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

PubMed

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

2016-06-01

Conventionally, workflows examining transcription regulation networks from gene expression data involve distinct analytical steps. There is a need for pipelines that unify data mining and inference deduction into a singular framework to enhance interpretation and hypotheses generation. We propose a workflow that merges network construction with gene expression data mining focusing on regulation processes in the context of transcription factor driven gene regulation. The pipeline implements pathway-based modularization of expression profiles into functional units to improve biological interpretation. The integrated workflow was implemented as a web application software (TransReguloNet) with functions that enable pathway visualization and comparison of transcription factor activity between sample conditions defined in the experimental design. The pipeline merges differential expression, network construction, pathway-based abstraction, clustering and visualization. The framework was applied in analysis of actual expression datasets related to lung, breast and prostrate cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

Amino Acids Regulate mTORC1 by an Obligate Two-step Mechanism*

PubMed Central

Dyachok, Julia; Earnest, Svetlana; Iturraran, Erica N.; Cobb, Melanie H.

2016-01-01

The mechanistic target of rapamycin complex 1 (mTORC1) coordinates cell growth with its nutritional, hormonal, energy, and stress status. Amino acids are critical regulators of mTORC1 that permit other inputs to mTORC1 activity. However, the roles of individual amino acids and their interactions in mTORC1 activation are not well understood. Here we demonstrate that activation of mTORC1 by amino acids includes two discrete and separable steps: priming and activation. Sensitizing mTORC1 activation by priming amino acids is a prerequisite for subsequent stimulation of mTORC1 by activating amino acids. Priming is achieved by a group of amino acids that includes l-asparagine, l-glutamine, l-threonine, l-arginine, l-glycine, l-proline, l-serine, l-alanine, and l-glutamic acid. The group of activating amino acids is dominated by l-leucine but also includes l-methionine, l-isoleucine, and l-valine. l-Cysteine predominantly inhibits priming but not the activating step. Priming and activating steps differ in their requirements for amino acid concentration and duration of treatment. Priming and activating amino acids use mechanisms that are distinct both from each other and from growth factor signaling. Neither step requires intact tuberous sclerosis complex of proteins to activate mTORC1. Concerted action of priming and activating amino acids is required to localize mTORC1 to lysosomes and achieve its activation. PMID:27587390

Prokaryotic Heme Biosynthesis: Multiple Pathways to a Common Essential Product

PubMed Central

Dailey, Tamara A.; Gerdes, Svetlana; Jahn, Dieter; O'Brian, Mark R.; Warren, Martin J.

2017-01-01

SUMMARY The advent of heme during evolution allowed organisms possessing this compound to safely and efficiently carry out a variety of chemical reactions that otherwise were difficult or impossible. While it was long assumed that a single heme biosynthetic pathway existed in nature, over the past decade, it has become clear that there are three distinct pathways among prokaryotes, although all three pathways utilize a common initial core of three enzymes to produce the intermediate uroporphyrinogen III. The most ancient pathway and the only one found in the Archaea converts siroheme to protoheme via an oxygen-independent four-enzyme-step process. Bacteria utilize the initial core pathway but then add one additional common step to produce coproporphyrinogen III. Following this step, Gram-positive organisms oxidize coproporphyrinogen III to coproporphyrin III, insert iron to make coproheme, and finally decarboxylate coproheme to protoheme, whereas Gram-negative bacteria first decarboxylate coproporphyrinogen III to protoporphyrinogen IX and then oxidize this to protoporphyrin IX prior to metal insertion to make protoheme. In order to adapt to oxygen-deficient conditions, two steps in the bacterial pathways have multiple forms to accommodate oxidative reactions in an anaerobic environment. The regulation of these pathways reflects the diversity of bacterial metabolism. This diversity, along with the late recognition that three pathways exist, has significantly slowed advances in this field such that no single organism's heme synthesis pathway regulation is currently completely characterized. PMID:28123057

Medicine and law as model professions: the heart of the matter (and how we have missed it).

PubMed

Atkinson, Rob

2013-01-01

This article has two coordinate goals: to undergird the functionalist understanding of professionalism with classical normative theory and to advance the classical theory of civic virtue with the insights of modern social science. More specifically, this article seeks to connect classical theories about the care of the body and the soul with modern theories of market and government failure. The first step is to distinguish two kinds of professions, caring professions like medicine and public professions like law, by identifying the distinctive virtue of each. The distinctive virtue of the caring professions is single-minded commitment to those in their care, their principals, to the virtual exclusion of all other concerns; the distinctive virtue of the public professions is commitment to the common good, sometimes even at the expense of their principals' self-defined interest. The next step is to show how these two distinctive professional virtues, the one principal-protecting, the other public-protecting, branch from the same root, the common function of all proper professions: guaranteeing the delivery of socially essential but necessarily esoteric knowledge when the usual protections of both private contracts and government regulation systematically fail. The third and final step is to map out the implications of this neo-classical understanding of professionalism, beginning at its core in the paradigmatic caring and public professions of medicine and law, through putative professions that take these as their models, to the kind of republican society that places care of individuals and concern for the public welfare at the center of its value system. The result of this analysis should be not only a fuller theoretical appreciation of professionalism's proper function, but also a practical guide to professionals themselves for better service to both the individuals in their care and the common good of all humankind.

The Rice Basic Helix-Loop-Helix Transcription Factor TDR INTERACTING PROTEIN2 Is a Central Switch in Early Anther Development[C][W

PubMed Central

Fu, Zhenzhen; Yu, Jing; Cheng, Xiaowei; Zong, Xu; Xu, Jie; Chen, Mingjiao; Li, Zongyun; Zhang, Dabing; Liang, Wanqi

2014-01-01

In male reproductive development in plants, meristemoid precursor cells possessing transient, stem cell–like features undergo cell divisions and differentiation to produce the anther, the male reproductive organ. The anther contains centrally positioned microsporocytes surrounded by four distinct layers of wall: the epidermis, endothecium, middle layer, and tapetum. Here, we report that the rice (Oryza sativa) basic helix-loop-helix (bHLH) protein TDR INTERACTING PROTEIN2 (TIP2) functions as a crucial switch in the meristemoid transition and differentiation during early anther development. The tip2 mutants display undifferentiated inner three anther wall layers and abort tapetal programmed cell death, causing complete male sterility. TIP2 has two paralogs in rice, TDR and EAT1, which are key regulators of tapetal programmed cell death. We revealed that TIP2 acts upstream of TDR and EAT1 and directly regulates the expression of TDR and EAT1. In addition, TIP2 can interact with TDR, indicating a role of TIP2 in later anther development. Our findings suggest that the bHLH proteins TIP2, TDR, and EAT1 play a central role in regulating differentiation, morphogenesis, and degradation of anther somatic cell layers, highlighting the role of paralogous bHLH proteins in regulating distinct steps of plant cell–type determination. PMID:24755456

Imaging Transcriptional Regulation of Eukaryotic mRNA Genes: Advances and Outlook.

PubMed

Yao, Jie

2017-01-06

Regulation of eukaryotic transcription in vivo occurs at distinct stages. Previous research has identified many active or repressive transcription factors (TFs) and core transcription components and studied their functions in vitro and in vivo. Nonetheless, how individual TFs act in concert to regulate mRNA gene expression in a single cell remains poorly understood. Direct observation of TF assembly and disassembly and various biochemical reactions during transcription of a single-copy gene in vivo is the ideal approach to study this problem. Research in this area requires developing novel techniques for single-cell transcription imaging and integrating imaging studies into understanding the molecular biology of transcription. In the past decade, advanced cell imaging has enabled unprecedented capabilities to visualize individual TF molecules, to track single transcription sites, and to detect individual mRNA in fixed and living cells. These studies have raised several novel insights on transcriptional regulation such as the "hit-and-run" model and transcription bursting that could not be obtained by in vitro biochemistry analysis. At this point, the key question is how to achieve deeper understandings or discover novel mechanisms of eukaryotic transcriptional regulation by imaging transcription in single cells. Meanwhile, further technical advancements are likely required for visualizing distinct kinetic steps of transcription on a single-copy gene in vivo. This review article summarizes recent progress in the field and describes the challenges and opportunities ahead. Copyright © 2016 Elsevier Ltd. All rights reserved.

Modular control of glutamatergic neuronal identity in C.elegans by distinct homeodomain proteins

PubMed Central

Serrano-Saiz, Esther; Poole, Richard J.; Felton, Terry; Zhang, Feifan; De La Cruz, Estanisla Daniel; Hobert, Oliver

2013-01-01

The choice of using one of many possible neurotransmitter systems is a critical step in defining the identity of an individual neuron type. We show here that the key defining feature of glutamatergic neurons, the vesicular glutamate transporter EAT-4/VGLUT is expressed in 38 of the 118 anatomically defined neuron classes of the C.elegans nervous system. We show that eat-4/VGLUT expression is controlled in a modular manner, with distinct cis-regulatory modules driving expression in distinct glutamatergic neuron classes. We identify 13 different transcription factors, 11 of them homeodomain proteins, that act in specific combinations in 25 different glutamatergic neuron classes to initiate and maintain eat-4/VGLUT expression. We show that the adoption of a glutamatergic phenotype is linked to the adoption of other terminal identity features of a neuron, including cotransmitter phenotypes. Examination of mouse orthologs of these homeodomain proteins resulted in the identification of mouse LHX1 as a regulator of glutamatergic neurons in the brainstem. PMID:24243022

Uncovering Hidden Layers of Cell Cycle Regulation through Integrative Multi-omic Analysis

PubMed Central

Aviner, Ranen; Shenoy, Anjana; Elroy-Stein, Orna; Geiger, Tamar

2015-01-01

Studying the complex relationship between transcription, translation and protein degradation is essential to our understanding of biological processes in health and disease. The limited correlations observed between mRNA and protein abundance suggest pervasive regulation of post-transcriptional steps and support the importance of profiling mRNA levels in parallel to protein synthesis and degradation rates. In this work, we applied an integrative multi-omic approach to study gene expression along the mammalian cell cycle through side-by-side analysis of mRNA, translation and protein levels. Our analysis sheds new light on the significant contribution of both protein synthesis and degradation to the variance in protein expression. Furthermore, we find that translation regulation plays an important role at S-phase, while progression through mitosis is predominantly controlled by changes in either mRNA levels or protein stability. Specific molecular functions are found to be co-regulated and share similar patterns of mRNA, translation and protein expression along the cell cycle. Notably, these include genes and entire pathways not previously implicated in cell cycle progression, demonstrating the potential of this approach to identify novel regulatory mechanisms beyond those revealed by traditional expression profiling. Through this three-level analysis, we characterize different mechanisms of gene expression, discover new cycling gene products and highlight the importance and utility of combining datasets generated using different techniques that monitor distinct steps of gene expression. PMID:26439921

Metabolic Genetic Screens Reveal Multidimensional Regulation of Virulence Gene Expression in Listeria monocytogenes and an Aminopeptidase That Is Critical for PrfA Protein Activation.

PubMed

Friedman, Sivan; Linsky, Marika; Lobel, Lior; Rabinovich, Lev; Sigal, Nadejda; Herskovits, Anat A

2017-06-01

Listeria monocytogenes is an environmental saprophyte and intracellular bacterial pathogen. Upon invading mammalian cells, the bacterium senses abrupt changes in its metabolic environment, which are rapidly transduced to regulation of virulence gene expression. To explore the relationship between L. monocytogenes metabolism and virulence, we monitored virulence gene expression dynamics across a library of genetic mutants grown under two metabolic conditions known to activate the virulent state: charcoal-treated rich medium containing glucose-1-phosphate and minimal defined medium containing limiting concentrations of branched-chain amino acids (BCAAs). We identified over 100 distinct mutants that exhibit aberrant virulence gene expression profiles, the majority of which mapped to nonessential metabolic genes. Mutants displayed enhanced, decreased, and early and late virulence gene expression profiles, as well as persistent levels, demonstrating a high plasticity in virulence gene regulation. Among the mutants, one was noteworthy for its particularly low virulence gene expression level and mapped to an X-prolyl aminopeptidase (PepP). We show that this peptidase plays a role in posttranslational activation of the major virulence regulator, PrfA. Specifically, PepP mediates recruitment of PrfA to the cytoplasmic membrane, a step identified as critical for PrfA protein activation. This study establishes a novel step in the complex mechanism of PrfA activation and further highlights the cross regulation of metabolism and virulence. Copyright © 2017 American Society for Microbiology.

Mechanisms of ribosome stalling by SecM at multiple elongation steps

PubMed Central

Zhang, Jun; Pan, Xijiang; Yan, Kaige; Sun, Shan; Gao, Ning; Sui, Sen-Fang

2015-01-01

Regulation of translating ribosomes is a major component of gene expression control network. In Escherichia coli, ribosome stalling by the C-terminal arrest sequence of SecM regulates the SecA-dependent secretion pathway. Previous studies reported many residues of SecM peptide and ribosome exit tunnel are critical for stalling. However, the underlying molecular mechanism is still not clear at the atomic level. Here, we present two cryo-EM structures of the SecM-stalled ribosomes at 3.3–3.7 Å resolution, which reveal two different stalling mechanisms at distinct elongation steps of the translation cycle: one is due to the inactivation of ribosomal peptidyl-transferase center which inhibits peptide bond formation with the incoming prolyl-tRNA; the other is the prolonged residence of the peptidyl-RNA at the hybrid A/P site which inhibits the full-scale tRNA translocation. These results demonstrate an elegant control of translation cycle by regulatory peptides through a continuous, dynamic reshaping of the functional center of the ribosome. DOI: http://dx.doi.org/10.7554/eLife.09684.001 PMID:26670735

Preparation of clinker from paper pulp industry wastes.

PubMed

Buruberri, Leire H; Seabra, M P; Labrincha, J A

2015-04-09

The production of paper pulp by the Kraft method generates considerable amounts of wastes. Namely, lime mud generated in the recovery circuit of chemical reagents, biological sludge from the wastewater treatment of wood digestion process and fly ash collected in the fluidized bed combustor used to generate electricity from biomass burning. The final destination of such wastes is an important concern, since environmental regulations are becoming stricter regarding their landfill. Driven by this fact, industries are looking for more sustainable solutions, such as the recycling in distinct products. This work tested these wastes as secondary raw materials to produce clinker/cement that was then experienced in mortar formulations. The first step involved the residues detailed characterization and a generated amounts survey. Then, specific but simple steps were suggested, aiming to facilitate transport and manipulation. Distinct blends were prepared and fired in order to get belitic and Portland clinkers. The Portland clinkers were processed at lower temperatures than the normally used in the industry due to the presence of mineralizing impurities in some wastes. Belite-based cements were used to produce mortars that developed satisfactory mechanical strength and did not reveal signs of deterioration or durability weaknesses. Copyright © 2014 Elsevier B.V. All rights reserved.

Phosphoproteomics in bacteria: towards a systemic understanding of bacterial phosphorylation networks.

PubMed

Jers, Carsten; Soufi, Boumediene; Grangeasse, Christophe; Deutscher, Josef; Mijakovic, Ivan

2008-08-01

Bacteria use protein phosphorylation to regulate all kinds of physiological processes. Protein phosphorylation plays a role in several key steps of the infection process of bacterial pathogens, such as adhesion to the host, triggering and regulation of pathogenic functions as well as biochemical warfare; scrambling the host signaling cascades and impairing its defense mechanisms. Recent phosphoproteomic studies indicate that the bacterial protein phosphorylation networks could be more complex than initially expected, comprising promiscuous kinases that regulate several distinct cellular functions by phosphorylating different protein substrates. Recent advances in protein labeling with stable isotopes in the field of quantitative mass spectrometry phosphoproteomics will enable us to chart the global phosphorylation networks and to understand the implication of protein phosphorylation in cellular regulation on the systems scale. For the study of bacterial pathogens, in particular, this research avenue will enable us to dissect phosphorylation-related events during different stages of infection and stimulate our efforts to find inhibitors for key kinases and phosphatases implicated therein.

GABA-CREB signalling regulates maturation and survival of newly generated neurons in the adult hippocampus

PubMed Central

Jagasia, Ravi; Steib, Kathrin; Englberger, Elisabeth; Herold, Sabine; Faus-Kessler, Theresa; Saxe, Michael; Gage, Fred H.; Song, Hongjun; Lie, D. Chichung

2009-01-01

Survival and integration of new neurons in the hippocampal circuit are rate-limiting steps in adult hippocampal neurogenesis. Neuronal network activity is a major regulator of these processes, yet little is known about the respective downstream signalling pathways. Here, we investigate the role of CREB signalling in adult hippocampal neurogenesis. CREB is activated in new granule neurons during a distinct developmental period. Loss of CREB function in a cell-autonomous fashion impairs dendritic development, decreases the expression of the neurogenic transcription factor NeuroD and of the neuronal microtubule associated protein, DCX, and compromises the survival of newborn neurons. In addition, GABA-mediated excitation regulates CREB activation at early developmental stages. Importantly, developmental defects following loss of GABA-mediated excitation can be compensated by enhanced CREB signalling. These results indicate that CREB signalling is a central pathway in adult hippocampal neurogenesis, regulating the development and survival of new hippocampal neurons downstream of GABA-mediated excitation. PMID:19553437

Regulation of chromatin organization and inducible gene expression by a Drosophila insulator

PubMed Central

Wood, Ashley M.; Van Bortle, Kevin; Ramos, Edward; Takenaka, Naomi; Rohrbaugh, Margaret; Jones, Brian C.; Jones, Keith C.; Corces, Victor G.

