-inducing interferon-β (TRIF) and Z-DNA-binding protein 1 (ZBP1)/DNA-dependent activator of IFN-regulatory factors (DAI) that contain receptor-interacting protein (RIP) homotypic interaction motifs (RHIM) play a key role in cell death and inflammatory signalling 1-3 . RHIM-dependent interactions help drive a caspase-independent form of cell death termed necroptosis 4,5 . Here, we report that the bacterial pathogen enteropathogenic Escherichia coli (EPEC) uses the type III secretion system (T3SS) effector EspL to degrade the RHIM-containing proteins RIPK1, RIPK3, TRIF and ZBP1/DAI during infection. This requires a previously unrecognized tripartite cysteine protease motif in EspL (Cys47, His131, Asp153) that cleaves within the RHIM of these proteins. Bacterial infection and/or ectopic expression of EspL leads to rapid inactivation of RIPK1, RIPK3, TRIF and ZBP1/DAI and inhibition of tumour necrosis factor (TNF), lipopolysaccharide or polyinosinic:polycytidylic acid (poly(I:C))-induced necroptosis and inflammatory signalling. Furthermore, EPEC infection inhibits TNF-induced phosphorylation and plasma membrane localization of mixed lineage kinase domain-like pseudokinase (MLKL). In vivo, EspL cysteine protease activity contributes to persistent colonization of mice by the EPEC-like mouse pathogen Citrobacter rodentium. The activity of EspL defines a family of T3SS cysteine protease effectors found in a range of bacteria and reveals a mechanism by which gastrointestinal pathogens directly target RHIM-dependent inflammatory and necroptotic signalling pathways.

Injuries to the vascular endothelium: vascular wall and endothelial dysfunction.

PubMed

Fisher, Mark

2008-01-01

Vascular endothelial injury has multiple elements, and this article focuses on ischemia-related processes that have particular relevance to ischemic stroke. Distinctions between necrotic and apoptotic cell death provide a basic science context in which to better understand the significance of classical core and penumbra concepts of acute stroke, with apoptotic processes particularly prominent in the penumbra. The mitochondria are understood to serve as a reservoir of proteins that mediate apoptosis. Oxidative stress pathways generating reactive oxygen species (ROS) are prominent in endothelial injury, both ischemic and nonischemic, with prominent roles of enzyme- and nonenzymemediated pathways; mitochondria once again have a critical role, particularly in the nonenzymatic pathways generating ROS. Inflammation also contributes to vascular endothelial injury, and endothelial cells have the capacity to rapidly increase expression of inflammatory mediators following ischemic challenge; this leads to enhanced leukocyte-endothelial interactions mediated by selectins and adhesion molecules. Preconditioning consists of a minor version of an injurious event, which in turn may protect vascular endothelium from injury following a more substantial event. Presence of the blood-brain barrier creates unique responses to endothelial injury, with permeability changes due to impairment of endothelial-matrix interactions compounding altered vasomotor tone and tissue perfusion mediated by nitric oxide. Pharmacological protection against vascular endothelial injury can be provided by several of the phosphodiesterases (cilostazol and dipyridamole), along with statins. Optimal clinical responses for protection of brain vascular endothelium may use preconditioning as a model, and will likely require combined protection against apoptosis, ROS, and inflammation.

ERManI (Endoplasmic Reticulum Class I α-Mannosidase) Is Required for HIV-1 Envelope Glycoprotein Degradation via Endoplasmic Reticulum-associated Protein Degradation Pathway.

PubMed

Zhou, Tao; Frabutt, Dylan A; Moremen, Kelley W; Zheng, Yong-Hui

2015-09-04

Previously, we reported that the mitochondrial translocator protein (TSPO) induces HIV-1 envelope (Env) degradation via the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway, but the mechanism was not clear. Here we investigated how the four ER-associated glycoside hydrolase family 47 (GH47) α-mannosidases, ERManI, and ER-degradation enhancing α-mannosidase-like (EDEM) proteins 1, 2, and 3, are involved in the Env degradation process. Ectopic expression of these four α-mannosidases uncovers that only ERManI inhibits HIV-1 Env expression in a dose-dependent manner. In addition, genetic knock-out of the ERManI gene MAN1B1 using CRISPR/Cas9 technology disrupts the TSPO-mediated Env degradation. Biochemical studies show that HIV-1 Env interacts with ERManI, and between the ERManI cytoplasmic, transmembrane, lumenal stem, and lumenal catalytic domains, the catalytic domain plays a critical role in the Env-ERManI interaction. In addition, functional studies show that inactivation of the catalytic sites by site-directed mutagenesis disrupts the ERManI activity. These studies identify ERManI as a critical GH47 α-mannosidase in the ER-associated protein degradation pathway that initiates the Env degradation and suggests that its catalytic domain and enzymatic activity play an important role in this process. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Functional characterization of the first filamentous fungal tRNA-isopentenyltransferase and its role in the virulence of Claviceps purpurea.

PubMed

Hinsch, Janine; Galuszka, Petr; Tudzynski, Paul

2016-08-01

In plants, cytokinins (CKs) are synthesized de novo or by the degradation of modified tRNAs. Recently, the first fungal de novo pathway was identified within the plant pathogen Claviceps purpurea. As the deletion of the de novo pathway did not lead to a complete loss of CKs, this work focuses on the tRNA-modifying protein tRNA-isopentenyltransferase (CptRNA-IPT). The contribution of this enzyme to the CK pool of Claviceps and the role of CKs in the host-pathogen interaction are emphasized. The effects of the deletion of cptRNA-ipt and the double deletion of cptRNA-ipt and the key gene of de novo biosynthesis cpipt-log on growth, CK biosynthesis and virulence were analyzed. In addition, the sites of action of CptRNA-IPT were visualized using reporter gene fusions. In addition to CK-independent functions, CptRNA-IPT was essential for the biosynthesis of cis-zeatin (cZ) and contributed to the formation of isopentenyladenine (iP) and trans-zeatin (tZ). Although ΔcptRNA-ipt was reduced in virulence, the 'CK-free' double deletion mutant was nearly apathogenic. The results prove a redundancy of the CK biosynthesis pathway in C. purpurea for iP and tZ formation. Moreover, we show, for the first time, that CKs are required for the successful establishment of a host-fungus interaction. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

PPARα siRNA–Treated Expression Profiles Uncover the Causal Sufficiency Network for Compound-Induced Liver Hypertrophy

PubMed Central

Dai, Xudong; Souza, Angus T. De; Dai, Hongyue; Lewis, David L; Lee, Chang-kyu; Spencer, Andy G; Herweijer, Hans; Hagstrom, Jim E; Linsley, Peter S; Bassett, Douglas E; Ulrich, Roger G; He, Yudong D

2007-01-01

Uncovering pathways underlying drug-induced toxicity is a fundamental objective in the field of toxicogenomics. Developing mechanism-based toxicity biomarkers requires the identification of such novel pathways and the order of their sufficiency in causing a phenotypic response. Genome-wide RNA interference (RNAi) phenotypic screening has emerged as an effective tool in unveiling the genes essential for specific cellular functions and biological activities. However, eliciting the relative contribution of and sufficiency relationships among the genes identified remains challenging. In the rodent, the most widely used animal model in preclinical studies, it is unrealistic to exhaustively examine all potential interactions by RNAi screening. Application of existing computational approaches to infer regulatory networks with biological outcomes in the rodent is limited by the requirements for a large number of targeted permutations. Therefore, we developed a two-step relay method that requires only one targeted perturbation for genome-wide de novo pathway discovery. Using expression profiles in response to small interfering RNAs (siRNAs) against the gene for peroxisome proliferator-activated receptor α (Ppara), our method unveiled the potential causal sufficiency order network for liver hypertrophy in the rodent. The validity of the inferred 16 causal transcripts or 15 known genes for PPARα-induced liver hypertrophy is supported by their ability to predict non-PPARα–induced liver hypertrophy with 84% sensitivity and 76% specificity. Simulation shows that the probability of achieving such predictive accuracy without the inferred causal relationship is exceedingly small (p < 0.005). Five of the most sufficient causal genes have been previously disrupted in mouse models; the resulting phenotypic changes in the liver support the inferred causal roles in liver hypertrophy. Our results demonstrate the feasibility of defining pathways mediating drug-induced toxicity from siRNA-treated expression profiles. When combined with phenotypic evaluation, our approach should help to unleash the full potential of siRNAs in systematically unveiling the molecular mechanism of biological events. PMID:17335344

Activation of β-catenin and Yap1 in human hepatoblastoma and induction of hepatocarcinogenesis in mice.

PubMed

Tao, Junyan; Calvisi, Diego F; Ranganathan, Sarangarajan; Cigliano, Antonio; Zhou, Lili; Singh, Sucha; Jiang, Lijie; Fan, Biao; Terracciano, Luigi; Armeanu-Ebinger, Sorin; Ribback, Silvia; Dombrowski, Frank; Evert, Matthias; Chen, Xin; Monga, Satdarshan P S

2014-09-01

Aberrant activation of β-catenin and Yes-associated protein 1 (Yap1) signaling pathways have been associated with the development of multiple tumor types. Yap functions as a transcriptional coactivator by interacting with TEA domain DNA binding proteins. We investigated the interactions among these pathways during hepatic tumorigenesis. We used immunohistochemical analysis to determine expression of β-catenin and Yap1 in liver cancer specimens collected from patients in Europe and the United States, consisting of 104 hepatocellular carcinoma, 62 intrahepatic cholangiocarcinoma, and 94 hepatoblastoma samples. We assessed β-catenin and Yap1 signaling and interactions in hepatoblastoma cell lines ((HuH6, HepG2, HepT1, HC-AFW1, HepG2, and HC-AFW1); proteins were knocked down with small interfering RNAs, and effects on proliferation and cell death were measured. Sleeping beauty-mediated hydrodynamic transfection was used to overexpress constitutively active forms of β-catenin (ΔN90/β-catenin) and Yap1 (YapS127A) in livers of mice; tissues were collected, and histological and immunohistochemical analyses were performed. We observed nuclear localization of β-catenin and Yap1 in 79% of hepatoblastoma samples but not in most hepatocellular carcinoma or intrahepatic cholangiocarcinoma samples. Yap1 and β-catenin coprecipitated in hepatoblastoma but not hepatocellular carcinoma cells. Small interfering RNA-mediated knockdown of Yap1 or β-catenin in hepatoblastoma cells reduced proliferation in an additive manner. Knockdown of Yap1 reduced its ability to coactivate transcription with β-catenin; β-catenin inhibitors inactivated Yap1. Overexpression of constitutively active forms of Yap1 and β-catenin in mouse liver led to rapid tumorigenesis, with 100% mortality by 11 weeks. Tumor cells expressed both proteins, and human hepatoblastoma cells expressed common targets of their 2 signaling pathways. Yap1 binding of TEA domain factors was required for tumorigenesis in mice. β-catenin and the transcriptional regulator Yap1 interact physically and are activated in most human hepatoblastoma tissues; overexpression of activated forms of these proteins in livers of mice leads to rapid tumor development. Further analysis of these mice will allow further studies of these pathways in hepatoblastoma pathogenesis and could lead to the identification of new therapeutic targets. Copyright © 2014 AGA Institute. Published by Elsevier Inc. All rights reserved.

Activation of β Catenin and Yap1 in Human Hepatoblastoma and Induction of Hepatocarcinogenesis in Mice

PubMed Central

Cigliano, Antonio; Zhou, Lili; Singh, Sucha; Jiang, Lijie; Fan, Biao; Terracciano, Luigi; Armeanu-Ebinger, Sorin; Ribback, Silvia; Dombrowski, Frank; Evert, Matthias; Chen, Xin; Monga, Satdarshan P. S.

2014-01-01

Background & Aims Aberrant activation of βcatenin and Yes-associated protein 1 (Yap1) signaling pathways have been associated with development of multiple tumor types. Yap functions as a transcriptional co-activator by interacting with TEAD DNA binding proteins. We investigated the interactions among these pathways during hepatic tumorigenesis. Methods We used immunohistochemical analysis to determine expression of β-catenin and Yap1 in liver cancer specimens collected from patients in Europe and the US, consisting of 104 hepatocellular carcinoma (HCC), 62 intrahepatic cholangiocarcinoma (ICC), and 94 hepatoblastoma samples. We assessed βcatenin and Yap1 signaling and interactions in hepatoblastoma cell lines ((HuH6, HepG2, HepT1, HC-AFW1, HepG2, and HC-AFW1); proteins were knocked down with small interfering (si)RNAs and effects on proliferation and cell death were measured. Sleeping beauty-mediated hydrodynamic transfection was used to overexpress constitutively active forms of β catenin ( N90-βcatenin) and Yap1 (YapS127A) in livers of mice; tissues were collected and histologic and immunohistochemical analyses were performed. Results We observed nuclear localization of βcatenin and Yap1 in 79% of hepatoblastoma samples, but not in most HCC or ICC tissues. Yap1 and β catenin co-precipitated in hepatoblastoma but not HCC cells. siRNA-mediated knockdown of Yap1 or β catenin in hepatoblastoma cells reduced proliferation in an additive manner. Knockdown of Yap1 reduced its ability to co-activate transcription with βcatenin; βcatenin inhibitors inactivated Yap1. Overexpression of constitutively active forms of Yap1 and βcatenin in mouse liver led to rapid tumorigenesis, with 100% mortality by 11 weeks. Tumors cells expressed both proteins, and human hepatoblastoma cells expressed common targets of their 2 signaling pathways. Yap1 binding of TEAD factors was required for tumorigenesis in mice. Conclusions β catenin and the transcriptional regulator Yap1 interact physically and are activated in most human hepatoblastoma tissues; overexpression of activated forms of these proteins in livers of mice leads to rapid tumor development. Further analysis of these mice will allow further studies of these pathways in hepatoblastoma pathogenesis and could lead to the identification of new therapeutic targets. PMID:24837480

IKKγ-Mimetic Peptides Block the Resistance to Apoptosis Associated with Kaposi's Sarcoma-Associated Herpesvirus Infection.

PubMed

Briggs, Louise C; Chan, A W Edith; Davis, Christopher A; Whitelock, Nicholas; Hotiana, Hajira A; Baratchian, Mehdi; Bagnéris, Claire; Selwood, David L; Collins, Mary K; Barrett, Tracey E

2017-12-01

Primary effusion lymphoma (PEL) is a lymphogenic disorder associated with Kaposi's sarcoma-associated herpesvirus (KSHV) infection. Key to the survival and proliferation of PEL is the canonical NF-κB pathway, which becomes constitutively activated following overexpression of the viral oncoprotein KSHV vFLIP (ks-vFLIP). This arises from its capacity to form a complex with the modulatory subunit of the IκB kinase (IKK) kinase, IKKγ (or NEMO), resulting in the overproduction of proteins that promote cellular survival and prevent apoptosis, both of which are important drivers of tumorigenesis. Using a combination of cell-based and biophysical assays together with structural techniques, we showed that the observed resistance to cell death is largely independent of autophagy or major death receptor signaling pathways and demonstrated that direct targeting of the ks-vFLIP-IKKγ interaction both in cells and in vitro can be achieved using IKKγ-mimetic peptides. Our results further reveal that these peptides not only induce cell killing but also potently sensitize PEL to the proapoptotic agents tumor necrosis factor alpha and etoposide and are the first to confirm ks-vFLIP as a tractable target for the treatment of PEL and related disorders. IMPORTANCE KSHV vFLIP (ks-vFLIP) has been shown to have a crucial role in cellular transformation, in which it is vital for the survival and proliferation of primary effusion lymphoma (PEL), an aggressive malignancy associated with infection that is resistant to the majority of chemotherapeutic drugs. It operates via subversion of the canonical NF-κB pathway, which requires a physical interaction between ks-vFLIP and the IKK kinase modulatory subunit IKKγ. While this interaction has been directly linked to protection against apoptosis, it is unclear whether the suppression of other cell death pathways implicated in ks-vFLIP pathogenesis is an additional contributor. We demonstrate that the interaction between ks-vFLIP and IKKγ is pivotal in conferring resistance to apoptosis. Additionally, we show that the ks-vFLIP-IKKγ complex can be disrupted using peptides leading to direct killing and the sensitization of PEL cells to proapoptotic agents. Our studies thus provide a framework for future therapeutic interventions. Copyright © 2017 Briggs et al.