2011-01-01

SUMMARY Insulators are multi-protein-DNA complexes thought to affect gene expression by mediating inter- and intra-chromosomal interactions. Drosophila insulators contain specific DNA binding proteins plus common components, such as CP190, that facilitate these interactions. Here we examine changes in the distribution of Drosophila insulator proteins during the heat-shock and ecdysone responses. We find that CP190 recruitment to insulator sites is the main regulatable step in controlling insulator function during heat shock. In contrast, both CP190 and DNA binding protein recruitment are regulated during the ecdysone response. CP190 is necessary to stabilize specific chromatin loops and for proper activation of transcription of genes regulated by this hormone. These findings suggest that cells may regulate recruitment of insulator proteins to the DNA in order to activate insulator activity at specific sites and create distinct patterns of nuclear organization that are necessary to achieve proper gene expression in response to different stimuli. PMID:21981916

Regulation of chromatin organization and inducible gene expression by a Drosophila insulator.

PubMed

Wood, Ashley M; Van Bortle, Kevin; Ramos, Edward; Takenaka, Naomi; Rohrbaugh, Margaret; Jones, Brian C; Jones, Keith C; Corces, Victor G

2011-10-07

Insulators are multiprotein-DNA complexes thought to affect gene expression by mediating inter- and intrachromosomal interactions. Drosophila insulators contain specific DNA-binding proteins plus common components, such as CP190, that facilitate these interactions. Here, we examine changes in the distribution of Drosophila insulator proteins during the heat-shock and ecdysone responses. We find that CP190 recruitment to insulator sites is the main regulatable step in controlling insulator function during heat shock. In contrast, both CP190 and DNA-binding protein recruitment are regulated during the ecdysone response. CP190 is necessary to stabilize specific chromatin loops and for proper activation of transcription of genes regulated by this hormone. These findings suggest that cells may regulate recruitment of insulator proteins to DNA to activate insulator activity at specific sites and create distinct patterns of nuclear organization that are necessary to achieve proper gene expression in response to different stimuli. Copyright © 2011 Elsevier Inc. All rights reserved.

The Cryptococcus neoformans Capsule: a Sword and a Shield

PubMed Central

O'Meara, Teresa R.

2012-01-01

Summary: The human fungal pathogen Cryptococcus neoformans is characterized by its ability to induce a distinct polysaccharide capsule in response to a number of host-specific environmental stimuli. The induction of capsule is a complex biological process encompassing regulation at multiple steps, including the biosynthesis, transport, and maintenance of the polysaccharide at the cell surface. By precisely regulating the composition of its cell surface and secreted polysaccharides, C. neoformans has developed intricate ways to establish chronic infection and dormancy in the human host. The plasticity of the capsule structure in response to various host conditions also underscores the complex relationship between host and parasite. Much of this precise regulation of capsule is achieved through the transcriptional responses of multiple conserved signaling pathways that have been coopted to regulate this C. neoformans-specific virulence-associated phenotype. This review focuses on specific host stimuli that trigger the activation of the signal transduction cascades and on the downstream transcriptional responses that are required for robust encapsulation around the cell. PMID:22763631

N-terminus of Cardiac Myosin Essential Light Chain Modulates Myosin Step-Size

PubMed Central

Wang, Yihua; Ajtai, Katalin; Kazmierczak, Katarzyna; Szczesna-Cordary, Danuta; Burghardt, Thomas P.

2016-01-01

Muscle myosin cyclically hydrolyzes ATP to translate actin. Ventricular cardiac myosin (βmys) moves actin with three distinct unitary step-sizes resulting from its lever-arm rotation and with step-frequencies that are modulated in a myosin regulation mechanism. The lever-arm associated essential light chain (vELC) binds actin by its 43 residue N-terminal extension. Unitary steps were proposed to involve the vELC N-terminal extension with the 8 nm step engaging the vELC/actin bond facilitating an extra ~19 degrees of lever-arm rotation while the predominant 5 nm step forgoes vELC/actin binding. A minor 3 nm step is the unlikely conversion of the completed 5 to the 8 nm step. This hypothesis was tested using a 17 residue N-terminal truncated vELC in porcine βmys (Δ17βmys) and a 43 residue N-terminal truncated human vELC expressed in transgenic mouse heart (Δ43αmys). Step-size and step-frequency were measured using the Qdot motility assay. Both Δ17βmys and Δ43αmys had significantly increased 5 nm step-frequency and coincident loss in the 8 nm step-frequency compared to native proteins suggesting the vELC/actin interaction drives step-size preference. Step-size and step-frequency probability densities depend on the relative fraction of truncated vELC and relate linearly to pure myosin species concentrations in a mixture containing native vELC homodimer, two truncated vELCs in the modified homodimer, and one native and one truncated vELC in the heterodimer. Step-size and step-frequency, measured for native homodimer and at two or more known relative fractions of truncated vELC, are surmised for each pure species by using a new analytical method. PMID:26671638

43 CFR 17.314 - Age distinctions contained in DOI regulations.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 43 Public Lands: Interior 1 2011-10-01 2011-10-01 false Age distinctions contained in DOI... Age Standards for Determining Age Discrimination § 17.314 Age distinctions contained in DOI regulations. Any age distinctions contained in a rule or regulation issued by DOI shall be presumed to be...

43 CFR 17.314 - Age distinctions contained in DOI regulations.

Code of Federal Regulations, 2013 CFR

2013-10-01

... 43 Public Lands: Interior 1 2013-10-01 2013-10-01 false Age distinctions contained in DOI... Age Standards for Determining Age Discrimination § 17.314 Age distinctions contained in DOI regulations. Any age distinctions contained in a rule or regulation issued by DOI shall be presumed to be...

43 CFR 17.314 - Age distinctions contained in DOI regulations.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 43 Public Lands: Interior 1 2014-10-01 2014-10-01 false Age distinctions contained in DOI... Age Standards for Determining Age Discrimination § 17.314 Age distinctions contained in DOI regulations. Any age distinctions contained in a rule or regulation issued by DOI shall be presumed to be...

43 CFR 17.314 - Age distinctions contained in DOI regulations.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 43 Public Lands: Interior 1 2012-10-01 2011-10-01 true Age distinctions contained in DOI... Age Standards for Determining Age Discrimination § 17.314 Age distinctions contained in DOI regulations. Any age distinctions contained in a rule or regulation issued by DOI shall be presumed to be...

43 CFR 17.314 - Age distinctions contained in DOI regulations.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Age distinctions contained in DOI... Age Standards for Determining Age Discrimination § 17.314 Age distinctions contained in DOI regulations. Any age distinctions contained in a rule or regulation issued by DOI shall be presumed to be...

Swing-leg trajectory of running guinea fowl suggests task-level priority of force regulation rather than disturbance rejection.

PubMed

Blum, Yvonne; Vejdani, Hamid R; Birn-Jeffery, Aleksandra V; Hubicki, Christian M; Hurst, Jonathan W; Daley, Monica A

2014-01-01

To achieve robust and stable legged locomotion in uneven terrain, animals must effectively coordinate limb swing and stance phases, which involve distinct yet coupled dynamics. Recent theoretical studies have highlighted the critical influence of swing-leg trajectory on stability, disturbance rejection, leg loading and economy of walking and running. Yet, simulations suggest that not all these factors can be simultaneously optimized. A potential trade-off arises between the optimal swing-leg trajectory for disturbance rejection (to maintain steady gait) versus regulation of leg loading (for injury avoidance and economy). Here we investigate how running guinea fowl manage this potential trade-off by comparing experimental data to predictions of hypothesis-based simulations of running over a terrain drop perturbation. We use a simple model to predict swing-leg trajectory and running dynamics. In simulations, we generate optimized swing-leg trajectories based upon specific hypotheses for task-level control priorities. We optimized swing trajectories to achieve i) constant peak force, ii) constant axial impulse, or iii) perfect disturbance rejection (steady gait) in the stance following a terrain drop. We compare simulation predictions to experimental data on guinea fowl running over a visible step down. Swing and stance dynamics of running guinea fowl closely match simulations optimized to regulate leg loading (priorities i and ii), and do not match the simulations optimized for disturbance rejection (priority iii). The simulations reinforce previous findings that swing-leg trajectory targeting disturbance rejection demands large increases in stance leg force following a terrain drop. Guinea fowl negotiate a downward step using unsteady dynamics with forward acceleration, and recover to steady gait in subsequent steps. Our results suggest that guinea fowl use swing-leg trajectory consistent with priority for load regulation, and not for steadiness of gait. Swing-leg trajectory optimized for load regulation may facilitate economy and injury avoidance in uneven terrain.

Swing-Leg Trajectory of Running Guinea Fowl Suggests Task-Level Priority of Force Regulation Rather than Disturbance Rejection

PubMed Central

Blum, Yvonne; Vejdani, Hamid R.; Birn-Jeffery, Aleksandra V.; Hubicki, Christian M.; Hurst, Jonathan W.; Daley, Monica A.

2014-01-01

To achieve robust and stable legged locomotion in uneven terrain, animals must effectively coordinate limb swing and stance phases, which involve distinct yet coupled dynamics. Recent theoretical studies have highlighted the critical influence of swing-leg trajectory on stability, disturbance rejection, leg loading and economy of walking and running. Yet, simulations suggest that not all these factors can be simultaneously optimized. A potential trade-off arises between the optimal swing-leg trajectory for disturbance rejection (to maintain steady gait) versus regulation of leg loading (for injury avoidance and economy). Here we investigate how running guinea fowl manage this potential trade-off by comparing experimental data to predictions of hypothesis-based simulations of running over a terrain drop perturbation. We use a simple model to predict swing-leg trajectory and running dynamics. In simulations, we generate optimized swing-leg trajectories based upon specific hypotheses for task-level control priorities. We optimized swing trajectories to achieve i) constant peak force, ii) constant axial impulse, or iii) perfect disturbance rejection (steady gait) in the stance following a terrain drop. We compare simulation predictions to experimental data on guinea fowl running over a visible step down. Swing and stance dynamics of running guinea fowl closely match simulations optimized to regulate leg loading (priorities i and ii), and do not match the simulations optimized for disturbance rejection (priority iii). The simulations reinforce previous findings that swing-leg trajectory targeting disturbance rejection demands large increases in stance leg force following a terrain drop. Guinea fowl negotiate a downward step using unsteady dynamics with forward acceleration, and recover to steady gait in subsequent steps. Our results suggest that guinea fowl use swing-leg trajectory consistent with priority for load regulation, and not for steadiness of gait. Swing-leg trajectory optimized for load regulation may facilitate economy and injury avoidance in uneven terrain. PMID:24979750

BLOC-1 and BLOC-3 regulate VAMP7 cycling to and from melanosomes via distinct tubular transport carriers

PubMed Central

Delevoye, Cédric; Romao, Maryse; Owen, David J.; Raposo, Graça

2016-01-01

Endomembrane organelle maturation requires cargo delivery via fusion with membrane transport intermediates and recycling of fusion factors to their sites of origin. Melanosomes and other lysosome-related organelles obtain cargoes from early endosomes, but the fusion machinery involved and its recycling pathway are unknown. Here, we show that the v-SNARE VAMP7 mediates fusion of melanosomes with tubular transport carriers that also carry the cargo protein TYRP1 and that require BLOC-1 for their formation. Using live-cell imaging, we identify a pathway for VAMP7 recycling from melanosomes that employs distinct tubular carriers. The recycling carriers also harbor the VAMP7-binding scaffold protein VARP and the tissue-restricted Rab GTPase RAB38. Recycling carrier formation is dependent on the RAB38 exchange factor BLOC-3. Our data suggest that VAMP7 mediates fusion of BLOC-1–dependent transport carriers with melanosomes, illuminate SNARE recycling from melanosomes as a critical BLOC-3–dependent step, and likely explain the distinct hypopigmentation phenotypes associated with BLOC-1 and BLOC-3 deficiency in Hermansky–Pudlak syndrome variants. PMID:27482051

41 CFR 101-8.710 - Age distinctions contained in General Services Administration regulation.

Code of Federal Regulations, 2010 CFR

2010-07-01

... contained in General Services Administration regulation. 101-8.710 Section 101-8.710 Public Contracts and... Prohibited on the Basis of Age § 101-8.710 Age distinctions contained in General Services Administration regulation. Any age distinctions contained in a rule or regulation issued by GSA are presumed to be necessary...

Specialized sorting of GLUT4 and its recruitment to the cell surface are independently regulated by distinct Rabs

PubMed Central

Sadacca, L. Amanda; Bruno, Joanne; Wen, Jennifer; Xiong, Wenyong; McGraw, Timothy E.

2013-01-01

Adipocyte glucose uptake in response to insulin is essential for physiological glucose homeostasis: stimulation of adipocytes with insulin results in insertion of the glucose transporter GLUT4 into the plasma membrane and subsequent glucose uptake. Here we establish that RAB10 and RAB14 are key regulators of GLUT4 trafficking that function at independent, sequential steps of GLUT4 translocation. RAB14 functions upstream of RAB10 in the sorting of GLUT4 to the specialized transport vesicles that ferry GLUT4 to the plasma membrane. RAB10 and its GTPase-activating protein (GAP) AS160 comprise the principal signaling module downstream of insulin receptor activation that regulates the accumulation of GLUT4 transport vesicles at the plasma membrane. Although both RAB10 and RAB14 are regulated by the GAP activity of AS160 in vitro, only RAB10 is under the control of AS160 in vivo. Insulin regulation of the pool of RAB10 required for GLUT4 translocation occurs through regulation of AS160, since activation of RAB10 by DENND4C, its GTP exchange factor, does not require insulin stimulation. PMID:23804653

Dynamics of DNA methylomes underlie oyster development

PubMed Central

Sourdaine, Pascal; Guo, Ximing; Favrel, Pascal

2017-01-01

DNA methylation is a critical epigenetic regulator of development in mammals and social insects, but its significance in development outside these groups is not understood. Here we investigated the genome-wide dynamics of DNA methylation in a mollusc model, the oyster Crassostrea gigas, from the egg to the completion of organogenesis. Large-scale methylation maps reveal that the oyster genome displays a succession of methylated and non methylated regions, which persist throughout development. Differentially methylated regions (DMRs) are strongly regulated during cleavage and metamorphosis. The distribution and levels of methylated DNA within genomic features (exons, introns, promoters, repeats and transposons) show different developmental lansdscapes marked by a strong increase in the methylation of exons against introns after metamorphosis. Kinetics of methylation in gene-bodies correlate to their transcription regulation and to distinct functional gene clusters, and DMRs at cleavage and metamorphosis bear the genes functionally related to these steps, respectively. This study shows that DNA methylome dynamics underlie development through transcription regulation in the oyster, a lophotrochozoan species. To our knowledge, this is the first demonstration of such epigenetic regulation outside vertebrates and ecdysozoan models, bringing new insights into the evolution and the epigenetic regulation of developmental processes. PMID:28594821

Dynamics of DNA methylomes underlie oyster development.

PubMed

Riviere, Guillaume; He, Yan; Tecchio, Samuele; Crowell, Elizabeth; Gras, Michaël; Sourdaine, Pascal; Guo, Ximing; Favrel, Pascal

2017-06-01

DNA methylation is a critical epigenetic regulator of development in mammals and social insects, but its significance in development outside these groups is not understood. Here we investigated the genome-wide dynamics of DNA methylation in a mollusc model, the oyster Crassostrea gigas, from the egg to the completion of organogenesis. Large-scale methylation maps reveal that the oyster genome displays a succession of methylated and non methylated regions, which persist throughout development. Differentially methylated regions (DMRs) are strongly regulated during cleavage and metamorphosis. The distribution and levels of methylated DNA within genomic features (exons, introns, promoters, repeats and transposons) show different developmental lansdscapes marked by a strong increase in the methylation of exons against introns after metamorphosis. Kinetics of methylation in gene-bodies correlate to their transcription regulation and to distinct functional gene clusters, and DMRs at cleavage and metamorphosis bear the genes functionally related to these steps, respectively. This study shows that DNA methylome dynamics underlie development through transcription regulation in the oyster, a lophotrochozoan species. To our knowledge, this is the first demonstration of such epigenetic regulation outside vertebrates and ecdysozoan models, bringing new insights into the evolution and the epigenetic regulation of developmental processes.

Characterization of Autophagic Responses in Drosophila melanogaster.

PubMed

Xu, T; Kumar, S; Denton, D

2017-01-01

Drosophila is an excellent model system for studying autophagy during animal development due to the availability of genetic reagents and opportunity for in vivo cell biological analysis. The regulation and mechanism of autophagy are highly evolutionarily conserved and the role of autophagy has been characterized during various stages of Drosophila development as well as following starvation. Studies in Drosophila have revealed novel insights into the role of distinct components of the autophagy machinery. This chapter describes protocols for examining autophagy during Drosophila development. A crucial step in the induction of autophagy is the incorporation of Atg8a into the autophagosome. This can be measured as autophagic puncta using live fluorescent imaging, immunostaining, or immunoblot analysis of LC3/Atg8a processing. The level of autophagy can also be examined using other specific components of the autophagy pathway as markers detected by immunofluorescent imaging. Based on the distinct morphology of autophagy, it can also be examined by transmission electron microscopy. In addition, one of the advantages of using Drosophila as a model is the ability to undertake genetic analysis of individual components of the autophagy machinery. Current approaches that can be used to monitor autophagy, including the overall flux and individual steps in Drosophila melanogaster, will be discussed. © 2017 Elsevier Inc. All rights reserved.