Cellular Assays for Ferredoxins: A Strategy for Understanding Electron Flow through Protein Carriers That Link Metabolic Pathways.

PubMed

Atkinson, Joshua T; Campbell, Ian; Bennett, George N; Silberg, Jonathan J

2016-12-27

The ferredoxin (Fd) protein family is a structurally diverse group of iron-sulfur proteins that function as electron carriers, linking biochemical pathways important for energy transduction, nutrient assimilation, and primary metabolism. While considerable biochemical information about individual Fd protein electron carriers and their reactions has been acquired, we cannot yet anticipate the proportion of electrons shuttled between different Fd-partner proteins within cells using biochemical parameters that govern electron flow, such as holo-Fd concentration, midpoint potential (driving force), molecular interactions (affinity and kinetics), conformational changes (allostery), and off-pathway electron leakage (chemical oxidation). Herein, we describe functional and structural gaps in our Fd knowledge within the context of a sequence similarity network and phylogenetic tree, and we propose a strategy for improving our understanding of Fd sequence-function relationships. We suggest comparing the functions of divergent Fds within cells whose growth, or other measurable output, requires electron transfer between defined electron donor and acceptor proteins. By comparing Fd-mediated electron transfer with biochemical parameters that govern electron flow, we posit that models that anticipate energy flow across Fd interactomes can be built. This approach is expected to transform our ability to anticipate Fd control over electron flow in cellular settings, an obstacle to the construction of synthetic electron transfer pathways and rational optimization of existing energy-conserving pathways.

The Nuclear Dbf2-Related Kinase COT1 and the Mitogen-Activated Protein Kinases MAK1 and MAK2 Genetically Interact to Regulate Filamentous Growth, Hyphal Fusion and Sexual Development in Neurospora crassa

PubMed Central

Maerz, Sabine; Ziv, Carmit; Vogt, Nico; Helmstaedt, Kerstin; Cohen, Nourit; Gorovits, Rena; Yarden, Oded; Seiler, Stephan

2008-01-01

Ndr kinases, such as Neurospora crassa COT1, are important for cell differentiation and polar morphogenesis, yet their input signals as well as their integration into a cellular signaling context are still elusive. Here, we identify the cot-1 suppressor gul-4 as mak-2 and show that mutants of the gul-4/mak-2 mitogen-activated protein (MAP) kinase pathway suppress cot-1 phenotypes along with a concomitant reduction in protein kinase A (PKA) activity. Furthermore, mak-2 pathway defects are partially overcome in a cot-1 background and are associated with increased MAK1 MAPK signaling. A comparative characterization of N. crassa MAPKs revealed that they act as three distinct modules during vegetative growth and asexual development. In addition, common functions of MAK1 and MAK2 signaling during maintenance of cell-wall integrity distinguished the two ERK-type pathways from the p38-type OS2 osmosensing pathway. In contrast to separate functions during vegetative growth, the concerted activity of the three MAPK pathways is essential for cell fusion and for the subsequent formation of multicellular structures that are required for sexual development. Taken together, our data indicate a functional link between COT1 and MAPK signaling in regulating filamentous growth, hyphal fusion, and sexual development. PMID:18562669

FANCJ Helicase Operates in the Fanconi Anemia DNA Repair Pathway and the Response to Replicational Stress

PubMed Central

Wu, Yuliang; Brosh, Robert M.

2009-01-01

Fanconi anemia (FA) is an autosomal recessive disorder characterized by multiple congenital anomalies, progressive bone marrow failure, and high cancer risk. Cells from FA patients exhibit spontaneous chromosomal instability and hypersensitivity to DNA interstrand cross-linking (ICL) agents. Although the precise mechanistic details of the FA/BRCA pathway of ICL-repair are not well understood, progress has been made in the identification of the FA proteins that are required for the pathway. Among the 13 FA complementation groups from which all the FA genes have been cloned, only a few of the FA proteins are predicted to have direct roles in DNA metabolism. One of the more recently identified FA proteins, shown to be responsible for complementation of the FA complementation group J, is the BRCA1 Associated C-terminal Helicase (BACH1, designated FANCJ), originally identified as a protein associated with breast cancer. FANCJ has been proposed to function downstream of FANCD2 monoubiquitination, a critical event in the FA pathway. Evidence supports a role for FANCJ in a homologous recombination (HR) pathway of double strand break (DSB) repair. In this review, we will summarize the current knowledge in terms of FANCJ functions through its enzymatic activities and protein interactions. The molecular roles of FANCJ in DNA repair and the response to replicational stress will be discussed. PMID:19519404

Role of Rho/ROCK and p38 MAP kinase pathways in transforming growth factor-beta-mediated Smad-dependent growth inhibition of human breast carcinoma cells in vivo.

PubMed

Kamaraju, Anil K; Roberts, Anita B

2005-01-14

TGF-beta is a multifunctional cytokine known to exert its biological effects through a variety of signaling pathways of which Smad signaling is considered to be the main mediator. At present, the Smad-independent pathways, their interactions with each other, and their roles in TGF-beta-mediated growth inhibitory effects are not well understood. To address these questions, we have utilized a human breast cancer cell line MCF10CA1h and demonstrate that p38 MAP kinase and Rho/ROCK pathways together with Smad2 and Smad3 are necessary for TGF-beta-mediated growth inhibition of this cell line. We show that Smad2/3 are indispensable for TGF-beta-mediated growth inhibition, and that both p38 and Rho/ROCK pathways affect the linker region phosphorylation of Smad2/3. Further, by using Smad3 mutated at the putative phosphorylation sites in the linker region, we demonstrate that phosphorylation at Ser203 and Ser207 residues is required for the full transactivation potential of Smad3, and that these residues are targets of the p38 and Rho/ROCK pathways. We demonstrate that activation of the p38 MAP kinase pathway is necessary for the full transcriptional activation potential of Smad2/Smad3 by TGF-beta, whereas activity of Rho/ROCK is necessary for both down-regulation of c-Myc protein and up-regulation of p21waf1 protein, directly interfering with p21waf1 transcription. Our results not only implicate Rho/ROCK and p38 MAPK pathways as necessary for TGF-beta-mediated growth inhibition, but also demonstrate their individual contributions and the basis for their cooperation with each other.

Multiple cytoskeletal pathways and PI3K signaling mediate CDC-42-induced neuronal protrusion in C. elegans.

PubMed

Alan, Jamie K; Struckhoff, Eric C; Lundquist, Erik A

2013-01-01

Rho GTPases are key regulators of cellular protrusion and are involved in many developmental events including axon guidance during nervous system development. Rho GTPase pathways display functional redundancy in developmental events, including axon guidance. Therefore, their roles can often be masked when using simple loss-of-function genetic approaches. As a complement to loss-of-function genetics, we constructed a constitutively activated CDC-42(G12V) expressed in C. elegans neurons. CDC-42(G12V) drove the formation of ectopic lamellipodial and filopodial protrusions in the PDE neurons, which resembled protrusions normally found on migrating growth cones of axons. We then used a candidate gene approach to identify molecules that mediate CDC-42(G12V)-induced ectopic protrusions by determining if loss of function of the genes could suppress CDC-42(G12V). Using this approach, we identified 3 cytoskeletal pathways previously implicated in axon guidance, the Arp2/3 complex, UNC-115/abLIM, and UNC-43/Ena. We also identified the Nck-interacting kinase MIG-15/NIK and p21-activated kinases (PAKs), also implicated in axon guidance. Finally, PI3K signaling was required, specifically the Rictor/mTORC2 branch but not the mTORC1 branch that has been implicated in other aspects of PI3K signaling including stress and aging. Our results indicate that multiple pathways can mediate CDC-42-induced neuronal protrusions that might be relevant to growth cone protrusions during axon pathfinding. Each of these pathways involves Rac GTPases, which might serve to integrate the pathways and coordinate the multiple CDC-42 pathways. These pathways might be relevant to developmental events such as axon pathfinding as well as disease states such as metastatic melanoma.

Multiple cytoskeletal pathways and PI3K signaling mediate CDC-42-induced neuronal protrusion in C. elegans

PubMed Central

Alan, Jamie K; Struckhoff, Eric C; Lundquist, Erik A

2013-01-01

Rho GTPases are key regulators of cellular protrusion and are involved in many developmental events including axon guidance during nervous system development. Rho GTPase pathways display functional redundancy in developmental events, including axon guidance. Therefore, their roles can often be masked when using simple loss-of-function genetic approaches. As a complement to loss-of-function genetics, we constructed a constitutively activated CDC-42(G12V) expressed in C. elegans neurons. CDC-42(G12V) drove the formation of ectopic lamellipodial and filopodial protrusions in the PDE neurons, which resembled protrusions normally found on migrating growth cones of axons. We then used a candidate gene approach to identify molecules that mediate CDC-42(G12V)-induced ectopic protrusions by determining if loss of function of the genes could suppress CDC-42(G12V). Using this approach, we identified 3 cytoskeletal pathways previously implicated in axon guidance, the Arp2/3 complex, UNC-115/abLIM, and UNC-43/Ena. We also identified the Nck-interacting kinase MIG-15/NIK and p21-activated kinases (PAKs), also implicated in axon guidance. Finally, PI3K signaling was required, specifically the Rictor/mTORC2 branch but not the mTORC1 branch that has been implicated in other aspects of PI3K signaling including stress and aging. Our results indicate that multiple pathways can mediate CDC-42-induced neuronal protrusions that might be relevant to growth cone protrusions during axon pathfinding. Each of these pathways involves Rac GTPases, which might serve to integrate the pathways and coordinate the multiple CDC-42 pathways. These pathways might be relevant to developmental events such as axon pathfinding as well as disease states such as metastatic melanoma. PMID:24149939

Interactions between the jasmonic and salicylic acid pathway modulate the plant metabolome and affect herbivores of different feeding types.

PubMed

Schweiger, R; Heise, A-M; Persicke, M; Müller, C

2014-07-01

The phytohormones jasmonic acid (JA) and salicylic acid (SA) mediate induced plant defences and the corresponding pathways interact in a complex manner as has been shown on the transcript and proteine level. Downstream, metabolic changes are important for plant-herbivore interactions. This study investigated metabolic changes in leaf tissue and phloem exudates of Plantago lanceolata after single and combined JA and SA applications as well as consequences on chewing-biting (Heliothis virescens) and piercing-sucking (Myzus persicae) herbivores. Targeted metabolite profiling and untargeted metabolic fingerprinting uncovered different categories of plant metabolites, which were influenced in a specific manner, indicating points of divergence, convergence, positive crosstalk and pronounced mutual antagonism between the signaling pathways. Phytohormone-specific decreases of primary metabolite pool sizes in the phloem exudates may indicate shifts in sink-source relations, resource allocation, nutrient uptake or photosynthesis. Survival of both herbivore species was significantly reduced by JA and SA treatments. However, the combined application of JA and SA attenuated the negative effects at least against H. virescens suggesting that mutual antagonism between the JA and SA pathway may be responsible. Pathway interactions provide a great regulatory potential for the plant that allows triggering of appropriate defences when attacked by different antagonist species. © 2013 John Wiley & Sons Ltd.

BRCA1 interacts directly with the Fanconi anemia protein FANCA.

PubMed

Folias, Alexandra; Matkovic, Mara; Bruun, Donald; Reid, Sonja; Hejna, James; Grompe, Markus; D'Andrea, Alan; Moses, Robb

2002-10-01

Fanconi anemia (FA) is a rare autosomal recessive disease characterized by skeletal defects, anemia, chromosomal instability and increased risk of leukemia. At the cellular level FA is characterized by increased sensitivity to agents forming interstrand crosslinks (ICL) in DNA. Six FA genes have been cloned and interactions among individual FANC proteins have been found. The FANCD2 protein co-localizes in nuclear foci with the BRCA1 protein following DNA damage and during S-phase, requiring the FANCA, C, E and G proteins to do so. This finding may reflect a direct role for the BRCA1 protein in double strand break (DSB) repair and interaction with the FANC proteins. Therefore interactions between BRCA1 and the FANC proteins were investigated. Among the known FANC proteins, we find evidence for direct interaction only between the FANCA protein and BRCA1. The evidence rests on three different tests: yeast two-hybrid analysis, coimmunoprecipitation from in vitro synthesis, and coimmunoprecipitation from cell extracts. The amino terminal portion of FANCA and the central part (aa 740-1083) of BRCA1 contain the sites of interaction. The interaction does not depend on DNA damage, thus FANCA and BRCA1 are constitutively interacting. The demonstrated interaction directly connects BRCA1 to the FA pathway of DNA repair.

Common drug-drug interactions in antifungal treatments for superficial fungal infections.

PubMed

Gupta, Aditya K; Versteeg, Sarah G; Shear, Neil H

2018-04-01

Antifungal agents can be co-administered alongside several other medications for a variety of reasons such as the presence of comorbidities. Pharmacodynamic interactions such as synergistic and antagonistic interactions could be the result of co-administered medications. Pharmacokinetic interactions could also transpire through the inhibition of metabolizing enzymes and drug transport systems, altering the absorption, metabolism and excretion of co-administered medications. Both pharmacodynamic and pharmacokinetic interactions can result in hospitalization due to serious adverse effects associated with antifungal agents, lower therapeutic doses required to achieve desired antifungal activity, and prevent antifungal resistance. Areas covered: The objective of this review is to summarize pharmacodynamic and pharmacokinetic interactions associated with common antifungal agents used to treat superficial fungal infections. Pharmacodynamic and pharmacokinetic interactions that impact the therapeutic effects of antifungal agents and drugs that are influenced by the presence of antifungal agents was the context to which these antifungal agents were addressed. Expert opinion: The potential for drug-drug interactions is minimal for topical antifungals as opposed to oral antifungals as they have minimal exposure to other co-administered medications. Developing non-lipophilic antifungals that have unique metabolizing pathways and are topical applied are suggested properties that could help limit drug-drug interactions associated with future treatments.

Integrated Interactive Chart as a Tool for Teaching Metabolic Pathways

ERIC Educational Resources Information Center

Kalogiannis, Stavros; Pagkalos, Ioannis; Koufoudakis, Panagiotis; Dashi, Ino; Pontikeri, Kyriaki; Christodoulou, Constantina

2014-01-01

An interactive chart of energy metabolism with didactic function, complementary to the already existing metabolic maps, located at the URL www.metpath.teithe.gr is being presented. The chart illustrates the major catabolic and biosynthetic pathways of glucose, fatty acids, and aminoacids, individually as well as in an integrated view. For every…

Not just black and white: pigment pattern development and evolution in vertebrates

PubMed Central

Mills, Margaret G.; Patterson, Larissa B.