A Review of Current and Prospective Factors for Classification of Civil Unmanned Aircraft Systems

NASA Technical Reports Server (NTRS)

Hayhurst, Kelly J.; Maddalon, Jeffrey M.; Morris, A. Terry; Neogi, Natasha; Verstynen, Harry A.

2014-01-01

While progress is being made on integrating unmanned aircraft systems (UAS) into our national airspace on a broad scale, much work remains to establish appropriate certification standards and operational procedures, particularly with respect to routine commercial operations. This paper summarizes research to examine the extent to which today's civil aircraft taxonomy applies to UAS, and, if needed, how that taxonomy could be amended to better cover different UAS designs and operations. Factors that shape the current taxonomy, as defined in the Federal Aviation Regulations, were assessed for applicability to UAS, potential incompatibilities were identified, and additional factors were proposed that might be useful for an updated aircraft taxonomy intended to cover UAS. The results suggest the possibility of constructing new groups in the taxonomy for UAS under a restricted category that share common airworthiness standards. Establishing distinct groups for UAS and associated standards that enable low risk operations for compensation or hire could be a timely step toward full integration. Such a step would allow the civil aviation industry and regulators to gain valuable experience with UAS while carefully controlling access and potential harm to the aviation system as a whole.

The head module of Mediator directs activation of preloaded RNAPII in vivo.

PubMed

Lee, Sarah K; Chen, Xu; Huang, Liangqun; Stargell, Laurie A

2013-12-01

The successful synthesis of a transcript by RNA polymerase II (RNAPII) is a multistage process with distinct rate-limiting steps that can vary depending on the particular gene. A growing number of genes in a variety of organisms are regulated at steps after the recruitment of RNAPII. The best-characterized Saccharomyces cerevisiae gene regulated in this manner is CYC1. This gene has high occupancy of RNAPII under non-inducing conditions, defining it as a poised gene. Here, we find that subunits of the head module of Mediator, Med18 and Med20, and Med19 are required for activation of transcription at the CYC1 promoter in response to environmental cues. These subunits of Mediator are required at the preloaded promoter for normal levels of recruitment and activity of the general transcription factor TFIIH. Strikingly, these Mediator components are dispensable for activation by the same activator at a different gene, which lacks a preloaded polymerase in the promoter region. Based on these results and other studies, we speculate that Mediator plays an essential role in triggering an inactive polymerase at CYC1 into a productively elongating form.

Sequence-dependent nucleosome sliding in rotation-coupled and uncoupled modes revealed by molecular simulations

PubMed Central

Tan, Cheng; Takada, Shoji

2017-01-01

While nucleosome positioning on eukaryotic genome play important roles for genetic regulation, molecular mechanisms of nucleosome positioning and sliding along DNA are not well understood. Here we investigated thermally-activated spontaneous nucleosome sliding mechanisms developing and applying a coarse-grained molecular simulation method that incorporates both long-range electrostatic and short-range hydrogen-bond interactions between histone octamer and DNA. The simulations revealed two distinct sliding modes depending on the nucleosomal DNA sequence. A uniform DNA sequence showed frequent sliding with one base pair step in a rotation-coupled manner, akin to screw-like motions. On the contrary, a strong positioning sequence, the so-called 601 sequence, exhibits rare, abrupt transitions of five and ten base pair steps without rotation. Moreover, we evaluated the importance of hydrogen bond interactions on the sliding mode, finding that strong and weak bonds favor respectively the rotation-coupled and -uncoupled sliding movements. PMID:29194442

Single-Molecule View of Small RNA-Guided Target Search and Recognition.

PubMed

Globyte, Viktorija; Kim, Sung Hyun; Joo, Chirlmin

2018-05-20

Most everyday processes in life involve a necessity for an entity to locate its target. On a cellular level, many proteins have to find their target to perform their function. From gene-expression regulation to DNA repair to host defense, numerous nucleic acid-interacting proteins use distinct target search mechanisms. Several proteins achieve that with the help of short RNA strands known as guides. This review focuses on single-molecule advances studying the target search and recognition mechanism of Argonaute and CRISPR (clustered regularly interspaced short palindromic repeats) systems. We discuss different steps involved in search and recognition, from the initial complex prearrangement into the target-search competent state to the final proofreading steps. We focus on target search mechanisms that range from weak interactions, to one- and three-dimensional diffusion, to conformational proofreading. We compare the mechanisms of Argonaute and CRISPR with a well-studied target search system, RecA.

Real-Time Label-Free Direct Electronic Monitoring of Topoisomerase Enzyme Binding Kinetics on Graphene.

PubMed

Zuccaro, Laura; Tesauro, Cinzia; Kurkina, Tetiana; Fiorani, Paola; Yu, Hak Ki; Knudsen, Birgitta R; Kern, Klaus; Desideri, Alessandro; Balasubramanian, Kannan

2015-11-24

Monolayer graphene field-effect sensors operating in liquid have been widely deployed for detecting a range of analyte species often under equilibrium conditions. Here we report on the real-time detection of the binding kinetics of the essential human enzyme, topoisomerase I interacting with substrate molecules (DNA probes) that are immobilized electrochemically on to monolayer graphene strips. By monitoring the field-effect characteristics of the graphene biosensor in real-time during the enzyme-substrate interactions, we are able to decipher the surface binding constant for the cleavage reaction step of topoisomerase I activity in a label-free manner. Moreover, an appropriate design of the capture probes allows us to distinctly follow the cleavage step of topoisomerase I functioning in real-time down to picomolar concentrations. The presented results are promising for future rapid screening of drugs that are being evaluated for regulating enzyme activity.

Endosidin2 targets conserved exocyst complex subunit EXO70 to inhibit exocytosis

DOE PAGES

Zhang, Chunhua; Brown, Michelle Q.; van de Ven, Wilhelmina; ...

2015-11-25

The exocyst complex regulates the last steps of exocytosis, which is essential to organisms across kingdoms. In humans, its dysfunction is correlated with several significant diseases, such as diabetes and cancer progression. Investigation of the dynamic regulation of the evolutionarily conserved exocyst-related processes using mutants in genetically tractable organisms such as Arabidopsis thaliana is limited by the lethality or the severity of phenotypes. We discovered that the small molecule Endosidin2 (ES2) binds to the EXO70 (exocyst component of 70 kDa) subunit of the exocyst complex, resulting in inhibition of exocytosis and endosomal recycling in both plant and human cells andmore » enhancement of plant vacuolar trafficking. An EXO70 protein with a C-terminal truncation results in dominant ES2 resistance, uncovering possible distinct regulatory roles for the N terminus of the protein. Ultimately, this study not only provides a valuable tool in studying exocytosis regulation but also offers a potentially new target for drugs aimed at addressing human disease.« less

Endosidin2 targets conserved exocyst complex subunit EXO70 to inhibit exocytosis

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

Zhang, Chunhua; Brown, Michelle Q.; van de Ven, Wilhelmina

The exocyst complex regulates the last steps of exocytosis, which is essential to organisms across kingdoms. In humans, its dysfunction is correlated with several significant diseases, such as diabetes and cancer progression. Investigation of the dynamic regulation of the evolutionarily conserved exocyst-related processes using mutants in genetically tractable organisms such as Arabidopsis thaliana is limited by the lethality or the severity of phenotypes. We discovered that the small molecule Endosidin2 (ES2) binds to the EXO70 (exocyst component of 70 kDa) subunit of the exocyst complex, resulting in inhibition of exocytosis and endosomal recycling in both plant and human cells andmore » enhancement of plant vacuolar trafficking. An EXO70 protein with a C-terminal truncation results in dominant ES2 resistance, uncovering possible distinct regulatory roles for the N terminus of the protein. Ultimately, this study not only provides a valuable tool in studying exocytosis regulation but also offers a potentially new target for drugs aimed at addressing human disease.« less

Glutamine synthetase in Medicago truncatula, unveiling new secrets of a very old enzyme

PubMed Central

Seabra, Ana R.; Carvalho, Helena G.

2015-01-01

Glutamine synthetase (GS) catalyzes the first step at which nitrogen is brought into cellular metabolism and is also involved in the reassimilation of ammonium released by a number of metabolic pathways. Due to its unique position in plant nitrogen metabolism, GS plays essential roles in all aspects of plant development, from germination to senescence, and is a key component of nitrogen use efficiency (NUE) and plant yield. Understanding the mechanisms regulating GS activity is therefore of utmost importance and a great effort has been dedicated to understand how GS is regulated in different plant species. The present review summarizes exciting recent developments concerning the structure and regulation of GS isoenzymes, using the model legume Medicago truncatula. These include the understanding of the structural determinants of both the cytosolic and plastid located isoenzymes, the existence of a seed-specific GS gene unique to M. truncatula and closely related species and the discovery that GS isoenzymes are regulated by nitric oxide at the post-translational level. The data is discussed and integrated with the potential roles of the distinct GS isoenzymes within the whole plant context. PMID:26284094

45 CFR 91.18 - Age distinctions contained in HHS regulations.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 45 Public Welfare 1 2014-10-01 2014-10-01 false Age distinctions contained in HHS regulations. 91... NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM HHS Standards for Determining Age Discrimination § 91.18 Age distinctions contained in HHS regulations. Any age...

45 CFR 91.18 - Age distinctions contained in HHS regulations.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 1 2010-10-01 2010-10-01 false Age distinctions contained in HHS regulations. 91... NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM HHS Standards for Determining Age Discrimination § 91.18 Age distinctions contained in HHS regulations. Any age...

45 CFR 91.18 - Age distinctions contained in HHS regulations.

Code of Federal Regulations, 2013 CFR

2013-10-01

... 45 Public Welfare 1 2013-10-01 2013-10-01 false Age distinctions contained in HHS regulations. 91... NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM HHS Standards for Determining Age Discrimination § 91.18 Age distinctions contained in HHS regulations. Any age...

45 CFR 91.18 - Age distinctions contained in HHS regulations.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 45 Public Welfare 1 2011-10-01 2011-10-01 false Age distinctions contained in HHS regulations. 91... NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM HHS Standards for Determining Age Discrimination § 91.18 Age distinctions contained in HHS regulations. Any age...

Feedback regulation of TGF-β signaling.

PubMed

Yan, Xiaohua; Xiong, Xiangyang; Chen, Ye-Guang

2018-01-01

Transforming growth factor beta (TGF-β) is a multi-functional polypeptide that plays a critical role in regulating a broad range of cellular functions and physiological processes. Signaling is initiated when TGF-β ligands bind to two types of cell membrane receptors with intrinsic Ser/Thr kinase activity and transmitted by the intracellular Smad proteins, which act as transcription factors to regulate gene expression in the nucleus. Although it is relatively simple and straight-forward, this TGF-β/Smad pathway is regulated by various feedback loops at different levels, including the ligand, the receptor, Smads and transcription, and is thus fine-tuned in terms of signaling robustness, duration, specificity, and plasticity. The precise control gives rise to versatile and context-dependent pathophysiological functions. In this review, we firstly give an overview of TGF-β signaling, and then discuss how each step of TGF-β signaling is finely controlled by distinct modes of feedback mechanisms, involving both protein regulators and miRNAs. © The Author 2017. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Structure and Dynamics of the tRNA-like Structure Domain of Brome Mosaic Virus

NASA Astrophysics Data System (ADS)

Vieweger, Mario; Nesbitt, David

2014-03-01

Conformational switching is widely accepted as regulatory mechanism in gene expression in bacterial systems. More recently, similar regulation mechanisms are emerging for viral systems. One of the most abundant and best studied systems is the tRNA-like structure domain that is found in a number of plant viruses across eight genera. In this work, the folding dynamics of the tRNA-like structure domain of Brome Mosaic Virus are investigated using single-molecule Fluorescence Resonance Energy Transfer techniques. In particular, Burst fluorescence is applied to observe metal-ion induced folding in freely diffusing RNA constructs resembling the 3'-terminal 169nt of BMV RNA3. Histograms of EFRET probabilities reveal a complex equilibrium of three distinct populations. A step-wise kinetic model for TLS folding is developed in accord with the evolution of conformational populations and structural information in the literature. In this mechanism, formation of functional TLS domains from unfolded RNAs requires two consecutive steps; 1) hybridization of a long-range stem interaction followed by 2) formation of a 3' pseudoknot. This three-state equilibrium is well described by step-wise dissociation constants K1(328(30) μM) and K2(1092(183) μM) for [Mg2+] and K1(74(6) mM) and K2(243(52) mM) for [Na+]-induced folding. The kinetic model is validated by oligo competition with the STEM interaction. Implications of this conformational folding mechanism are discussed in regards to regulation of virus replication.

Transcriptional regulation of cranial sensory placode development

PubMed Central

Moody, Sally A.; LaMantia, Anthony-Samuel

2015-01-01

Cranial sensory placodes derive from discrete patches of the head ectoderm, and give rise to numerous sensory structures. During gastrulation, a specialized “neural border zone” forms around the neural plate in response to interactions between the neural and non-neural ectoderm and signals from adjacent mesodermal and/or endodermal tissues. This zone subsequently gives rise to two distinct precursor populations of the peripheral nervous system: the neural crest and the pre-placodal ectoderm (PPE). The PPE is a common field from which all cranial sensory placodes arise (adenohypophyseal, olfactory, lens, trigeminal, epibranchial, otic). Members of the Six family of transcription factors are major regulators of PPE specification, in partnership with co-factor proteins such as Eya. Six gene activity also maintains tissue boundaries between the PPE, neural crest and epidermis by repressing genes that specify the fates of those adjacent ectodermally-derived domains. As the embryo acquires anterior-posterior identity, the PPE becomes transcriptionally regionalized, and it subsequently subdivides into specific placodes with distinct developmental fates in response to signaling from adjacent tissues. Each placode is characterized by a unique transcriptional program that leads to the differentiation of highly specialized cells, such as neurosecretory cells, somatic sensory receptor cells, chemosensory neurons, peripheral glia and supporting cells. In this review, we summarize the transcriptional and signaling factors that regulate key steps of placode development, influence subsequent sensory neuron specification, and discuss what is known about mutations in some of the essential PPE genes that underlie human congenital syndromes. PMID:25662264

Wind energy and Turkey.

PubMed

Coskun, Aynur Aydin; Türker, Yavuz Özhan

2012-03-01

The global energy requirement for sustaining economic activities, meeting social needs and social development is increasing daily. Environmentally friendly, renewable energy resources are an alternative to the primary non-renewable energy resources, which devastate ecosystems in order to meet increasing demand. Among renewable energy sources such as hydropower, biopower, geothermal power and solar power, wind power offers distinct advantages to Turkey. There is an increasing tendency toward wind globally and the European Union adjusted its legal regulations in this regard. As a potential EU Member state, Turkey is going through a similar process. The number of institutional and legal regulations concerning wind power has increased in recent years; technical infrastructure studies were completed, and some important steps were taken in this regard. This study examines the way in which Turkey has developed support for wind power, presents a SWOT analysis of the wind power sector in Turkey and a projection was made for the concrete success expected to be accomplished in the future.

Social Media Blurred the Distinction Between Author and Reader

NASA Astrophysics Data System (ADS)

Lambiotte, Renaud

The last few years have seen the emergence of the sharing economy. As social media blurred the distinction between author and reader, everyone can now offer or receive services thanks to the networking tools provided by new technological companies. Take Uber, and its billion of journeys in 2015 alone, with tens of thousands of vehicles crawling every moment in the globe's biggest cities. As often, when confronted with a technological change, we observe a polarization of society, and the search for an equilibrium characterized by new norms, rights, and obligations. Understanding the mechanisms behind this re-organization requires an integrated, interdisciplinary approach, covering an intricate web of legal, societal, economical, and computational issues which, we believe, could benefit from a complex systems perspective. As a first step, we are currently studying the dynamics of pricing in Uber. In this new de-regulated world, journey prices fluctuate in time depending on traffic but also on the service's perceived balance of passenger demand and driver supply...

Structural analysis of a 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase with an N-terminal chorismate mutase-like regulatory domain

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

Light, Samuel H.; Halavaty, Andrei S.; Minasov, George

2012-06-27

3-Deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAHPS) catalyzes the first step in the biosynthesis of a number of aromatic metabolites. Likely because this reaction is situated at a pivotal biosynthetic gateway, several DAHPS classes distinguished by distinct mechanisms of allosteric regulation have independently evolved. One class of DAHPSs contains a regulatory domain with sequence homology to chorismate mutase - an enzyme further downstream of DAHPS that catalyzes the first committed step in tyrosine/phenylalanine biosynthesis - and is inhibited by chorismate mutase substrate (chorismate) and product (prephenate). Described in this work, structures of the Listeria monocytogenes chorismate/prephenate regulated DAHPS in complex with Mn{sup 2+}more » and Mn{sup 2+} + phosphoenolpyruvate reveal an unusual quaternary architecture: DAHPS domains assemble as a tetramer, from either side of which chorismate mutase-like (CML) regulatory domains asymmetrically emerge to form a pair of dimers. This domain organization suggests that chorismate/prephenate binding promotes a stable interaction between the discrete regulatory and catalytic domains and supports a mechanism of allosteric inhibition similar to tyrosine/phenylalanine control of a related DAHPS class. We argue that the structural similarity of chorismate mutase enzyme and CML regulatory domain provides a unique opportunity for the design of a multitarget antibacterial.« less

Vesicle Docking Is a Key Target of Local PI(4,5)P2 Metabolism in the Secretory Pathway of INS-1 Cells.

PubMed

Ji, Chen; Fan, Fan; Lou, Xuelin

2017-08-08

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P 2 ) signaling is transient and spatially confined in live cells. How this pattern of signaling regulates transmitter release and hormone secretion has not been addressed. We devised an optogenetic approach to control PI(4,5)P 2 levels in time and space in insulin-secreting cells. Combining this approach with total internal reflection fluorescence microscopy, we examined individual vesicle-trafficking steps. Unlike long-term PI(4,5)P 2 perturbations, rapid and cell-wide PI(4,5)P 2 reduction in the plasma membrane (PM) strongly inhibits secretion and intracellular Ca 2+ concentration ([Ca 2+ ] i ) responses, but not sytaxin1a clustering. Interestingly, local PI(4,5)P 2 reduction selectively at vesicle docking sites causes remarkable vesicle undocking from the PM without affecting [Ca 2+ ] i . These results highlight a key role of local PI(4,5)P 2 in vesicle tethering and docking, coordinated with its role in priming and fusion. Thus, different spatiotemporal PI(4,5)P 2 signaling regulates distinct steps of vesicle trafficking, and vesicle docking may be a key target of local PI(4,5)P 2 signaling in vivo. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Metastasis Suppressor Genes: At the Interface Between the Environment and Tumor Cell Growth

PubMed Central

Hurst, Douglas R.; Welch, Danny R.

2013-01-01

The molecular mechanisms and genetic programs required for cancer metastasis are sometimes overlapping, but components are clearly distinct from those promoting growth of a primary tumor. Every sequential, rate-limiting step in the sequence of events leading to metastasis requires coordinated expression of multiple genes, necessary signaling events, and favorable environmental conditions or the ability to escape negative selection pressures. Metastasis suppressors are molecules that inhibit the process of metastasis without preventing growth of the primary tumor. The cellular processes regulated by metastasis suppressors are diverse and function at every step in the metastatic cascade. As we gain knowledge into the molecular mechanisms of metastasis suppressors and cofactors with which they interact, we learn more about the process, including appreciation that some are potential targets for therapy of metastasis, the most lethal aspect of cancer. Until now, metastasis suppressors have been described largely by their function. With greater appreciation of their biochemical mechanisms of action, the importance of context is increasingly recognized especially since tumor cells exist in myriad microenvironments. In this review, we assemble the evidence that selected molecules are indeed suppressors of metastasis, collate the data defining the biochemical mechanisms of action, and glean insights regarding how metastasis suppressors regulate tumor cell communication to–from microenvironments. PMID:21199781

Method of Simulating Flow-Through Area of a Pressure Regulator

NASA Technical Reports Server (NTRS)

Hass, Neal E. (Inventor); Schallhorn, Paul A. (Inventor)

2011-01-01

The flow-through area of a pressure regulator positioned in a branch of a simulated fluid flow network is generated. A target pressure is defined downstream of the pressure regulator. A projected flow-through area is generated as a non-linear function of (i) target pressure, (ii) flow-through area of the pressure regulator for a current time step and a previous time step, and (iii) pressure at the downstream location for the current time step and previous time step. A simulated flow-through area for the next time step is generated as a sum of (i) flow-through area for the current time step, and (ii) a difference between the projected flow-through area and the flow-through area for the current time step multiplied by a user-defined rate control parameter. These steps are repeated for a sequence of time steps until the pressure at the downstream location is approximately equal to the target pressure.

Bromodomain and extraterminal inhibitors block the Epstein-Barr virus lytic cycle at two distinct steps.

PubMed

Keck, Kristin M; Moquin, Stephanie A; He, Amanda; Fernandez, Samantha G; Somberg, Jessica J; Liu, Stephanie M; Martinez, Delsy M; Miranda, Jj L

2017-08-11

Lytic infection by the Epstein-Barr virus (EBV) poses numerous health risks, such as infectious mononucleosis and lymphoproliferative disorder. Proteins in the bromodomain and extraterminal (BET) family regulate multiple stages of viral life cycles and provide promising intervention targets. Synthetic small molecules can bind to the bromodomains and disrupt function by preventing recognition of acetylated lysine substrates. We demonstrate that JQ1 and other BET inhibitors block two different steps in the sequential cascade of the EBV lytic cycle. BET inhibitors prevent expression of the viral immediate-early protein BZLF1. JQ1 alters transcription of genes controlled by the host protein BACH1, and BACH1 knockdown reduces BZLF1 expression. BET proteins also localize to the lytic origin of replication (OriLyt) genetic elements, and BET inhibitors prevent viral late gene expression. There JQ1 reduces BRD4 recruitment during reactivation to preclude replication initiation. This represents a rarely observed dual mode of action for drugs.

Live-cell analysis of endogenous GFP-RPB1 uncovers rapid turnover of initiating and promoter-paused RNA Polymerase II.

PubMed

Steurer, Barbara; Janssens, Roel C; Geverts, Bart; Geijer, Marit E; Wienholz, Franziska; Theil, Arjan F; Chang, Jiang; Dealy, Shannon; Pothof, Joris; van Cappellen, Wiggert A; Houtsmuller, Adriaan B; Marteijn, Jurgen A

2018-05-08

Initiation and promoter-proximal pausing are key regulatory steps of RNA Polymerase II (Pol II) transcription. To study the in vivo dynamics of endogenous Pol II during these steps, we generated fully functional GFP-RPB1 knockin cells. GFP-RPB1 photobleaching combined with computational modeling revealed four kinetically distinct Pol II fractions and showed that on average 7% of Pol II are freely diffusing, while 10% are chromatin-bound for 2.4 seconds during initiation, and 23% are promoter-paused for only 42 seconds. This unexpectedly high turnover of Pol II at promoters is most likely caused by premature termination of initiating and promoter-paused Pol II and is in sharp contrast to the 23 minutes that elongating Pol II resides on chromatin. Our live-cell-imaging approach provides insights into Pol II dynamics and suggests that the continuous release and reinitiation of promoter-bound Pol II is an important component of transcriptional regulation. Copyright © 2018 the Author(s). Published by PNAS.

T Lymphocyte Migration: An Action Movie Starring the Actin and Associated Actors.

PubMed

Dupré, Loïc; Houmadi, Raïssa; Tang, Catherine; Rey-Barroso, Javier

2015-01-01

The actin cytoskeleton is composed of a dynamic filament meshwork that builds the architecture of the cell to sustain its fundamental properties. This physical structure is characterized by a continuous remodeling, which allows cells to accomplish complex motility steps such as directed migration, crossing of biological barriers, and interaction with other cells. T lymphocytes excel in these motility steps to ensure their immune surveillance duties. In particular, actin cytoskeleton remodeling is a key to facilitate the journey of T lymphocytes through distinct tissue environments and to tune their stop and go behavior during the scanning of antigen-presenting cells. The molecular mechanisms controlling actin cytoskeleton remodeling during T lymphocyte motility have been only partially unraveled, since the function of many actin regulators has not yet been assessed in these cells. Our review aims to integrate the current knowledge into a comprehensive picture of how the actin cytoskeleton drives T lymphocyte migration. We will present the molecular actors that control actin cytoskeleton remodeling, as well as their role in the different T lymphocyte motile steps. We will also highlight which challenges remain to be addressed experimentally and which approaches appear promising to tackle them.

Proteomic Analysis of the Cell Cycle of Procylic Form Trypanosoma brucei.

PubMed

Crozier, Thomas W M; Tinti, Michele; Wheeler, Richard J; Ly, Tony; Ferguson, Michael A J; Lamond, Angus I

2018-06-01

We describe a single-step centrifugal elutriation method to produce synchronous Gap1 (G1)-phase procyclic trypanosomes at a scale amenable for proteomic analysis of the cell cycle. Using ten-plex tandem mass tag (TMT) labeling and mass spectrometry (MS)-based proteomics technology, the expression levels of 5325 proteins were quantified across the cell cycle in this parasite. Of these, 384 proteins were classified as cell-cycle regulated and subdivided into nine clusters with distinct temporal regulation. These groups included many known cell cycle regulators in trypanosomes, which validates the approach. In addition, we identify 40 novel cell cycle regulated proteins that are essential for trypanosome survival and thus represent potential future drug targets for the prevention of trypanosomiasis. Through cross-comparison to the TrypTag endogenous tagging microscopy database, we were able to validate the cell-cycle regulated patterns of expression for many of the proteins of unknown function detected in our proteomic analysis. A convenient interface to access and interrogate these data is also presented, providing a useful resource for the scientific community. Data are available via ProteomeXchange with identifier PXD008741 (https://www.ebi.ac.uk/pride/archive/). © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

34 CFR 110.17 - Age distinctions contained in ED's regulations.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 34 Education 1 2013-07-01 2013-07-01 false Age distinctions contained in ED's regulations. 110.17..., DEPARTMENT OF EDUCATION NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Standards for Determining Age Discrimination § 110.17 Age distinctions contained in ED...

34 CFR 110.17 - Age distinctions contained in ED's regulations.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 34 Education 1 2014-07-01 2014-07-01 false Age distinctions contained in ED's regulations. 110.17..., DEPARTMENT OF EDUCATION NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Standards for Determining Age Discrimination § 110.17 Age distinctions contained in ED...

34 CFR 110.17 - Age distinctions contained in ED's regulations.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 34 Education 1 2010-07-01 2010-07-01 false Age distinctions contained in ED's regulations. 110.17..., DEPARTMENT OF EDUCATION NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Standards for Determining Age Discrimination § 110.17 Age distinctions contained in ED...

Expression, subcellular localization, and cis-regulatory structure of duplicated phytoene synthase genes in melon (Cucumis melo L.).

PubMed

Qin, Xiaoqiong; Coku, Ardian; Inoue, Kentaro; Tian, Li

2011-10-01

Carotenoids perform many critical functions in plants, animals, and humans. It is therefore important to understand carotenoid biosynthesis and its regulation in plants. Phytoene synthase (PSY) catalyzes the first committed and rate-limiting step in carotenoid biosynthesis. While PSY is present as a single copy gene in Arabidopsis, duplicated PSY genes have been identified in many economically important monocot and dicot crops. CmPSY1 was previously identified from melon (Cucumis melo L.), but was not functionally characterized. We isolated a second PSY gene, CmPSY2, from melon in this work. CmPSY2 possesses a unique intron/exon structure that has not been observed in other plant PSYs. Both CmPSY1 and CmPSY2 are functional in vitro, but exhibit distinct expression patterns in different melon tissues and during fruit development, suggesting differential regulation of the duplicated melon PSY genes. In vitro chloroplast import assays verified the plastidic localization of CmPSY1 and CmPSY2 despite the lack of an obvious plastid target peptide in CmPSY2. Promoter motif analysis of the duplicated melon and tomato PSY genes and the Arabidopsis PSY revealed distinctive cis-regulatory structures of melon PSYs and identified gibberellin-responsive motifs in all PSYs except for SlPSY1, which has not been reported previously. Overall, these data provide new insights into the evolutionary history of plant PSY genes and the regulation of PSY expression by developmental and environmental signals that may involve different regulatory networks.

Derivation of linearized transfer functions for switching-mode regulations. Phase A: Current step-up and voltage step-up converters

NASA Technical Reports Server (NTRS)

Wong, R. C.; Owen, H. A., Jr.; Wilson, T. G.

1981-01-01

Small-signal models are derived for the power stage of the voltage step-up (boost) and the current step-up (buck) converters. The modeling covers operation in both the continuous-mmf mode and the discontinuous-mmf mode. The power stage in the regulated current step-up converter on board the Dynamics Explorer Satellite is used as an example to illustrate the procedures in obtaining the small-signal functions characterizing a regulated converter.

X-Chromosome dosage compensation.

PubMed

Meyer, Barbara J

2005-06-25

In mammals, flies, and worms, sex is determined by distinctive regulatory mechanisms that cause males (XO or XY) and females (XX) to differ in their dose of X chromosomes. In each species, an essential X chromosome-wide process called dosage compensation ensures that somatic cells of either sex express equal levels of X-linked gene products. The strategies used to achieve dosage compensation are diverse, but in all cases, specialized complexes are targeted specifically to the X chromosome(s) of only one sex to regulate transcript levels. In C. elegans, this sex-specific targeting of the dosage compensation complex (DCC) is controlled by the same developmental signal that establishes sex, the ratio of X chromosomes to sets of autosomes (X:A signal). Molecular components of this chromosome counting process have been defined. Following a common step of regulation, sex determination and dosage compensation are controlled by distinct genetic pathways. C. elegans dosage compensation is implemented by a protein complex that binds both X chromosomes of hermaphrodites to reduce transcript levels by one-half. The dosage compensation complex resembles the conserved 13S condensin complex required for both mitotic and meiotic chromosome resolution and condensation, implying the recruitment of ancient proteins to the new task of regulating gene expression. Within each C. elegans somatic cell, one of the DCC components also participates in the separate mitotic/meiotic condensin complex. Other DCC components play pivotal roles in regulating the number and distribution of crossovers during meiosis. The strategy by which C. elegans X chromosomes attract the condensin-like DCC is known. Small, well-dispersed X-recognition elements act as entry sites to recruit the dosage compensation complex and to nucleate spreading of the complex to X regions that lack recruitment sites. In this manner, a repressed chromatin state is spread in cis over short or long distances, thus establishing the global, epigenetic regulation of X chromosomes that is maintained throughout the lifetime of hermaphrodites.

Different KChIPs compete for heteromultimeric assembly with pore-forming Kv4 subunits.

PubMed

Zhou, Jingheng; Tang, Yiquan; Zheng, Qin; Li, Meng; Yuan, Tianyi; Chen, Liangyi; Huang, Zhuo; Wang, KeWei

2015-06-02

Auxiliary Kv channel-interacting proteins 1-4 (KChIPs1-4) coassemble with pore-forming Kv4 α-subunits to form channel complexes underlying somatodendritic subthreshold A-type current that regulates neuronal excitability. It has been hypothesized that different KChIPs can competitively bind to Kv4 α-subunit to form variable channel complexes that can exhibit distinct biophysical properties for modulation of neural function. In this study, we use single-molecule subunit counting by total internal reflection fluorescence microscopy in combinations with electrophysiology and biochemistry to investigate whether different isoforms of auxiliary KChIPs, KChIP4a, and KChIP4bl, can compete for binding of Kv4.3 to coassemble heteromultimeric channel complexes for modulation of channel function. To count the number of photobleaching steps solely from cell membrane, we take advantage of a membrane tethered k-ras-CAAX peptide that anchors cytosolic KChIP4 proteins to the surface for reduction of background noise. Single-molecule subunit counting reveals that the number of KChIP4 isoforms in Kv4.3-KChIP4 complexes can vary depending on the KChIP4 expression level. Increasing the amount of KChIP4bl gradually reduces bleaching steps of KChIP4a isoform proteins, and vice versa. Further analysis of channel gating kinetics from different Kv4-KChIP4 subunit compositions confirms that both KChIP4a and KChIP4bl can modulate the channel complex function upon coassembly. Taken together, our findings show that auxiliary KChIPs can heteroassemble with Kv4 in a competitive manner to form heteromultimeric Kv4-KChIP4 channel complexes that are biophysically distinct and regulated under physiological or pathological conditions. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

DKWSLLL, a versatile DXXXLL-type signal with distinct roles in the Cu(+)-regulated trafficking of ATP7B.

PubMed

Lalioti, Vasiliki; Hernandez-Tiedra, Sonia; Sandoval, Ignacio V

2014-08-01

In the liver, the P-type ATPase and membrane pump ATP7B plays a crucial role in Cu(+) donation to cuproenzymes and in the elimination of excess Cu(+). ATP7B is endowed with a COOH-cytoplasmic (DE)XXXLL-type traffic signal. We find that accessory (Lys -3, Trp -2, Ser -1 and Leu +2) and canonical (D -4, Leu 0 and Leu +1) residues confer the DKWSLLL signal with the versatility required for the Cu(+)-regulated cycling of ATP7B between the trans-Golgi network (TGN) and the plasma membrane (PM). The separate mutation of these residues caused a disruption of the signal, resulting in different ATP7B distribution phenotypes. These phenotypes indicate the key roles of specific residues at separate steps of ATP7B trafficking, including sorting at the TGN, transport from the TGN to the PM and its endocytosis, and recycling to the TGN and PM. The distinct roles of ATP7B in the TGN and PM and the variety of phenotypes caused by the mutation of the canonical and accessory residues of the DKWSLLL signal can explain the separate or joined presentation of Wilson's cuprotoxicosis and the dysfunction of the cuproenzymes that accept Cu(+) at the TGN. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The general alcoholics anonymous tools of recovery: the adoption of 12-step practices and beliefs.