2009-01-01

Animals display diverse colors and patterns that vary within and between species. Similar phenotypes appear in both closely related and widely divergent taxa. Pigment patterns thus provide an opportunity to explore how development is altered to produce differences in form and whether similar phenotypes share a common genetic basis. Understanding the development and evolution of pigment patterns requires knowledge of the cellular interactions and signaling pathways that produce those patterns. These complex traits provide unparalleled opportunities for integrating studies from ecology and behavior to molecular biology and biophysics. PMID:19073271

Energetics and Dynamics Across the Bcl-2-Regulated Apoptotic Pathway Reveal Distinct Evolutionary Determinants of Specificity and Affinity.

PubMed

Ivanov, Stefan M; Huber, Roland G; Warwicker, Jim; Bond, Peter J

2016-11-01

Critical regulatory pathways are replete with instances of intra- and interfamily protein-protein interactions due to the pervasiveness of gene duplication throughout evolution. Discerning the specificity determinants within these systems has proven a challenging task. Here, we present an energetic analysis of the specificity determinants within the Bcl-2 family of proteins (key regulators of the intrinsic apoptotic pathway) via a total of ∼20 μs of simulation of 60 distinct protein-protein complexes. We demonstrate where affinity and specificity of protein-protein interactions arise across the family, and corroborate our conclusions with extensive experimental evidence. We identify energy and specificity hotspots that may offer valuable guidance in the design of targeted therapeutics for manipulating the protein-protein interactions within the apoptosis-regulating pathway. Moreover, we propose a conceptual framework that allows us to quantify the relationship between sequence, structure, and binding energetics. This approach may represent a general methodology for investigating other paralogous protein-protein interaction sites. Copyright © 2016 Elsevier Ltd. All rights reserved.

The Fibroblast Growth Factor signaling pathway

PubMed Central

Ornitz, David M; Itoh, Nobuyuki

2015-01-01

The signaling component of the mammalian Fibroblast Growth Factor (FGF) family is comprised of eighteen secreted proteins that interact with four signaling tyrosine kinase FGF receptors (FGFRs). Interaction of FGF ligands with their signaling receptors is regulated by protein or proteoglycan cofactors and by extracellular binding proteins. Activated FGFRs phosphorylate specific tyrosine residues that mediate interaction with cytosolic adaptor proteins and the RAS-MAPK, PI3K-AKT, PLCγ, and STAT intracellular signaling pathways. Four structurally related intracellular non-signaling FGFs interact with and regulate the family of voltage gated sodium channels. Members of the FGF family function in the earliest stages of embryonic development and during organogenesis to maintain progenitor cells and mediate their growth, differentiation, survival, and patterning. FGFs also have roles in adult tissues where they mediate metabolic functions, tissue repair, and regeneration, often by reactivating developmental signaling pathways. Consistent with the presence of FGFs in almost all tissues and organs, aberrant activity of the pathway is associated with developmental defects that disrupt organogenesis, impair the response to injury, and result in metabolic disorders, and cancer. © 2015 Wiley Periodicals, Inc. PMID:25772309

Wnt7a interaction with Fzd5 and detection of signaling activation using a split eGFP

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

Carmon, Kendra S.; Loose, David S.

2008-04-04

Wnts are secreted glycoproteins that regulate important cellular processes including proliferation, differentiation, and cell fate. In the {beta}-catenin/canonical pathway, Wnt interacts with Fzd receptors to inhibit degradation of {beta}-catenin and promote its translocation into the nucleus where it regulates transcription of a number of genes. Dysregulation of this pathway has been attributed to a host of diseases including cancer. As a result, components of the {beta}-catenin/canonical pathway have been gaining recognition as promising targets for the discovery of novel therapeutic agents. Here, we show, using an ELISA-based protein-protein binding assay that purified Wnt7a binds to the extracellular cysteine-rich domain ofmore » Fzd5 in the nanomolar range. We have developed a novel split eGFP complementation assay to visually detect Wnt7a-Fzd5 interactions and subsequent pathway activation in cells. These biological tools could help lead to a better understanding of Wnt-Fzd interactions and the identification of new modulators of Wnt signaling.« less

Illuminating the Reaction Pathways of Viromimetic Assembly.

PubMed

Cingil, Hande E; Boz, Emre B; Biondaro, Giovanni; de Vries, Renko; Cohen Stuart, Martien A; Kraft, Daniela J; van der Schoot, Paul; Sprakel, Joris

2017-04-05

The coassembly of well-defined biological nanostructures relies on a delicate balance between attractive and repulsive interactions between biomolecular building blocks. Viral capsids are a prototypical example, where coat proteins exhibit not only self-interactions but also interact with the cargo they encapsulate. In nature, the balance between antagonistic and synergistic interactions has evolved to avoid kinetic trapping and polymorphism. To date, it has remained a major challenge to experimentally disentangle the complex kinetic reaction pathways that underlie successful coassembly of biomolecular building blocks in a noninvasive approach with high temporal resolution. Here we show how macromolecular force sensors, acting as a genome proxy, allow us to probe the pathways through which a viromimetic protein forms capsids. We uncover the complex multistage process of capsid assembly, which involves recruitment and complexation, followed by allosteric growth of the proteinaceous coat. Under certain conditions, the single-genome particles condense into capsids containing multiple copies of the template. Finally, we derive a theoretical model that quantitatively describes the kinetics of recruitment and growth. These results shed new light on the origins of the pathway complexity in biomolecular coassembly.

ICE1 promotes the link between splicing and nonsense-mediated mRNA decay

PubMed Central

Baird, Thomas D; Cheng, Ken Chih-Chien; Chen, Yu-Chi; Buehler, Eugen; Martin, Scott E; Inglese, James

2018-01-01

The nonsense-mediated mRNA decay (NMD) pathway detects aberrant transcripts containing premature termination codons (PTCs) and regulates expression of 5–10% of non-aberrant human mRNAs. To date, most proteins involved in NMD have been identified by genetic screens in model organisms; however, the increased complexity of gene expression regulation in human cells suggests that additional proteins may participate in the human NMD pathway. To identify proteins required for NMD, we performed a genome-wide RNAi screen against >21,000 genes. Canonical members of the NMD pathway were highly enriched as top hits in the siRNA screen, along with numerous candidate NMD factors, including the conserved ICE1/KIAA0947 protein. RNAseq studies reveal that depletion of ICE1 globally enhances accumulation and stability of NMD-target mRNAs. Further, our data suggest that ICE1 uses a putative MIF4G domain to interact with exon junction complex (EJC) proteins and promotes the association of the NMD protein UPF3B with the EJC. PMID:29528287

piRNA pathway targets active LINE1 elements to establish the repressive H3K9me3 mark in germ cells

PubMed Central

Pezic, Dubravka; Manakov, Sergei A.; Sachidanandam, Ravi; Aravin, Alexei A.

2014-01-01

Transposable elements (TEs) occupy a large fraction of metazoan genomes and pose a constant threat to genomic integrity. This threat is particularly critical in germ cells, as changes in the genome that are induced by TEs will be transmitted to the next generation. Small noncoding piwi-interacting RNAs (piRNAs) recognize and silence a diverse set of TEs in germ cells. In mice, piRNA-guided transposon repression correlates with establishment of CpG DNA methylation on their sequences, yet the mechanism and the spectrum of genomic targets of piRNA silencing are unknown. Here we show that in addition to DNA methylation, the piRNA pathway is required to maintain a high level of the repressive H3K9me3 histone modification on long interspersed nuclear elements (LINEs) in germ cells. piRNA-dependent chromatin repression targets exclusively full-length elements of actively transposing LINE families, demonstrating the remarkable ability of the piRNA pathway to recognize active elements among the large number of genomic transposon fragments. PMID:24939875

Daple coordinates organ-wide and cell-intrinsic polarity to pattern inner-ear hair bundles

PubMed Central

Siletti, Kimberly; Hudspeth, A. J.