PubMed

Greenfield, Brenna L; Tonigan, J Scott

2013-09-01

Working the 12 steps is widely prescribed for Alcoholics Anonymous (AA) members although the relative merits of different methods for measuring step work have received minimal attention and even less is known about how step work predicts later substance use. The current study (1) compared endorsements of step work on an face-valid or direct measure, the Alcoholics Anonymous Inventory (AAI), with an indirect measure of step work, the General Alcoholics Anonymous Tools of Recovery (GAATOR); (2) evaluated the underlying factor structure of the GAATOR and changes in step work over time; (3) examined changes in the endorsement of step work over time; and (4) investigated how, if at all, 12-step work predicted later substance use. New AA affiliates (N = 130) completed assessments at intake, 3, 6, and 9 months. Significantly more participants endorsed step work on the GAATOR than on the AAI for nine of the 12 steps. An exploratory factor analysis revealed a two-factor structure for the GAATOR comprising behavioral step work and spiritual step work. Behavioral step work did not change over time, but was predicted by having a sponsor, while Spiritual step work decreased over time and increases were predicted by attending 12-step meetings or treatment. Behavioral step work did not prospectively predict substance use. In contrast, spiritual step work predicted percent days abstinent. Behavioral step work and spiritual step work appear to be conceptually distinct components of step work that have distinct predictors and unique impacts on outcomes. PsycINFO Database Record (c) 2013 APA, all rights reserved.

Myb permits multilineage airway epithelial cell differentiation

PubMed Central

Pan, Jie-hong; Adair-Kirk, Tracy L.; Patel, Anand C.; Huang, Tao; Yozamp, Nicholas S.; Xu, Jian; Reddy, E. Premkumar; Byers, Derek E.; Pierce, Richard A.; Holtzman, Michael J.; Brody, Steven L.

2014-01-01

The epithelium of the pulmonary airway is specially differentiated to provide defense against environmental insults, but also subject to dysregulated differentiation that results in lung disease. The current paradigm for airway epithelial differentiation is a one-step program whereby a p63+ basal epithelial progenitor cell generates a ciliated or secretory cell lineage, but the cue for this transition and whether there are intermediate steps is poorly defined. Here we identify transcription factor Myb as a key regulator that permits early multilineage differentiation of airway epithelial cells. Myb+ cells were identified as p63− and therefore distinct from basal progenitor cells, but were still negative for markers of differentiation. Myb RNAi treatment of primary-culture airway epithelial cells and Myb gene deletion in mice resulted in a p63− population with failed maturation of Foxj1+ ciliated cells, as well as Scbg1a1+ and Muc5ac+ secretory cells. Consistent with these findings, analysis of whole genome expression of Myb-deficient cells identified Myb-dependent programs for ciliated and secretory cell differentiation. Myb+ cells were rare in human airways but were increased in regions of ciliated cells and mucous cell hyperplasia in samples from subjects with chronic obstructive pulmonary disease. Together, the results show that a p63− Myb+ population of airway epithelial cells represents a distinct intermediate stage of differentiation that is required under normal conditions and may be heightened in airway disease. PMID:25103188

Two Forkhead transcription factors regulate the division of cardiac progenitor cells by a Polo-dependent pathway

PubMed Central

Ahmad, Shaad M.; Tansey, Terese R.; Busser, Brian W.; Nolte, Michael T.; Jeffries, Neal; Gisselbrecht, Stephen S.; Rusan, Nasser M.; Michelson, Alan M.

2012-01-01

SUMMARY The development of a complex organ requires the specification of appropriate numbers of each of its constituent cell types, as well as their proper differentiation and correct positioning relative to each other. During Drosophila cardiogenesis, all three of these processes are controlled by jumeau (jumu) and Checkpoint suppressor homologue (CHES-1-like), two genes encoding forkhead transcription factors that we discovered utilizing an integrated genetic, genomic and computational strategy for identifying genes expressed in the developing Drosophila heart. Both jumu and CHES-1-like are required during asymmetric cell division for the derivation of two distinct cardiac cell types from their mutual precursor, and in symmetric cell divisions that produce yet a third type of heart cell. jumu and CHES-1-like control the division of cardiac progenitors by regulating the activity of Polo, a kinase involved in multiple steps of mitosis. This pathway demonstrates how transcription factors integrate diverse developmental processes during organogenesis. PMID:22814603

Transcriptional regulation of development in heterocyst-forming cyanobacteria.

PubMed

Flores, Enrique; Picossi, Silvia; Valladares, Ana; Herrero, Antonia

2018-04-30

Filamentous, heterocyst-forming cyanobacteria are among the simplest multicellular systems in Nature. In the absence of combined nitrogen, the filaments consist of vegetative cells that fix CO 2 through oxygenic photosynthesis and micro-oxic heterocysts specialized for the fixation of N 2 in a proportion of about 10 to 1. The development of a heterocyst-containing filament involves differentiation of vegetative cells into heterocysts in a process that requires a distinct gene expression program. Two transcription factors are strictly required, NtcA and HetR. The CRP-family protein NtcA directly activates the expression of multiple genes during heterocyst differentiation - in some cases assisted by coactivators including HetR - and in mature heterocysts, whereas HetR is needed to build high NtcA levels in differentiating heterocysts and directly activates some particular genes. A few other regulators of gene expression participate at specific differentiation steps, and a specific transcription factor, CnfR, activates nif gene expression under the micro-oxic conditions of the heterocyst. Copyright © 2018 Elsevier B.V. All rights reserved.

Identification of Neuregulin-2 as a novel stress granule component.

PubMed

Kim, Jin Ah; Jayabalan, Aravinth Kumar; Kothandan, Vinoth Kumar; Mariappan, Ramesh; Kee, Younghoon; Ohn, Takbum

2016-08-01

Stress Granules (SGs) are microscopically visible, phase dense aggregates of translationally stalled messenger ribonucleoprotein (mRNP) complexes formed in response to distinct stress conditions. It is generally considered that SG formation is induced to protect cells from conditions of stress. The precise constituents of SGs and the mechanism through which SGs are dynamically regulated in response to stress are not completely understood. Hence, it is important to identify proteins which regulate SG assembly and disassembly. In the present study, we report Neuregulin-2 (NRG2) as a novel component of SGs; furthermore, depletion of NRG2 potently inhibits SG formation. We also demonstrate that NRG2 specifically localizes to SGs under various stress conditions. Knockdown of NRG2 has no effect on stress-induced polysome disassembly, suggesting that the component does not influence early step of SG formation. It was also observed that reduced expression of NRG2 led to marginal increase in cell survival under arsenite-induced stress. [BMB Reports 2016; 49(8): 449-454].

Identification of Neuregulin-2 as a novel stress granule component

PubMed Central

Kim, Jin Ah; Jayabalan, Aravinth Kumar; Kothandan, Vinoth Kumar; Mariappan, Ramesh; Kee, Younghoon; Ohn, Takbum

2016-01-01

Stress Granules (SGs) are microscopically visible, phase dense aggregates of translationally stalled messenger ribonucleoprotein (mRNP) complexes formed in response to distinct stress conditions. It is generally considered that SG formation is induced to protect cells from conditions of stress. The precise constituents of SGs and the mechanism through which SGs are dynamically regulated in response to stress are not completely understood. Hence, it is important to identify proteins which regulate SG assembly and disassembly. In the present study, we report Neuregulin-2 (NRG2) as a novel component of SGs; furthermore, depletion of NRG2 potently inhibits SG formation. We also demonstrate that NRG2 specifically localizes to SGs under various stress conditions. Knockdown of NRG2 has no effect on stress-induced polysome disassembly, suggesting that the component does not influence early step of SG formation. It was also observed that reduced expression of NRG2 led to marginal increase in cell survival under arsenite-induced stress. [BMB Reports 2016; 49(8): 449-454] PMID:27345716

Ubiquitinated Sirtuin 1 (SIRT1) Function Is Modulated during DNA Damage-induced Cell Death and Survival*

PubMed Central

Peng, Lirong; Yuan, Zhigang; Li, Yixuan; Ling, Hongbo; Izumi, Victoria; Fang, Bin; Fukasawa, Kenji; Koomen, John; Chen, Jiandong; Seto, Edward

2015-01-01

Downstream signaling of physiological and pathological cell responses depends on post-translational modification such as ubiquitination. The mechanisms regulating downstream DNA damage response (DDR) signaling are not completely elucidated. Sirtuin 1 (SIRT1), the founding member of Class III histone deacetylases, regulates multiple steps in DDR and is closely associated with many physiological and pathological processes. However, the role of post-translational modification or ubiquitination of SIRT1 during DDR is unclear. We show that SIRT1 is dynamically and distinctly ubiquitinated in response to DNA damage. SIRT1 was ubiquitinated by the MDM2 E3 ligase in vitro and in vivo. SIRT1 ubiquitination under normal conditions had no effect on its enzymatic activity or rate of degradation; hypo-ubiquitination, however, reduced SIRT1 nuclear localization. Ubiquitination of SIRT1 affected its function in cell death and survival in response to DNA damage. Our results suggest that ubiquitination is required for SIRT1 function during DDR. PMID:25670865

Integrin Beta 1 Suppresses Multilayering of a Simple Epithelium

PubMed Central

Chen, Jichao; Krasnow, Mark A.

2012-01-01

Epithelia are classified as either simple, a single cell layer thick, or stratified (multilayered). Stratified epithelia arise from simple epithelia during development, and transcription factor p63 functions as a key positive regulator of epidermal stratification. Here we show that deletion of integrin beta 1 (Itgb1) in the developing mouse airway epithelium abrogates airway branching and converts this monolayer epithelium into a multilayer epithelium with more than 10 extra layers. Mutant lung epithelial cells change mitotic spindle orientation to seed outer layers, and cells in different layers become molecularly and functionally distinct, hallmarks of normal stratification. However, mutant lung epithelial cells do not activate p63 and do not switch to the stratified keratin profile of epidermal cells. These data, together with previous data implicating Itgb1 in regulation of epidermal stratification, suggest that the simple-versus-stratified developmental decision may involve not only stratification inducers like p63 but suppressors like Itgb1 that prevent simple epithelia from inappropriately activating key steps in the stratification program. PMID:23285215

Common threads in cardiac fibrosis, infarct scar formation, and wound healing.

PubMed

Czubryt, Michael P

2012-11-01

Wound healing, cardiac fibrosis, and infarct scar development, while possessing distinct features, share a number of key functional similarities, including extracellular matrix synthesis and remodeling by fibroblasts and myofibroblasts. Understanding the underlying mechanisms that are common to these processes may suggest novel therapeutic approaches for pathologic situations such as fibrosis, or defective wound healing such as hypertrophic scarring or keloid formation. This manuscript will briefly review the major steps of wound healing, and will contrast this process with how cardiac infarct scar formation or interstitial fibrosis occurs. The feasibility of targeting common pro-fibrotic growth factor signaling pathways will be discussed. Finally, the potential exploitation of novel regulators of wound healing and fibrosis (ski and scleraxis), will be examined.

A One-Step Immunostaining Method to Visualize Rodent Muscle Fiber Type within a Single Specimen

PubMed Central

Sawano, Shoko; Komiya, Yusuke; Ichitsubo, Riho; Ohkawa, Yasuyuki; Nakamura, Mako; Tatsumi, Ryuichi; Ikeuchi, Yoshihide; Mizunoya, Wataru

2016-01-01

In this study, we present a quadruple immunostaining method for rapid muscle fiber typing of mice and rats using antibodies specific to the adult myosin heavy chain (MyHC) isoforms MyHC1, 2A, 2X, and 2B, which are common marker proteins of distinct muscle fiber types. We developed rat monoclonal antibodies specific to each MyHC isoform and conjugated these four antibodies to fluorophores with distinct excitation and emission wavelengths. By mixing the four types of conjugated antibodies, MyHC1, 2A, 2X, and 2B could be distinguished within a single specimen allowing for facile delineation of skeletal muscle fiber types. Furthermore, we could observe hybrid fibers expressing MyHC2X and MyHC2B together in single longitudinal muscle sections from mice and rats, that was not attained in previous techniques. This staining method is expected to be applied to study muscle fiber type transition in response to environmental factors, and to ultimately develop techniques to regulate animal muscle fiber types. PMID:27814384

The General Alcoholics Anonymous Tools of Recovery: The Adoption of 12-Step Practices and Beliefs

PubMed Central

Greenfield, Brenna L.; Tonigan, J. Scott

2013-01-01

Working the 12 steps is widely prescribed for Alcoholics Anonymous (AA) members although the relative merits of different methods for measuring step-work have received minimal attention and even less is known about how step-work predicts later substance use. The current study (1) compared endorsements of step-work on an face-valid or direct measure, the Alcoholics Anonymous Inventory (AAI), with an indirect measure of step-work, the General Alcoholics Anonymous Tools of Recovery (GAATOR), (2) evaluated the underlying factor structure of the GAATOR and changes in step-work over time, (3) examined changes in the endorsement of step-work over time, and (4) investigated how, if at all, 12-step-work predicted later substance use. New AA affiliates (N = 130) completed assessments at intake, 3, 6, and 9 months. Significantly more participants endorsed step-work on the GAATOR than on the AAI for nine of the 12 steps. An exploratory factor analysis revealed a two-factor structure for the GAATOR comprising Behavioral Step-Work and Spiritual Step-Work. Behavioral Step-Work did not change over time, but was predicted by having a sponsor, while Spiritual Step-Work decreased over time and increases were predicted by attending 12-step meetings or treatment. Behavioral Step-Work did not prospectively predict substance use. In contrast, Spiritual Step-Work predicted percent days abstinent, an effect that is consistent with recent work on the mediating effects of spiritual growth, AA, and increased abstinence. Behavioral and Spiritual Step-Work appear to be conceptually distinct components of step-work that have distinct predictors and unique impacts on outcomes. PMID:22867293

Individual Distinctive Features of Self-Regulation Processes Peculiar to Students of Different Profiles of Lateral Organization

ERIC Educational Resources Information Center

Korneeva, Svetlana A.; Zherebnenko, Oksana A.; Mukhamedzyanova, Flera G.; Moskalenko, Svetlana V.; Gorelikova, Olga N.

2016-01-01

The research paper presents an analysis of the interrelation between the lateral organisation profiles' indicators and self-regulation features. The existence of significant distinctions in the processes of self-regulation among respondents with different variants of lateral profiles of the interhemispheric asymmetry is proved, as well as the…

NEW TYPE OF STREPTOMYCIN RESISTANCE RESULTING FROM ACTION OF THE EPISOMELIKE MUTATOR FACTOR IN ESCHERICHIA COLI

PubMed Central

Gundersen, Wenche B.

1963-01-01

Gundersen, Wenche B. (Oslo University, Oslo, Norway). New type of streptomycin resistance resulting from action of the episomelike mutator factor in Escherichia coli. J. Bacteriol. 86:510–516. 1963.—Analyses have been performed to elucidate the genetic nature of the streptomycin resistance that results from the action of the previously described episomelike mutator factor in Escherichia coli. This streptomycin resistance has been found to differ from ordinary one-step streptomycin resistance. The new type of streptomycin resistance, “mutator resistance,” can be lost, either spontaneously or by treatment with ultraviolet light and acriflavine. It is more stable in a K-12 strain than in the original E. coli strain 635. Mutator resistance segregates like a chromosomal marker in genetic crosses, and is located near the ordinary streptomycin locus. The locus for mutator resistance is distinct from that of ordinary streptomycin resistance, apparently located further toward the threonine region. Mutator resistance, unlike ordinary one-step streptomycin resistance, appears as a dominant character. The possibilities of its being a suppressor or regulator mutation are discussed. PMID:14066430

A gene expression system offering multiple levels of regulation: the Dual Drug Control (DDC) system.

PubMed

Sudomoina, Marina; Latypova, Ekaterina; Favorova, Olga O; Golemis, Erica A; Serebriiskii, Ilya G

2004-04-29

Whether for cell culture studies of protein function, construction of mouse models to enable in vivo analysis of disease epidemiology, or ultimately gene therapy of human diseases, a critical enabling step is the ability to achieve finely controlled regulation of gene expression. Previous efforts to achieve this goal have explored inducible drug regulation of gene expression, and construction of synthetic promoters based on two-hybrid paradigms, among others. In this report, we describe the combination of dimerizer-regulated two-hybrid and tetracycline regulatory elements in an ordered cascade, placing expression of endpoint reporters under the control of two distinct drugs. In this Dual Drug Control (DDC) system, a first plasmid expresses fusion proteins to DBD and AD, which interact only in the presence of a small molecule dimerizer; a second plasmid encodes a cassette transcriptionally responsive to the first DBD, directing expression of the Tet-OFF protein; and a third plasmid encodes a reporter gene transcriptionally responsive to binding by Tet-OFF. We evaluate the dynamic range and specificity of this system in comparison to other available systems. This study demonstrates the feasibility of combining two discrete drug-regulated expression systems in a temporally sequential cascade, without loss of dynamic range of signal induction. The efficient layering of control levels allowed by this combination of elements provides the potential for the generation of complex control circuitry that may advance ability to regulate gene expression in vivo.

Performance goals in conflictual social interactions: towards the distinction between two modes of relational conflict regulation.

PubMed

Sommet, Nicolas; Darnon, Céline; Mugny, Gabriel; Quiamzade, Alain; Pulfrey, Caroline; Dompnier, Benoît; Butera, Fabrizio

2014-03-01

Socio-cognitive conflict has been defined as a situation of confrontation with a disagreeing other. Previous research suggests that individuals can regulate conflict in a relational way, namely by focusing on social comparison between relative levels of competences. Relational conflict regulation has been described as yielding particularly negative effects on social interactions and learning, but has been understudied. The present research addresses the question of the origin of relational conflict regulation by introducing a fundamental distinction between two types of regulation, one based on the affirmation of one's own point of view and the invalidation of the other's (i.e., 'competitive' regulation), the other corresponding to the protection of self-competence via compliance (i.e., 'protective' regulation). Three studies show that these modes of relational conflict regulation result from the endorsement of distinct performance goals, respectively, performance-approach goals (trying to outperform others) and performance-avoidance goals (avoiding performing more poorly than others). Theoretical implications for the literature on both conflict regulation and achievement goals are discussed. © 2012 The British Psychological Society.

Divide and conquer: The Pseudomonas aeruginosa two-component hybrid SagS enables biofilm formation and recalcitrance of biofilm cells to antimicrobial agents via distinct regulatory circuits

PubMed Central

Petrova, Olga E.; Gupta, Kajal; Liao, Julie; Goodwine, James S.; Sauer, Karin

2017-01-01

The opportunistic pathogen Pseudomonas aeruginosa forms antimicrobial resistant biofilms through sequential steps requiring several two-component regulatory systems. The sensor-regulator hybrid SagS plays a central role in biofilm development by enabling the switch from the planktonic to the biofilm mode of growth, and by facilitating the transition of biofilm cells to a highly tolerant state. However, the mechanism by which SagS accomplishes both functions is unknown. SagS harbors a periplasmic sensory HmsP, and phosphorelay HisKA and Rec domains. We used SagS domain constructs and site-directed mutagenesis to elucidate how SagS performs its dual functions. We demonstrate that HisKA-Rec and the phospho-signaling between SagS and BfiS contribute to the switch to the biofilm mode of growth, but not to the tolerant state. Instead, expression of SagS domain constructs harboring HmsP rendered ΔsagS biofilm cells as recalcitrant to antimicrobial agents as wild-type biofilms, likely by restoring BrlR production and cellular c-di-GMP levels to wild-type levels. Restoration of biofilm tolerance by HmsP was independent of biofilm biomass accumulation, RsmA, RsmYZ, HptB, and BfiSR-downstream targets. Our findings thus suggest that SagS likely makes use of a “divide-and-conquer” mechanism to regulate its dual switch function, by activating two distinct regulatory networks via its individual domains. PMID:28263038

Parallel Allostery by cAMP and PDE Coordinates Activation and Termination Phases in cAMP Signaling.

PubMed

Krishnamurthy, Srinath; Tulsian, Nikhil Kumar; Chandramohan, Arun; Anand, Ganesh S

2015-09-15

The second messenger molecule cAMP regulates the activation phase of the cAMP signaling pathway through high-affinity interactions with the cytosolic cAMP receptor, the protein kinase A regulatory subunit (PKAR). Phosphodiesterases (PDEs) are enzymes responsible for catalyzing hydrolysis of cAMP to 5' AMP. It was recently shown that PDEs interact with PKAR to initiate the termination phase of the cAMP signaling pathway. While the steps in the activation phase are well understood, steps in the termination pathway are unknown. Specifically, the binding and allosteric networks that regulate the dynamic interplay between PKAR, PDE, and cAMP are unclear. In this study, PKAR and PDE from Dictyostelium discoideum (RD and RegA, respectively) were used as a model system to monitor complex formation in the presence and absence of cAMP. Amide hydrogen/deuterium exchange mass spectrometry was used to monitor slow conformational transitions in RD, using disordered regions as conformational probes. Our results reveal that RD regulates its interactions with cAMP and RegA at distinct loci by undergoing slow conformational transitions between two metastable states. In the presence of cAMP, RD and RegA form a stable ternary complex, while in the absence of cAMP they maintain transient interactions. RegA and cAMP each bind at orthogonal sites on RD with resultant contrasting effects on its dynamics through parallel allosteric relays at multiple important loci. RD thus serves as an integrative node in cAMP termination by coordinating multiple allosteric relays and governing the output signal response. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Differential Regulation of Synaptic Vesicle Tethering and Docking by UNC-18 and TOM-1.

PubMed

Gracheva, Elena O; Maryon, Ed B; Berthelot-Grosjean, Martine; Richmond, Janet E

2010-01-01

The assembly of SNARE complexes between syntaxin, SNAP-25 and synaptobrevin is required to prime synaptic vesicles for fusion. Since Munc18 and tomosyn compete for syntaxin interactions, the interplay between these proteins is predicted to be important in regulating synaptic transmission. We explored this possibility, by examining genetic interactions between C. elegans unc-18(Munc18), unc-64(syntaxin) and tom-1(tomosyn). We have previously demonstrated that unc-18 mutants have reduced synaptic transmission, whereas tom-1 mutants exhibit enhanced release. Here we show that the unc-18 mutant release defect is associated with loss of two morphologically distinct vesicle pools; those tethered within 25 nm of the plasma membrane and those docked with the plasma membrane. In contrast, priming defective unc-13 mutants accumulate tethered vesicles, while docked vesicles are greatly reduced, indicating tethering is UNC-18-dependent and occurs in the absence of priming. C. elegans unc-64 mutants phenocopy unc-18 mutants, losing both tethered and docked vesicles, whereas overexpression of open syntaxin preferentially increases vesicle docking, suggesting UNC-18/closed syntaxin interactions are responsible for vesicle tethering. Given the competition between vertebrate tomosyn and Munc18, for syntaxin binding, we hypothesized that C. elegans TOM-1 may inhibit both UNC-18-dependent vesicle targeting steps. Consistent with this hypothesis, tom-1 mutants exhibit enhanced UNC-18 plasma membrane localization and a concomitant increase in both tethered and docked synaptic vesicles. Furthermore, in tom-1;unc-18 double mutants the docked, primed vesicle pool is preferentially rescued relative to unc-18 single mutants. Together these data provide evidence for the differential regulation of two vesicle targeting steps by UNC-18 and TOM-1 through competitive interactions with syntaxin.

7 CFR 51.2842 - Dormant.

Code of Federal Regulations, 2011 CFR

2011-01-01

... Regulations of the Department of Agriculture AGRICULTURAL MARKETING SERVICE (Standards, Inspections, Marketing Practices), DEPARTMENT OF AGRICULTURE REGULATIONS AND STANDARDS UNDER THE AGRICULTURAL MARKETING ACT OF 1946... indicated by distinct elongation of the growing point or distinct yellow or green color in the tip of the...

STriatal-Enriched protein tyrosine Phosphatase (STEP) Regulates the PTPα/Fyn Signaling Pathway

PubMed Central

Xu, Jian; Kurup, Pradeep; Foscue, Ethan; Lombroso, Paul J.

2015-01-01

The tyrosine kinase Fyn has two regulatory tyrosine residues that when phosphorylated either activate (Tyr420) or inhibit (Tyr531) Fyn activity. Within the central nervous system, two protein tyrosine phosphatases (PTPs) target these regulatory tyrosines in Fyn. PTPα dephosphorylates Tyr531 and activates Fyn, while STEP (STriatal-Enriched protein tyrosine Phosphatase) dephosphorylates Tyr420 and inactivates Fyn. Thus, PTPα and STEP have opposing functions in the regulation of Fyn; however, whether there is cross talk between these two PTPs remains unclear. Here, we used molecular techniques in primary neuronal cultures and in vivo to demonstrate that STEP negatively regulates PTPα by directly dephosphorylating PTPα at its regulatory Tyr789. Dephosphorylation of Tyr789 prevents the translocation of PTPα to synaptic membranes, blocking its ability to interact with and activate Fyn. Genetic or pharmacologic reduction of STEP61 activity increased the phosphorylation of PTPα at Tyr789, as well as increased translocation of PTPα to synaptic membranes. Activation of PTPα and Fyn and trafficking of GluN2B to synaptic membranes are necessary for ethanol intake behaviors in rodents. We tested the functional significance of STEP61 in this signaling pathway by ethanol administration to primary cultures as well as in vivo, and demonstrated that the inactivation of STEP61 by ethanol leads to the activation of PTPα, its translocation to synaptic membranes, and the activation of Fyn. These findings indicate a novel mechanism by which STEP61 regulates PTPα and suggest that STEP and PTPα coordinate the regulation of Fyn. PMID:25951993

STRIATAL-ENRICHED PROTEIN TYROSINE PHOSPHATASE (STEP) KNOCKOUT MICE HAVE ENHANCED HIPPOCAMPAL MEMORY

PubMed Central

Venkitaramani, Deepa V.; Moura, Paula J.; Picciotto, Marina R.; Lombroso, Paul J.

2011-01-01

STEP is a brain-specific phosphatase that opposes synaptic strengthening by the regulation of key synaptic signaling proteins. Previous studies suggest a possible role for STriatal-Enriched protein tyrosine Phosphatase (STEP) in learning and memory. To demonstrate the functional importance of STEP in learning and memory, we generated STEP knockout (KO) mice and examined the effect of deletion of STEP on behavioral performance, as well as the phosphorylation and expression of its substrates. Here we report that loss of STEP leads to significantly enhanced performance in hippocampal-dependent learning and memory tasks. In addition, STEP KO mice displayed greater dominance behavior, although they were normal in their motivation, motor coordination, visual acuity and social interactions. STEP KO mice displayed enhanced tyrosine phosphorylation of extracellular-signal regulated kinase 1/2 (ERK1/2), the NR2B subunit of the N-methyl-D-aspartate receptor (NMDAR), Proline-rich tyrosine kinase (Pyk2), as well as an increased phosphorylation of ERK1/2 substrates. Concomitant to the increased phosphorylation of NR2B, synaptosomal expression of NR1/NR2B NMDARs was increased in STEP KO mice, as was the GluR1/GluR2 containing α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptors (AMPAR), providing a potential molecular mechanism for the improved cognitive performance. The data support a role for STEP in the regulation of synaptic strengthening. The absence of STEP improves cognitive performance, and may do so by the regulation of downstream effectors necessary for synaptic transmission. PMID:21501258

29 CFR 35.17 - Age distinctions in DOL regulations.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 29 Labor 1 2010-07-01 2010-07-01 true Age distinctions in DOL regulations. 35.17 Section 35.17 Labor Office of the Secretary of Labor NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF LABOR Standards for Determining Age...

29 CFR 35.17 - Age distinctions in DOL regulations.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 29 Labor 1 2014-07-01 2013-07-01 true Age distinctions in DOL regulations. 35.17 Section 35.17 Labor Office of the Secretary of Labor NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF LABOR Standards for Determining Age...

29 CFR 35.17 - Age distinctions in DOL regulations.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 29 Labor 1 2013-07-01 2013-07-01 false Age distinctions in DOL regulations. 35.17 Section 35.17 Labor Office of the Secretary of Labor NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF LABOR Standards for Determining Age...

29 CFR 35.17 - Age distinctions in DOL regulations.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 29 Labor 1 2012-07-01 2012-07-01 false Age distinctions in DOL regulations. 35.17 Section 35.17 Labor Office of the Secretary of Labor NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF LABOR Standards for Determining Age...

29 CFR 35.17 - Age distinctions in DOL regulations.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 29 Labor 1 2011-07-01 2011-07-01 false Age distinctions in DOL regulations. 35.17 Section 35.17 Labor Office of the Secretary of Labor NONDISCRIMINATION ON THE BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF LABOR Standards for Determining Age...

Thyroid hormone induction of human cholesterol 7 alpha-hydroxylase (Cyp7a1) in vitro.

PubMed

Lammel Lindemann, Jan A; Angajala, Anusha; Engler, David A; Webb, Paul; Ayers, Stephen D

2014-05-05

Thyroid hormone (TH) modulates serum cholesterol by acting on TH receptor β1 (TRβ1) in liver to regulate metabolic gene sets. In rodents, one important TH regulated step involves induction of Cyp7a1, an enzyme in the cytochrome P450 family, which enhances cholesterol to bile acid conversion and plays a crucial role in regulation of serum cholesterol levels. Current models suggest, however, that Cyp7a1 has lost the capacity to respond to THs in humans. We were prompted to re-examine TH effects on cholesterol metabolic genes in human liver cells by a recent study of a synthetic TH mimetic which showed that serum cholesterol reductions were accompanied by increases in a marker for bile acid synthesis in humans. Here, we show that TH effects upon cholesterol metabolic genes are almost identical in mouse liver, mouse and human liver primary cells and human hepatocyte cell lines. Moreover, Cyp7a1 is a direct TR target gene that responds to physiologic TR levels through a set of distinct response elements in its promoter. These findings suggest that THs regulate cholesterol to bile acid conversion in similar ways in humans and rodent experimental models and that manipulation of hormone signaling pathways could provide a strategy to enhance Cyp7a1 activity in human patients. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

The DSIF subunits Spt4 and Spt5 have distinct roles at various phases of immunoglobulin class switch recombination.

PubMed

Stanlie, Andre; Begum, Nasim A; Akiyama, Hideo; Honjo, Tasuku

2012-01-01

Class-switch recombination (CSR), induced by activation-induced cytidine deaminase (AID), can be divided into two phases: DNA cleavage of the switch (S) regions and the joining of the cleaved ends of the different S regions. Here, we show that the DSIF complex (Spt4 and Spt5), a transcription elongation factor, is required for CSR in a switch-proficient B cell line CH12F3-2A cells, and Spt4 and Spt5 carry out independent functions in CSR. While neither Spt4 nor Spt5 is required for transcription of S regions and AID, expression array analysis suggests that Spt4 and Spt5 regulate a distinct subset of transcripts in CH12F3-2A cells. Curiously, Spt4 is critically important in suppressing cryptic transcription initiating from the intronic Sμ region. Depletion of Spt5 reduced the H3K4me3 level and DNA cleavage at the Sα region, whereas Spt4 knockdown did not perturb the H3K4me3 status and S region cleavage. H3K4me3 modification level thus correlated well with the DNA breakage efficiency. Therefore we conclude that Spt5 plays a role similar to the histone chaperone FACT complex that regulates H3K4me3 modification and DNA cleavage in CSR. Since Spt4 is not involved in the DNA cleavage step, we suspected that Spt4 might be required for DNA repair in CSR. We examined whether Spt4 or Spt5 is essential in non-homologous end joining (NHEJ) and homologous recombination (HR) as CSR utilizes general repair pathways. Both Spt4 and Spt5 are required for NHEJ and HR as determined by assay systems using synthetic repair substrates that are actively transcribed even in the absence of Spt4 and Spt5. Taken together, Spt4 and Spt5 can function independently in multiple transcription-coupled steps of CSR.

The Atypical Response Regulator Protein ChxR Has Structural Characteristics and Dimer Interface Interactions That Are Unique within the OmpR/PhoB Subfamily

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

Hickey, John M.; Lovell, Scott; Battaile, Kevin P.

2013-05-29

Typically as a result of phosphorylation, OmpR/PhoB response regulators form homodimers through a receiver domain as an integral step in transcriptional activation. Phosphorylation stabilizes the ionic and hydrophobic interactions between monomers. Recent studies have shown that some response regulators retain functional activity in the absence of phosphorylation and are termed atypical response regulators. The two currently available receiver domain structures of atypical response regulators are very similar to their phospho-accepting homologs, and their propensity to form homodimers is generally retained. An atypical response regulator, ChxR, from Chlamydia trachomatis, was previously reported to form homodimers; however, the residues critical to thismore » interaction have not been elucidated. We hypothesize that the intra- and intermolecular interactions involved in forming a transcriptionally competent ChxR are distinct from the canonical phosphorylation (activation) paradigm in the OmpR/PhoB response regulator subfamily. To test this hypothesis, structural and functional studies were performed on the receiver domain of ChxR. Two crystal structures of the receiver domain were solved with the recently developed method using triiodo compound I3C. These structures revealed many characteristics unique to OmpR/PhoB subfamily members: typical or atypical. Included was the absence of two {alpha}-helices present in all other OmpR/PhoB response regulators. Functional studies on various dimer interface residues demonstrated that ChxR forms relatively stable homodimers through hydrophobic interactions, and disruption of these can be accomplished with the introduction of a charged residue within the dimer interface. A gel shift study with monomeric ChxR supports that dimerization through the receiver domain is critical for interaction with DNA.« less

Biotechnology regulation: is policy transfer an appropriate answer?