2017-01-01

The establishment of planar polarization by mammalian cells necessitates the integration of diverse signaling pathways. In the inner ear, at least two systems regulate the planar polarity of sensory hair bundles. The core planar cell polarity (PCP) proteins coordinate the orientations of hair cells across the epithelial plane. The cell-intrinsic patterning of hair bundles is implemented independently by the G protein complex classically known for orienting the mitotic spindle. Although the primary cilium also participates in each of these pathways, its role and the integration of the two systems are poorly understood. We show that Dishevelled-associating protein with a high frequency of leucine residues (Daple) interacts with PCP and cell-intrinsic signals. Regulated by the cell-intrinsic pathway, Daple is required to maintain the polarized distribution of the core PCP protein Dishevelled and to position the primary cilium at the abneural edge of the apical surface. Our results suggest that the primary cilium or an associated structure influences the domain of cell-intrinsic signals that shape the hair bundle. Daple is therefore essential to orient and pattern sensory hair bundles. PMID:29229865

Dynamic pathway modeling of signal transduction networks: a domain-oriented approach.

PubMed

Conzelmann, Holger; Gilles, Ernst-Dieter

2008-01-01

Mathematical models of biological processes become more and more important in biology. The aim is a holistic understanding of how processes such as cellular communication, cell division, regulation, homeostasis, or adaptation work, how they are regulated, and how they react to perturbations. The great complexity of most of these processes necessitates the generation of mathematical models in order to address these questions. In this chapter we provide an introduction to basic principles of dynamic modeling and highlight both problems and chances of dynamic modeling in biology. The main focus will be on modeling of s transduction pathways, which requires the application of a special modeling approach. A common pattern, especially in eukaryotic signaling systems, is the formation of multi protein signaling complexes. Even for a small number of interacting proteins the number of distinguishable molecular species can be extremely high. This combinatorial complexity is due to the great number of distinct binding domains of many receptors and scaffold proteins involved in signal transduction. However, these problems can be overcome using a new domain-oriented modeling approach, which makes it possible to handle complex and branched signaling pathways.

Retinal Determination genes function along with cell-cell signals to regulate Drosophila eye development: examples of multi-layered regulation by Master Regulators

PubMed Central

Baker, Nicholas E.; Firth, Lucy C.

2015-01-01

It is thought that Retinal Determination gene products define the response made to cell-cell signals within the eye developmental field by binding to enhancers of genes that are also regulated by cell-cell signaling pathways. In Drosophila, Retinal Determination genes including Eyeless, teashirt, eyes absent, dachsous and sine oculis, are required for normal eye development and can induce ectopic eyes when mis-expressed. Characterization of the enhancers responsible for eye expression of the hedgehog, shaven, and atonal genes, as well as the dynamics of Retinal Determination gene expression themselves, now suggest a multilayered network whereby transcriptional regulation by either Retinal Determination genes or cell-cell signaling pathways can sometimes be indirect and mediated by other transcription factor intermediates. In this updated view of the interaction between extracellular information and cell intrinsic programs during development, regulation of individual genes might sometimes be several steps removed from either the Retinal Determination genes or cell-cell signaling pathways that nevertheless govern their expression. PMID:21607995

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

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

PubMed

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

2007-12-12

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 alpha-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.

Effect of occupational exposures on lung cancer susceptibility: a study of gene-environment interaction analysis.

PubMed

Malhotra, Jyoti; Sartori, Samantha; Brennan, Paul; Zaridze, David; Szeszenia-Dabrowska, Neonila; Świątkowska, Beata; Rudnai, Peter; Lissowska, Jolanta; Fabianova, Eleonora; Mates, Dana; Bencko, Vladimir; Gaborieau, Valerie; Stücker, Isabelle; Foretova, Lenka; Janout, Vladimir; Boffetta, Paolo

2015-03-01

Occupational exposures are known risk factors for lung cancer. Role of genetically determined host factors in occupational exposure-related lung cancer is unclear. We used genome-wide association (GWA) data from a case-control study conducted in 6 European countries from 1998 to 2002 to identify gene-occupation interactions and related pathways for lung cancer risk. GWA analysis was performed for each exposure using logistic regression and interaction term for genotypes, and exposure was included in this model. Both SNP-based and gene-based interaction P values were calculated. Pathway analysis was performed using three complementary methods, and analyses were adjusted for multiple comparisons. We analyzed 312,605 SNPs and occupational exposure to 70 agents from 1,802 lung cancer cases and 1,725 cancer-free controls. Mean age of study participants was 60.1 ± 9.1 years and 75% were male. Largest number of significant associations (P ≤ 1 × 10(-5)) at SNP level was demonstrated for nickel, brick dust, concrete dust, and cement dust, and for brick dust and cement dust at the gene-level (P ≤ 1 × 10(-4)). Approximately 14 occupational exposures showed significant gene-occupation interactions with pathways related to response to environmental information processing via signal transduction (P < 0.001 and FDR < 0.05). Other pathways that showed significant enrichment were related to immune processes and xenobiotic metabolism. Our findings suggest that pathways related to signal transduction, immune process, and xenobiotic metabolism may be involved in occupational exposure-related lung carcinogenesis. Our study exemplifies an integrative approach using pathway-based analysis to demonstrate the role of genetic variants in occupational exposure-related lung cancer susceptibility. Cancer Epidemiol Biomarkers Prev; 24(3); 570-9. ©2015 AACR. ©2015 American Association for Cancer Research.

N-Glycosylation of the alpha subunit does not influence trafficking or functional activity of the human organic solute transporter alpha/beta

PubMed Central

Soroka, Carol J; Xu, Shuhua; Mennone, Albert; Lam, Ping; Boyer, James L

2008-01-01

Background The organic solute transporter (OSTα-OSTβ) is a heteromeric transporter that is expressed on the basolateral membrane of epithelium in intestine, kidney, liver, testis and adrenal gland and facilitates efflux of bile acids and other steroid solutes. Both subunits are required for plasma membrane localization of the functional transporter but it is unclear how and where the subunits interact and whether glycosylation is required for functional activity. We sought to examine these questions for the human OSTα-OSTβ transporter using the human hepatoma cell line, HepG2, and COS7 cells transfected with constructs of human OSTα-FLAG and OSTβ-Myc. Results Tunicamycin treatment demonstrated that human OSTα is glycosylated. In COS7 cells Western blotting identified the unglycosylated form (~31 kD), the core precursor form (~35 kD), and the mature, complex glycoprotein (~40 kD). Immunofluorescence of both cells indicated that, in the presence of OSTβ, the alpha subunit could still be expressed on the plasma membrane after tunicamycin treatment. Furthermore, the functional uptake of 3H-estrone sulfate was unchanged in the absence of N-glycosylation. Co-immunoprecipitation indicates that the immature form of OSTα interact with OSTβ. However, immunoprecipitation of OSTβ using an anti-Myc antibody did not co-precipitate the mature, complex glycosylated form of OSTα, suggesting that the primary interaction occurs early in the biosynthetic pathway and may be transient. Conclusion In conclusion, human OSTα is a glycoprotein that requires interaction with OSTβ to reach the plasma membrane. However, glycosylation of OSTα is not necessary for interaction with the beta subunit or for membrane localization or function of the heteromeric transporter. PMID:18847488

Caenorhabditis elegans fibroblast growth factor receptor signaling can occur independently of the multi-substrate adaptor FRS2.