PubMed

Cárdenas-Gómez, Olga Carolina; Létourneau, Lyne

2010-01-01

In the world of biotechnology regulation, one often encounters the suggestion that the legislation of other countries should be consulted. Known as "policy transfer" in the field of public policy analysis, the purpose of such a recommendation is for policymakers to use the experiences of other States as a basis for developing appropriate regulatory frameworks in a timely manner. This paper examines whether policy transfer is relevant as an instrument for biotechnology regulation, and if it is, to what extent. Our analysis uses the example of Assisted Reproductive Technologies (ART), and unfolds according to the following argumentative steps. We will begin by discussing policy transfer as a recognized feature of policymaking in the literature pertaining to public policy analysis. We will then introduce a distinction between the technical dimension of policymaking and its political component. We will refer to "morality policy" as an illustration of policymaking directed toward its political component. We will show that, in the case of morality policy, States have moved away from a policy transfer approach. We will then establish that ART qualifies as morality policy, suggesting that policy transfer is most likely not the optimal policymaking tool for dealing with biotechnology regulation. Moving beyond the issue of ART in order to expand our reasoning to biotechnology regulation as a whole, we will conclude that, although the experiences of other States may be useful, policy transfer does not suffice in terms of informing policymaking in the case of biotechnology advances.

The Mediator co-activator complex regulates Ty1 retromobility by controlling the balance between Ty1i and Ty1 promoters.

PubMed

Salinero, Alicia C; Knoll, Elisabeth R; Zhu, Z Iris; Landsman, David; Curcio, M Joan; Morse, Randall H

2018-02-01

The Ty1 retrotransposons present in the genome of Saccharomyces cerevisiae belong to the large class of mobile genetic elements that replicate via an RNA intermediary and constitute a significant portion of most eukaryotic genomes. The retromobility of Ty1 is regulated by numerous host factors, including several subunits of the Mediator transcriptional co-activator complex. In spite of its known function in the nucleus, previous studies have implicated Mediator in the regulation of post-translational steps in Ty1 retromobility. To resolve this paradox, we systematically examined the effects of deleting non-essential Mediator subunits on the frequency of Ty1 retromobility and levels of retromobility intermediates. Our findings reveal that loss of distinct Mediator subunits alters Ty1 retromobility positively or negatively over a >10,000-fold range by regulating the ratio of an internal transcript, Ty1i, to the genomic Ty1 transcript. Ty1i RNA encodes a dominant negative inhibitor of Ty1 retromobility that blocks virus-like particle maturation and cDNA synthesis. These results resolve the conundrum of Mediator exerting sweeping control of Ty1 retromobility with only minor effects on the levels of Ty1 genomic RNA and the capsid protein, Gag. Since the majority of characterized intrinsic and extrinsic regulators of Ty1 retromobility do not appear to effect genomic Ty1 RNA levels, Mediator could play a central role in integrating signals that influence Ty1i expression to modulate retromobility.

Persistence of the cell-cycle checkpoint kinase Wee1 in SadA- and SadB-deficient neurons disrupts neuronal polarity.

PubMed

Müller, Myriam; Lutter, Daniela; Püschel, Andreas W

2010-01-15

Wee1 is well characterized as a cell-cycle checkpoint kinase that regulates the entry into mitosis in dividing cells. Here we identify a novel function of Wee1 in postmitotic neurons during the establishment of distinct axonal and dendritic compartments, which is an essential step during neuronal development. Wee1 is expressed in unpolarized neurons but is downregulated after neurons have extended an axon. Suppression of Wee1 impairs the formation of minor neurites but does not interfere with axon formation. However, neuronal polarity is disrupted when neurons fail to downregulate Wee1. The kinases SadA and SadB (Sad kinases) phosphorylate Wee1 and are required to initiate its downregulation in polarized neurons. Wee1 expression persists in neurons that are deficient in SadA and SadB and disrupts neuronal polarity. Knockdown of Wee1 rescues the Sada(-/-);Sadb(-/-) mutant phenotype and restores normal polarity in these neurons. Our results demonstrate that the regulation of Wee1 by SadA and SadB kinases is essential for the differentiation of polarized neurons.

Crystal structure of coelenterazine-binding protein from Renilla muelleri at 1.7 A: why it is not a calcium-regulated photoprotein.

PubMed

Stepanyuk, Galina A; Liu, Zhi-Jie; Markova, Svetlana S; Frank, Ludmila A; Lee, John; Vysotski, Eugene S; Wang, Bi-Cheng

2008-04-01

Bioluminescence in the sea pansy Renilla involves two distinct proteins, a Ca2+-triggered coelenterazine-binding protein (CBP), and Renilla luciferase. CBP contains one tightly bound coelenterazine molecule, which becomes available for reaction with luciferase and O2 only subsequent to Ca2+ binding. CBP belongs to the EF-hand superfamily of Ca2+-binding proteins and contains three "EF-hand" Ca2+-binding sites. The overall spatial structure of recombinant selenomethionine-labeled CBP determined at 1.7 A, is found to approximate the protein scaffold characteristic of the class of Ca2+-regulated photoproteins. Photoproteins however, catalyze molecular oxygen addition to coelenterazine producing a 2-hydroperoxycoelenterazine intermediate, which is stabilized within the binding cavity in the absence of Ca2+. Addition of Ca2+ triggers the bioluminescence reaction. However in CBP this first step of oxygen addition is not allowed. The different amino acid environments and hydrogen bond interactions within the binding cavity, are proposed to account for the different properties of the two classes of proteins.

An MRM-based workflow for quantifying cardiac mitochondrial protein phosphorylation in murine and human tissue.

PubMed

Lam, Maggie P Y; Scruggs, Sarah B; Kim, Tae-Young; Zong, Chenggong; Lau, Edward; Wang, Ding; Ryan, Christopher M; Faull, Kym F; Ping, Peipei

2012-08-03

The regulation of mitochondrial function is essential for cardiomyocyte adaptation to cellular stress. While it has long been understood that phosphorylation regulates flux through metabolic pathways, novel phosphorylation sites are continually being discovered in all functionally distinct areas of the mitochondrial proteome. Extracting biologically meaningful information from these phosphorylation sites requires an adaptable, sensitive, specific and robust method for their quantification. Here we report a multiple reaction monitoring-based mass spectrometric workflow for quantifying site-specific phosphorylation of mitochondrial proteins. Specifically, chromatographic and mass spectrometric conditions for 68 transitions derived from 23 murine and human phosphopeptides, and their corresponding unmodified peptides, were optimized. These methods enabled the quantification of endogenous phosphopeptides from the outer mitochondrial membrane protein VDAC, and the inner membrane proteins ANT and ETC complexes I, III and V. The development of this quantitative workflow is a pivotal step for advancing our knowledge and understanding of the regulatory effects of mitochondrial protein phosphorylation in cardiac physiology and pathophysiology. This article is part of a Special Issue entitled: Translational Proteomics. Copyright © 2012 Elsevier B.V. All rights reserved.

Study designs to investigate Nox1 acceleration of neoplastic progression in immortalized human epithelial cells by selection of differentiation resistant cells.

PubMed

Sattayakhom, Apsorn; Chunglok, Warangkana; Ittarat, Wanida; Chamulitrat, Walee

2014-01-01

To investigate the role of NADPH oxidase homolog Nox1 at an early step of cell transformation, we utilized human gingival mucosal keratinocytes immortalized by E6/E7 of human papillomavirus (HPV) type 16 (GM16) to generate progenitor cell lines either by chronic ethanol exposure or overexpression with Nox1. Among several cobblestone epithelial cell lines obtained, two distinctive spindle cell lines - FIB and NuB1 cells were more progressively transformed exhibiting tubulogenesis and anchorage-independent growth associated with increased invasiveness. These spindle cells acquired molecular markers of epithelial mesenchymal transition (EMT) including mesenchymal vimentin and simple cytokeratins (CK) 8 and 18 as well as myogenic alpha-smooth muscle actin and caldesmon. By overexpression and knockdown experiments, we showed that Nox1 on a post-translational level regulated the stability of CK18 in an ROS-, phosphorylation- and PKCepilon-dependent manner. PKCepilon may thus be used as a therapeutic target for EMT inhibition. Taken together, Nox1 accelerates neoplastic progression by regulating structural intermediate filaments leading to EMT of immortalized human gingival epithelial cells.

An HDAC2-TET1 switch at distinct chromatin regions significantly promotes the maturation of pre-iPS to iPS cells

PubMed Central

Wei, Tingyi; Chen, Wen; Wang, Xiukun; Zhang, Man; Chen, Jiayu; Zhu, Songcheng; Chen, Long; Yang, Dandan; Wang, Guiying; Jia, Wenwen; Yu, Yangyang; Duan, Tao; Wu, Minjuan; Liu, Houqi; Gao, Shaorong; Kang, Jiuhong

2015-01-01

The maturation of induced pluripotent stem cells (iPS) is one of the limiting steps of somatic cell reprogramming, but the underlying mechanism is largely unknown. Here, we reported that knockdown of histone deacetylase 2 (HDAC2) specifically promoted the maturation of iPS cells. Further studies showed that HDAC2 knockdown significantly increased histone acetylation, facilitated TET1 binding and DNA demethylation at the promoters of iPS cell maturation-related genes during the transition of pre-iPS cells to a fully reprogrammed state. We also found that HDAC2 competed with TET1 in the binding of the RbAp46 protein at the promoters of maturation genes and knockdown of TET1 markedly prevented the activation of these genes. Collectively, our data not only demonstrated a novel intrinsic mechanism that the HDAC2-TET1 switch critically regulates iPS cell maturation, but also revealed an underlying mechanism of the interplay between histone acetylation and DNA demethylation in gene regulation. PMID:25934799

Integrated Post-GWAS Analysis Sheds New Light on the Disease Mechanisms of Schizophrenia

PubMed Central

Lin, Jhih-Rong; Cai, Ying; Zhang, Quanwei; Zhang, Wen; Nogales-Cadenas, Rubén; Zhang, Zhengdong D.

2016-01-01

Schizophrenia is a severe mental disorder with a large genetic component. Recent genome-wide association studies (GWAS) have identified many schizophrenia-associated common variants. For most of the reported associations, however, the underlying biological mechanisms are not clear. The critical first step for their elucidation is to identify the most likely disease genes as the source of the association signals. Here, we describe a general computational framework of post-GWAS analysis for complex disease gene prioritization. We identify 132 putative schizophrenia risk genes in 76 risk regions spanning 120 schizophrenia-associated common variants, 78 of which have not been recognized as schizophrenia disease genes by previous GWAS. Even more significantly, 29 of them are outside the risk regions, likely under regulation of transcriptional regulatory elements contained therein. These putative schizophrenia risk genes are transcriptionally active in both brain and the immune system, and highly enriched among cellular pathways, consistent with leading pathophysiological hypotheses about the pathogenesis of schizophrenia. With their involvement in distinct biological processes, these putative schizophrenia risk genes, with different association strengths, show distinctive temporal expression patterns, and play specific biological roles during brain development. PMID:27754856

Differential utilization of decapping enzymes in mammalian mRNA decay pathways

PubMed Central

Li, You; Song, Mangen; Kiledjian, Megerditch

2011-01-01

mRNA decapping is a crucial step in the regulation of mRNA stability and gene expression. Dcp2 is an mRNA decapping enzyme that has been widely studied. We recently reported the presence of a second mammalian cytoplasmic decapping enzyme, Nudt16. Here we address the differential utilization of the two decapping enzymes in specified mRNA decay processes. Using mouse embryonic fibroblast (MEF) cell lines derived from a hypomorphic knockout of the Dcp2 gene with undetectable levels of Dcp2 or MEF cell lines harboring a Nudt16-directed shRNA to generate reduced levels of Nudt16, we demonstrate the distinct roles for Dcp2 and Nudt16 in nonsense-mediated mRNA decay (NMD), decay of ARE-containing mRNA and miRNA-mediated silencing. Our results indicated that NMD preferentially utilizes Dcp2 rather than Nudt16; Dcp2 and Nudt16 are redundant in miRNA-mediated silencing; and Dcp2 and Nudt16 are differentially utilized for ARE-mRNA decay. These data demonstrate that the two distinct decapping enzymes can uniquely function in specific mRNA decay processes in mammalian cells. PMID:21224379

41 CFR 102-74.590 - What steps must agencies take to implement these laws and policies?

Code of Federal Regulations, 2011 CFR

2011-01-01

... Management Federal Property Management Regulations System (Continued) FEDERAL MANAGEMENT REGULATION REAL PROPERTY 74-FACILITY MANAGEMENT Telework § 102-74.590 What steps must agencies take to implement these laws... 41 Public Contracts and Property Management 3 2011-01-01 2011-01-01 false What steps must agencies...

41 CFR 102-74.590 - What steps must agencies take to implement these laws and policies?

Code of Federal Regulations, 2012 CFR

2012-01-01

... Management Federal Property Management Regulations System (Continued) FEDERAL MANAGEMENT REGULATION REAL PROPERTY 74-FACILITY MANAGEMENT Telework § 102-74.590 What steps must agencies take to implement these laws... 41 Public Contracts and Property Management 3 2012-01-01 2012-01-01 false What steps must agencies...

41 CFR 102-74.590 - What steps must agencies take to implement these laws and policies?

Code of Federal Regulations, 2014 CFR

2014-01-01

... Management Federal Property Management Regulations System (Continued) FEDERAL MANAGEMENT REGULATION REAL PROPERTY 74-FACILITY MANAGEMENT Telework § 102-74.590 What steps must agencies take to implement these laws... 41 Public Contracts and Property Management 3 2014-01-01 2014-01-01 false What steps must agencies...

41 CFR 102-74.590 - What steps must agencies take to implement these laws and policies?

Code of Federal Regulations, 2013 CFR

2013-07-01

... Management Federal Property Management Regulations System (Continued) FEDERAL MANAGEMENT REGULATION REAL PROPERTY 74-FACILITY MANAGEMENT Telework § 102-74.590 What steps must agencies take to implement these laws... 41 Public Contracts and Property Management 3 2013-07-01 2013-07-01 false What steps must agencies...

15 CFR 732.5 - Steps regarding Shipper's Export Declaration or Automated Export System record, Destination...

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Steps regarding Shipper's Export... Section 732.5 Commerce and Foreign Trade Regulations Relating to Commerce and Foreign Trade (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS STEPS FOR USING...

Do lightning positive leaders really "step"?

NASA Astrophysics Data System (ADS)

Petersen, D.

2015-12-01

It has been known for some time that positive leaders exhibit impulsive charge motion and optical emissions as they extend. However, laboratory and field observations have not produced any evidence of a process analogous to the space leader mechanism of negative leader extension. Instead, observations have suggested that the positive leader tip undergoes a continuous to intermittent series of corona streamer bursts, each burst resulting in a small forward extension of the positive leader channel. Traditionally, it has been held that lightning positive leaders extend in a continuous or quasi-continuous fashion. Lately, however, many have become concerned that this position is incongruous with observations of impulsive activity during lightning positive leader extension. It is increasingly suggested that this impulsive activity is evidence that positive leaders also undergo "stepping". There are two issues that must be addressed. The first issue concerns whether or not the physical processes underlying impulsive extension in negative and positive leaders are distinct. We argue that these processes are in fact physically distinct, and offer new high-speed video evidence to support this position. The second issue regards the proper use of the term "step" as an identifier for the impulsive forward extension of a leader. Traditional use of this term has been applied only to negative leaders, due primarily to their stronger impulsive charge motions and photographic evidence of clearly discontinuous forward progression of the luminous channel. Recently, due to the increasing understanding of the distinct "space leader" process of negative leader extension, the term "step" has increasingly come to be associated with the space leader process itself. Should this emerging association, "step" = space leader attachment, be canonized? If not, then it seems reasonable to use the term "step" to describe impulsive positive leader extension. If, however, we do wish to associate the term "step" with space leader attachment, a process unique to negative leaders, should we devise a term for those process(es) that underly impulsive positive leader extension?

Chaperone expression profiles correlate with distinct physiological states of Plasmodium falciparum in malaria patients

PubMed Central

2010-01-01

Background Molecular chaperones have been shown to be important in the growth of the malaria parasite Plasmodium falciparum and inhibition of chaperone function by pharmacological agents has been shown to abrogate parasite growth. A recent study has demonstrated that clinical isolates of the parasite have distinct physiological states, one of which resembles environmental stress response showing up-regulation of specific molecular chaperones. Methods Chaperone networks operational in the distinct physiological clusters in clinical malaria parasites were constructed using cytoscape by utilizing their clinical expression profiles. Results Molecular chaperones show distinct profiles in the previously defined physiologically distinct states. Further, expression profiles of the chaperones from different cellular compartments correlate with specific patient clusters. While cluster 1 parasites, representing a starvation response, show up-regulation of organellar chaperones, cluster 2 parasites, which resemble active growth based on glycolysis, show up-regulation of cytoplasmic chaperones. Interestingly, cytoplasmic Hsp90 and its co-chaperones, previously implicated as drug targets in malaria, cluster in the same group. Detailed analysis of chaperone expression in the patient cluster 2 reveals up-regulation of the entire Hsp90-dependent pro-survival circuitries. In addition, cluster 2 also shows up-regulation of Plasmodium export element (PEXEL)-containing Hsp40s thought to have regulatory and host remodeling roles in the infected erythrocyte. Conclusion In all, this study demonstrates an intimate involvement of parasite-encoded chaperones, PfHsp90 in particular, in defining pathogenesis of malaria. PMID:20719001

The logistics of myelin biogenesis in the central nervous system.