PubMed

Lo, Te-Wen; Bennett, Daniel C; Goodman, S Jay; Stern, Michael J

2010-06-01

The components of receptor tyrosine kinase signaling complexes help to define the specificity of the effects of their activation. The Caenorhabditis elegans fibroblast growth factor receptor (FGFR), EGL-15, regulates a number of processes, including sex myoblast (SM) migration guidance and fluid homeostasis, both of which require a Grb2/Sos/Ras cassette of signaling components. Here we show that SEM-5/Grb2 can bind directly to EGL-15 to mediate SM chemoattraction. A yeast two-hybrid screen identified SEM-5 as able to interact with the carboxy-terminal domain (CTD) of EGL-15, a domain that is specifically required for SM chemoattraction. This interaction requires the SEM-5 SH2-binding motifs present in the CTD (Y(1009) and Y(1087)), and these sites are required for the CTD role of EGL-15 in SM chemoattraction. SEM-5, but not the SEM-5 binding sites located in the CTD, is required for the fluid homeostasis function of EGL-15, indicating that SEM-5 can link to EGL-15 through an alternative mechanism. The multi-substrate adaptor protein FRS2 serves to link vertebrate FGFRs to Grb2. In C. elegans, an FRS2-like gene, rog-1, functions upstream of a Ras/MAPK pathway for oocyte maturation but is not required for EGL-15 function. Thus, unlike the vertebrate FGFRs, which require the multi-substrate adaptor FRS2 to recruit Grb2, EGL-15 can recruit SEM-5/Grb2 directly.

Diet and Colorectal Cancer: Analysis of a Candidate Pathway Using SNPS, Haplotypes, and Multi-Gene Assessment

PubMed Central

Slattery, Martha L.; Lundgreen, Abbie; Herrick, Jennifer S.; Caan, Bette J.; Potter, John D.; Wolff, Roger K.

2012-01-01

There is considerable biologic plausibility to the hypothesis that genetic variability in pathways involved in insulin signaling and energy homeostasis may modulate dietary risk associated with colorectal cancer. We utilized data from 2 population-based case-control studies of colon (n = 1,574 cases, 1,970 controls) and rectal (n = 791 cases, 999 controls) cancer to evaluate genetic variation in candidate SNPs identified from 9 genes in a candidate pathway: PDK1, RP6KA1, RPS6KA2, RPS6KB1, RPS6KB2, PTEN, FRAP1 (mTOR), TSC1, TSC2, Akt1, PIK3CA, and PRKAG2 with dietary intake of total energy, carbohydrates, fat, and fiber. We employed SNP, haplotype, and multiple-gene analysis to evaluate associations. PDK1 interacted with dietary fat for both colon and rectal cancer and with dietary carbohydrates for colon cancer. Statistically significant interaction with dietary carbohydrates and rectal cancer was detected by haplotype analysis of PDK1. Evaluation of dietary interactions with multiple genes in this candidate pathway showed several interactions with pairs of genes: Akt1 and PDK1, PDK1 and PTEN, PDK1 and TSC1, and PRKAG2 and PTEN. Analyses show that genetic variation influences risk of colorectal cancer associated with diet and illustrate the importance of evaluating dietary interactions beyond the level of single SNPs or haplotypes when a biologically relevant candidate pathway is examined. PMID:21999454

Interactions between Casein Kinase Iε (CKIε) and Two Substrates from Disparate Signaling Pathways Reveal Mechanisms for Substrate-Kinase Specificity

PubMed Central

Dahlberg, Caroline Lund; Nguyen, Elizabeth Z.; Goodlett, David; Kimelman, David

2009-01-01

Background Members of the Casein Kinase I (CKI) family of serine/threonine kinases regulate diverse biological pathways. The seven mammalian CKI isoforms contain a highly conserved kinase domain and divergent amino- and carboxy-termini. Although they share a preferred target recognition sequence and have overlapping expression patterns, individual isoforms often have specific substrates. In an effort to determine how substrates recognize differences between CKI isoforms, we have examined the interaction between CKIε and two substrates from different signaling pathways. Methodology/Principal Findings CKIε, but not CKIα, binds to and phosphorylates two proteins: Period, a transcriptional regulator of the circadian rhythms pathway, and Disheveled, an activator of the planar cell polarity pathway. We use GST-pull-down assays data to show that two key residues in CKIα's kinase domain prevent Disheveled and Period from binding. We also show that the unique C-terminus of CKIε does not determine Dishevelled's and Period's preference for CKIε nor is it essential for binding, but instead plays an auxillary role in stabilizing the interactions of CKIε with its substrates. We demonstrate that autophosphorylation of CKIε's C-terminal tail prevents substrate binding, and use mass spectrometry and chemical crosslinking to reveal how a phosphorylation-dependent interaction between the C-terminal tail and the kinase domain prevents substrate phosphorylation and binding. Conclusions/Significance The biochemical interactions between CKIε and Disheveled, Period, and its own C-terminus lead to models that explain CKIε's specificity and regulation. PMID:19274088

A comprehensive pathway map of epidermal growth factor receptor signaling

PubMed Central

Oda, Kanae; Matsuoka, Yukiko; Funahashi, Akira; Kitano, Hiroaki

2005-01-01

The epidermal growth factor receptor (EGFR) signaling pathway is one of the most important pathways that regulate growth, survival, proliferation, and differentiation in mammalian cells. Reflecting this importance, it is one of the best-investigated signaling systems, both experimentally and computationally, and several computational models have been developed for dynamic analysis. A map of molecular interactions of the EGFR signaling system is a valuable resource for research in this area. In this paper, we present a comprehensive pathway map of EGFR signaling and other related pathways. The map reveals that the overall architecture of the pathway is a bow-tie (or hourglass) structure with several feedback loops. The map is created using CellDesigner software that enables us to graphically represent interactions using a well-defined and consistent graphical notation, and to store it in Systems Biology Markup Language (SBML). PMID:16729045

"Which Pathway Am I?" Using a Game Approach to Teach Students about Biochemical Pathways

ERIC Educational Resources Information Center

Ooi, Beng Guat; Sanger, Michael J.

2009-01-01

This game was designed to provide students with an alternative way to learn biochemical pathways through an interactive approach. In this game, students worked in pairs to help each other identify pathways taped to each other's backs by asking simple "yes or no" questions related to these pathways. This exercise was conducted after the traditional…

Learning cellular sorting pathways using protein interactions and sequence motifs.

PubMed

Lin, Tien-Ho; Bar-Joseph, Ziv; Murphy, Robert F

2011-11-01

Proper subcellular localization is critical for proteins to perform their roles in cellular functions. Proteins are transported by different cellular sorting pathways, some of which take a protein through several intermediate locations until reaching its final destination. The pathway a protein is transported through is determined by carrier proteins that bind to specific sequence motifs. In this article, we present a new method that integrates protein interaction and sequence motif data to model how proteins are sorted through these sorting pathways. We use a hidden Markov model (HMM) to represent protein sorting pathways. The model is able to determine intermediate sorting states and to assign carrier proteins and motifs to the sorting pathways. In simulation studies, we show that the method can accurately recover an underlying sorting model. Using data for yeast, we show that our model leads to accurate prediction of subcellular localization. We also show that the pathways learned by our model recover many known sorting pathways and correctly assign proteins to the path they utilize. The learned model identified new pathways and their putative carriers and motifs and these may represent novel protein sorting mechanisms. Supplementary results and software implementation are available from http://murphylab.web.cmu.edu/software/2010_RECOMB_pathways/.

VisANT 3.0: new modules for pathway visualization, editing, prediction and construction.

PubMed

Hu, Zhenjun; Ng, David M; Yamada, Takuji; Chen, Chunnuan; Kawashima, Shuichi; Mellor, Joe; Linghu, Bolan; Kanehisa, Minoru; Stuart, Joshua M; DeLisi, Charles

2007-07-01

With the integration of the KEGG and Predictome databases as well as two search engines for coexpressed genes/proteins using data sets obtained from the Stanford Microarray Database (SMD) and Gene Expression Omnibus (GEO) database, VisANT 3.0 supports exploratory pathway analysis, which includes multi-scale visualization of multiple pathways, editing and annotating pathways using a KEGG compatible visual notation and visualization of expression data in the context of pathways. Expression levels are represented either by color intensity or by nodes with an embedded expression profile. Multiple experiments can be navigated or animated. Known KEGG pathways can be enriched by querying either coexpressed components of known pathway members or proteins with known physical interactions. Predicted pathways for genes/proteins with unknown functions can be inferred from coexpression or physical interaction data. Pathways produced in VisANT can be saved as computer-readable XML format (VisML), graphic images or high-resolution Scalable Vector Graphics (SVG). Pathways in the format of VisML can be securely shared within an interested group or published online using a simple Web link. VisANT is freely available at http://visant.bu.edu.

Genome-wide gene–environment interaction analysis for asbestos exposure in lung cancer susceptibility

PubMed Central

Wei, Qingyi Wei

2012-01-01

Asbestos exposure is a known risk factor for lung cancer. Although recent genome-wide association studies (GWASs) have identified some novel loci for lung cancer risk, few addressed genome-wide gene–environment interactions. To determine gene–asbestos interactions in lung cancer risk, we conducted genome-wide gene–environment interaction analyses at levels of single nucleotide polymorphisms (SNPs), genes and pathways, using our published Texas lung cancer GWAS dataset. This dataset included 317 498 SNPs from 1154 lung cancer cases and 1137 cancer-free controls. The initial SNP-level P-values for interactions between genetic variants and self-reported asbestos exposure were estimated by unconditional logistic regression models with adjustment for age, sex, smoking status and pack-years. The P-value for the most significant SNP rs13383928 was 2.17×10–6, which did not reach the genome-wide statistical significance. Using a versatile gene-based test approach, we found that the top significant gene was C7orf54, located on 7q32.1 (P = 8.90×10–5). Interestingly, most of the other significant genes were located on 11q13. When we used an improved gene-set-enrichment analysis approach, we found that the Fas signaling pathway and the antigen processing and presentation pathway were most significant (nominal P < 0.001; false discovery rate < 0.05) among 250 pathways containing 17 572 genes. We believe that our analysis is a pilot study that first describes the gene–asbestos interaction in lung cancer risk at levels of SNPs, genes and pathways. Our findings suggest that immune function regulation-related pathways may be mechanistically involved in asbestos-associated lung cancer risk. Abbreviations:CIconfidence intervalEenvironmentFDRfalse discovery rateGgeneGSEAgene-set-enrichment analysisGWASgenome-wide association studiesi-GSEAimproved gene-set-enrichment analysis approachORodds ratioSNPsingle nucleotide polymorphism PMID:22637743

The Role of Herbivory in Structuring Tropical Seagrass Ecosystem Service Delivery

PubMed Central

Scott, Abigail L.; York, Paul H.; Duncan, Clare; Macreadie, Peter I.; Connolly, Rod M.; Ellis, Megan T.; Jarvis, Jessie C.; Jinks, Kristin I.; Marsh, Helene; Rasheed, Michael A.

2018-01-01

Seagrass meadows support key ecosystem services, via provision of food directly for herbivores, and indirectly to their predators. The importance of herbivores in seagrass meadows has been well-documented, but the links between food webs and ecosystem services in seagrass meadows have not previously been made explicit. Herbivores interact with ecosystem services – including carbon sequestration, cultural values, and coastal protection. Interactions can be positive or negative and depend on a range of factors including the herbivore identity and the grazing type and intensity. There can be unintended consequences from management actions based on a poor understanding of trade-offs that occur with complex seagrass-herbivore interactions. Tropical seagrass meadows support a diversity of grazers spanning the meso-, macro-, and megaherbivore scales. We present a conceptual model to describe how multiple ecosystem services are influenced by herbivore pressure in tropical seagrass meadows. Our model suggests that a balanced ecosystem, incorporating both seagrass and herbivore diversity, is likely to sustain the broadest range of ecosystem services. Our framework suggests the pathway to achieve desired ecosystem services outcomes requires knowledge on four key areas: (1) how size classes of herbivores interact to structure seagrass; (2) desired community and management values; (3) seagrass responses to top–down and bottom–up controls; (4) the pathway from intermediate to final ecosystem services and human benefits. We suggest research should be directed to these areas. Herbivory is a major structuring influence in tropical seagrass systems and needs to be considered for effective management of these critical habitats and their services. PMID:29487606

Cortical and subcortical interactions during action reprogramming and their related white matter pathways

PubMed Central

Neubert, Franz-Xaver; Mars, Rogier B.; Buch, Ethan R.; Olivier, Etienne; Rushworth, Matthew F. S.

2010-01-01

The right inferior frontal gyrus (rIFG) and the presupplementary motor area (pre-SMA) have been identified with cognitive control—the top-down influence on other brain areas when nonroutine behavior is required. It has been argued that they “inhibit” habitual motor responses when environmental changes mean a different response should be made. However, whether such “inhibition” can be equated with inhibitory physiological interactions has been unclear, as has the areas’ relationship with each other and the anatomical routes by which they influence movement execution. Paired-pulse transcranial magnetic stimulation (ppTMS) was applied over rIFG and primary motor cortex (M1) or over pre-SMA and M1 to measure their interactions, at a subsecond scale, during either inhibition and reprogramming of actions or during routine action selection. Distinct patterns of functional interaction between pre-SMA and M1 and between rIFG and M1 were found that were specific to action reprogramming trials; at a physiological level, direct influences of pre-SMA and rIFG on M1 were predominantly facilitatory and inhibitory, respectively. In a subsequent experiment, it was shown that the rIFG's inhibitory influence was dependent on pre-SMA. A third experiment showed that pre-SMA and rIFG influenced M1 at two time scales. By regressing white matter fractional anisotropy from diffusion-weighted magnetic resonance images against TMS-measured functional connectivity, it was shown that short-latency (6 ms) and longer latency (12 ms) influences were mediated by cortico-cortical and subcortical pathways, respectively, with the latter passing close to the subthalamic nucleus. PMID:20622155

Cortical and subcortical interactions during action reprogramming and their related white matter pathways.

PubMed

Neubert, Franz-Xaver; Mars, Rogier B; Buch, Ethan R; Olivier, Etienne; Rushworth, Matthew F S

2010-07-27

The right inferior frontal gyrus (rIFG) and the presupplementary motor area (pre-SMA) have been identified with cognitive control-the top-down influence on other brain areas when nonroutine behavior is required. It has been argued that they "inhibit" habitual motor responses when environmental changes mean a different response should be made. However, whether such "inhibition" can be equated with inhibitory physiological interactions has been unclear, as has the areas' relationship with each other and the anatomical routes by which they influence movement execution. Paired-pulse transcranial magnetic stimulation (ppTMS) was applied over rIFG and primary motor cortex (M1) or over pre-SMA and M1 to measure their interactions, at a subsecond scale, during either inhibition and reprogramming of actions or during routine action selection. Distinct patterns of functional interaction between pre-SMA and M1 and between rIFG and M1 were found that were specific to action reprogramming trials; at a physiological level, direct influences of pre-SMA and rIFG on M1 were predominantly facilitatory and inhibitory, respectively. In a subsequent experiment, it was shown that the rIFG's inhibitory influence was dependent on pre-SMA. A third experiment showed that pre-SMA and rIFG influenced M1 at two time scales. By regressing white matter fractional anisotropy from diffusion-weighted magnetic resonance images against TMS-measured functional connectivity, it was shown that short-latency (6 ms) and longer latency (12 ms) influences were mediated by cortico-cortical and subcortical pathways, respectively, with the latter passing close to the subthalamic nucleus.