PubMed

Snaidero, Nicolas; Simons, Mikael

2017-07-01

Rapid nerve conduction depends on myelin, but not all axons in the central nervous system (CNS) are myelinated to the same extent. Here, we review our current understanding of the biology of myelin biogenesis in the CNS. We focus on how the different steps of myelination are interconnected and how distinct patterns of myelin are generated. Possibly, a "basal" mode of myelination is laying the groundwork in areas devoted to basic homeostasis early in development, whereas a "targeted" mode generates myelin in regions controlling more complex tasks throughout adulthood. Such mechanisms may explain why myelination progresses in some areas according to a typical chronological and topographic sequence, while in other regions it is regulated by environmental stimuli contributing to interindividual variability of myelin structure. GLIA 2017;65:1021-1031. © 2017 Wiley Periodicals, Inc.

Structure of Mycobacterium tuberculosis phosphopantetheine adenylyltransferase in complex with the feedback inhibitor CoA reveals only one active-site conformation

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

Wubben, T.; Mesecar, A.D.; UIC)

Phosphopantetheine adenylyltransferase (PPAT) catalyzes the penultimate step in the coenzyme A (CoA) biosynthetic pathway, reversibly transferring an adenylyl group from ATP to 4'-phosphopantetheine to form dephosphocoenzyme A (dPCoA). To complement recent biochemical and structural studies on Mycobacterium tuberculosis PPAT (MtPPAT) and to provide further insight into the feedback regulation of MtPPAT by CoA, the X-ray crystal structure of the MtPPAT enzyme in complex with CoA was determined to 2.11 {angstrom} resolution. Unlike previous X-ray crystal structures of PPAT-CoA complexes from other bacteria, which showed two distinct CoA conformations bound to the active site, only one conformation of CoA is observedmore » in the MtPPAT-CoA complex.« less

Activity and growth efficiency of heterotrophic bacteria in a salt marsh (Ria de Aveiro, Portugal).

PubMed

Cunha, M A; Pedro, R; Almeida, M A; Silva, M H

2005-01-01

Bacterial utilization of monomers is recognized as an important step in the biogeochemical cycling of organic matter. In this study we have compared the heterotrophic activity of bacterial communities from different micro-habitats within a salt marsh environment (Ria de Aveiro, Portugal) in order to establish spatial patterns of bacterial abundance, monomer turnover rates (Tr) and bacterial growth efficiency (BGE). Differences in bacterial abundance and activity could be found between distinct plant rhizospheres. BGE tended to be lower at Halimione portulacoides banks, when compared to Sarcocornia perennis subsp. perennis banks which, on the contrary, showed the highest bacterial densities. Experiments of amendment of natural samples with organic and inorganic supplements indicated that salt marsh bacteria are not strongly regulated by salinity but the increased availability of labile organic matter causes a significant metabolic shift towards mineralization.

How to use and integrate bioinformatics tools to compare proteomic data from distinct conditions? A tutorial using the pathological similarities between Aortic Valve Stenosis and Coronary Artery Disease as a case-study.

PubMed

Trindade, Fábio; Ferreira, Rita; Magalhães, Beatriz; Leite-Moreira, Adelino; Falcão-Pires, Inês; Vitorino, Rui

2018-01-16

Nowadays we are surrounded by a plethora of bioinformatics tools, powerful enough to deal with the large amounts of data arising from proteomic studies, but whose application is sometimes hard to find. Therefore, we used a specific clinical problem - to discriminate pathophysiology and potential biomarkers between two similar cardiovascular diseases, aortic valve stenosis (AVS) and coronary artery disease (CAD) - to make a step-by-step guide through four bioinformatics tools: STRING, DisGeNET, Cytoscape and ClueGO. Proteome data was collected from articles available on PubMed centered on proteomic studies enrolling subjects with AVS or CAD. Through the analysis of gene ontology provided by STRING and ClueGO we could find specific biological phenomena associated with AVS, such as down-regulation of elastic fiber assembly, and with CAD, such as up-regulation of plasminogen activation. Moreover, through Cytoscape and DisGeNET we could pinpoint surrogate markers either for AVS (e.g. popeye domain containing protein 2 and 28S ribosomal protein S36, mitochondrial) or for CAD (e.g. ankyrin repeat and SOCS box protein 7) which deserve future validation. Data recycling and integration as well as research orientation are among the main advantages of resorting to bioinformatics analysis, hence these tutorials can be of great convenience for proteomics investigators. As we saw for aortic valve stenosis and coronary artery disease, it can be of great relevance to perform preliminary bioinformatics analysis with already published proteomics data. It not only saves us time in the lab (avoiding work duplication) as it points out new hypothesis to explain the phenotypical presentation of the diseases as well as new surrogate markers with clinical relevance, deserving future scrutiny. These essential steps can be easily overcome if one follows the steps proposed in our tutorial for STRING, DisGeNET, Cytoscape and ClueGO utilization. Copyright © 2017 Elsevier B.V. All rights reserved.

Coordination of the recruitment of the FANCD2 and PALB2 Fanconi anemia proteins by an ubiquitin signaling network.

PubMed

Bick, Gregory; Zhang, Fan; Meetei, A Ruhikanta; Andreassen, Paul R

2017-06-01

Fanconi anemia (FA) is a chromosome instability syndrome and the 20 identified FA proteins are organized into two main arms which are thought to function at distinct steps in the repair of DNA interstrand crosslinks (ICLs). These two arms include the upstream FA pathway, which culminates in the monoubiquitination of FANCD2 and FANCI, and downstream breast cancer (BRCA)-associated proteins that interact in protein complexes. How, and whether, these two groups of FA proteins are integrated is unclear. Here, we show that FANCD2 and PALB2, as indicators of the upstream and downstream arms, respectively, colocalize independently of each other in response to DNA damage induced by mitomycin C (MMC). We also show that ubiquitin chains are induced by MMC and colocalize with both FANCD2 and PALB2. Our finding that the RNF8 E3 ligase has a role in recruiting FANCD2 and PALB2 also provides support for the hypothesis that the two branches of the FA-BRCA pathway are coordinated by ubiquitin signaling. Interestingly, we find that the RNF8 partner, MDC1, as well as the ubiquitin-binding protein, RAP80, specifically recruit PALB2, while a different ubiquitin-binding protein, FAAP20, functions only in the recruitment of FANCD2. Thus, FANCD2 and PALB2 are not recruited in a single linear pathway, rather we define how their localization is coordinated and integrated by a network of ubiquitin-related proteins. We propose that such regulation may enable upstream and downstream FA proteins to act at distinct steps in the repair of ICLs.

Coordination of Hepatitis C Virus Assembly by Distinct Regulatory Regions in Nonstructural Protein 5A

PubMed Central

Zayas, Margarita; Long, Gang; Madan, Vanesa; Bartenschlager, Ralf

2016-01-01

Hepatitis C virus (HCV) nonstructural protein (NS)5A is a RNA-binding protein composed of a N-terminal membrane anchor, a structured domain I (DI) and two intrinsically disordered domains (DII and DIII) interacting with viral and cellular proteins. While DI and DII are essential for RNA replication, DIII is required for assembly. How these processes are orchestrated by NS5A is poorly understood. In this study, we identified a highly conserved basic cluster (BC) at the N-terminus of DIII that is critical for particle assembly. We generated BC mutants and compared them with mutants that are blocked at different stages of the assembly process: a NS5A serine cluster (SC) mutant blocked in NS5A-core interaction and a mutant lacking the envelope glycoproteins (ΔE1E2). We found that BC mutations did not affect core-NS5A interaction, but strongly impaired core–RNA association as well as virus particle envelopment. Moreover, BC mutations impaired RNA-NS5A interaction arguing that the BC might be required for loading of core protein with viral RNA. Interestingly, RNA-core interaction was also reduced with the ΔE1E2 mutant, suggesting that nucleocapsid formation and envelopment are coupled. These findings argue for two NS5A DIII determinants regulating assembly at distinct, but closely linked steps: (i) SC-dependent recruitment of replication complexes to core protein and (ii) BC-dependent RNA genome delivery to core protein, triggering encapsidation that is tightly coupled to particle envelopment. These results provide a striking example how a single viral protein exerts multiple functions to coordinate the steps from RNA replication to the assembly of infectious virus particles. PMID:26727512

Permeability and kinetic coefficients for mesoscale BCF surface step dynamics: Discrete two-dimensional deposition-diffusion equation analysis

DOE PAGES

Zhao, Renjie; Evans, James W.; Oliveira, Tiago J.

2016-04-08

Here, a discrete version of deposition-diffusion equations appropriate for description of step flow on a vicinal surface is analyzed for a two-dimensional grid of adsorption sites representing the stepped surface and explicitly incorporating kinks along the step edges. Model energetics and kinetics appropriately account for binding of adatoms at steps and kinks, distinct terrace and edge diffusion rates, and possible additional barriers for attachment to steps. Analysis of adatom attachment fluxes as well as limiting values of adatom densities at step edges for nonuniform deposition scenarios allows determination of both permeability and kinetic coefficients. Behavior of these quantities is assessedmore » as a function of key system parameters including kink density, step attachment barriers, and the step edge diffusion rate.« less

Permeability and kinetic coefficients for mesoscale BCF surface step dynamics: Discrete two-dimensional deposition-diffusion equation analysis

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

Zhao, Renjie; Evans, James W.; Oliveira, Tiago J.

Here, a discrete version of deposition-diffusion equations appropriate for description of step flow on a vicinal surface is analyzed for a two-dimensional grid of adsorption sites representing the stepped surface and explicitly incorporating kinks along the step edges. Model energetics and kinetics appropriately account for binding of adatoms at steps and kinks, distinct terrace and edge diffusion rates, and possible additional barriers for attachment to steps. Analysis of adatom attachment fluxes as well as limiting values of adatom densities at step edges for nonuniform deposition scenarios allows determination of both permeability and kinetic coefficients. Behavior of these quantities is assessedmore » as a function of key system parameters including kink density, step attachment barriers, and the step edge diffusion rate.« less

How do physicians become medical experts? A test of three competing theories: distinct domains, independent influence and encapsulation models.

PubMed

Violato, Claudio; Gao, Hong; O'Brien, Mary Claire; Grier, David; Shen, E

2018-05-01

The distinction between basic sciences and clinical knowledge which has led to a theoretical debate on how medical expertise is developed has implications for medical school and lifelong medical education. This longitudinal, population based observational study was conducted to test the fit of three theories-knowledge encapsulation, independent influence, distinct domains-of the development of medical expertise employing structural equation modelling. Data were collected from 548 physicians (292 men-53.3%; 256 women-46.7%; mean age = 24.2 years on admission) who had graduated from medical school 2009-2014. They included (1) Admissions data of undergraduate grade point average and Medical College Admission Test sub-test scores, (2) Course performance data from years 1, 2, and 3 of medical school, and (3) Performance on the NBME exams (i.e., Step 1, Step 2 CK, and Step 3). Statistical fit indices (Goodness of Fit Index-GFI; standardized root mean squared residual-SRMR; root mean squared error of approximation-RSMEA) and comparative fit [Formula: see text] of three theories of cognitive development of medical expertise were used to assess model fit. There is support for the knowledge encapsulation three factor model of clinical competency (GFI = 0.973, SRMR = 0.043, RSMEA = 0.063) which had superior fit indices to both the independent influence and distinct domains theories ([Formula: see text] vs [Formula: see text] [[Formula: see text

Picking Deep Filter Responses for Fine-Grained Image Recognition (Open Access Author’s Manuscript)

DTIC Science & Technology

2016-12-16

stages. Our method explores a unified framework based on two steps of deep filter response picking. The first picking step is to find distinctive... filters which respond to specific patterns significantly and consistently, and learn a set of part detectors via iteratively alternating between new...positive sample mining and part model retraining. The second picking step is to pool deep filter responses via spatially weighted combination of Fisher

Dynamical Scaling Implications of Ferrari, Prähofer, and Spohn's Remarkable Spatial Scaling Results for Facet-Edge Fluctuations

NASA Astrophysics Data System (ADS)

Einstein, T. L.; Pimpinelli, Alberto

2014-06-01

Spurred by theoretical predictions from Ferrari et al. (Phys Rev E 69:035102(R), 2004), we rederived and extended their result heuristically. With experimental colleagues we used STM line scans to corroborate their prediction that the fluctuations of the step bounding a facet exhibit scaling properties distinct from those of isolated steps or steps on vicinal surfaces. The correlation functions was shown to go as , decidedly different from the behavior for fluctuations of isolated steps.

Differences in the Regulation of K-Ras and H-Ras Isoforms by Monoubiquitination*

PubMed Central

Baker, Rachael; Wilkerson, Emily M.; Sumita, Kazutaka; Isom, Daniel G.; Sasaki, Atsuo T.; Dohlman, Henrik G.; Campbell, Sharon L.

2013-01-01

Ras GTPases are signaling switches that control critical cellular processes including gene expression, differentiation, and apoptosis. The major Ras isoforms (K, H, and N) contain a conserved core GTPase domain, but have distinct biological functions. Among the three Ras isoforms there are clear differences in post-translational regulation, which contribute to differences in localization and signaling output. Modification by ubiquitination was recently reported to activate Ras signaling in cells, but the mechanisms of activation are not well understood. Here, we show that H-Ras is activated by monoubiquitination and that ubiquitination at Lys-117 accelerates intrinsic nucleotide exchange, thereby promoting GTP loading. This mechanism of Ras activation is distinct from K-Ras monoubiquitination at Lys-147, which leads to impaired regulator-mediated GTP hydrolysis. These findings reveal that different Ras isoforms are monoubiquitinated at distinct sites, with distinct mechanisms of action, but with a common ability to chronically activate the protein in the absence of a receptor signal or oncogenic mutation. PMID:24247240

Zn2+-dependent Activation of the Trk Signaling Pathway Induces Phosphorylation of the Brain-enriched Tyrosine Phosphatase STEP

PubMed Central

Poddar, Ranjana; Rajagopal, Sathyanarayanan; Shuttleworth, C. William; Paul, Surojit

2016-01-01

Excessive release of Zn2+ in the brain is implicated in the progression of acute brain injuries. Although several signaling cascades have been reported to be involved in Zn2+-induced neurotoxicity, a potential contribution of tyrosine phosphatases in this process has not been well explored. Here we show that exposure to high concentrations of Zn2+ led to a progressive increase in phosphorylation of the striatal-enriched phosphatase (STEP), a component of the excitotoxic-signaling pathway that plays a role in neuroprotection. Zn2+-mediated phosphorylation of STEP61 at multiple sites (hyperphosphorylation) was induced by the up-regulation of brain-derived neurotropic factor (BDNF), tropomyosin receptor kinase (Trk) signaling, and activation of cAMP-dependent PKA (protein kinase A). Mutational studies further show that differential phosphorylation of STEP61 at the PKA sites, Ser-160 and Ser-221 regulates the affinity of STEP61 toward its substrates. Consistent with these findings we also show that BDNF/Trk/PKA mediated signaling is required for Zn2+-induced phosphorylation of extracellular regulated kinase 2 (ERK2), a substrate of STEP that is involved in Zn2+-dependent neurotoxicity. The strong correlation between the temporal profile of STEP61 hyperphosphorylation and ERK2 phosphorylation indicates that loss of function of STEP61 through phosphorylation is necessary for maintaining sustained ERK2 phosphorylation. This interpretation is further supported by the findings that deletion of the STEP gene led to a rapid and sustained increase in ERK2 phosphorylation within minutes of exposure to Zn2+. The study provides further insight into the mechanisms of regulation of STEP61 and also offers a molecular basis for the Zn2+-induced sustained activation of ERK2. PMID:26574547

The Mediator co-activator complex regulates Ty1 retromobility by controlling the balance between Ty1i and Ty1 promoters

PubMed Central

Salinero, Alicia C.; Knoll, Elisabeth R.; Zhu, Z. Iris

2018-01-01

The Ty1 retrotransposons present in the genome of Saccharomyces cerevisiae belong to the large class of mobile genetic elements that replicate via an RNA intermediary and constitute a significant portion of most eukaryotic genomes. The retromobility of Ty1 is regulated by numerous host factors, including several subunits of the Mediator transcriptional co-activator complex. In spite of its known function in the nucleus, previous studies have implicated Mediator in the regulation of post-translational steps in Ty1 retromobility. To resolve this paradox, we systematically examined the effects of deleting non-essential Mediator subunits on the frequency of Ty1 retromobility and levels of retromobility intermediates. Our findings reveal that loss of distinct Mediator subunits alters Ty1 retromobility positively or negatively over a >10,000-fold range by regulating the ratio of an internal transcript, Ty1i, to the genomic Ty1 transcript. Ty1i RNA encodes a dominant negative inhibitor of Ty1 retromobility that blocks virus-like particle maturation and cDNA synthesis. These results resolve the conundrum of Mediator exerting sweeping control of Ty1 retromobility with only minor effects on the levels of Ty1 genomic RNA and the capsid protein, Gag. Since the majority of characterized intrinsic and extrinsic regulators of Ty1 retromobility do not appear to effect genomic Ty1 RNA levels, Mediator could play a central role in integrating signals that influence Ty1i expression to modulate retromobility. PMID:29462141

Gender difference in older adult's utilization of gravitational and ground reaction force in regulation of angular momentum during stair descent.

PubMed

Singhal, Kunal; Kim, Jemin; Casebolt, Jeffrey; Lee, Sangwoo; Han, Ki-Hoon; Kwon, Young-Hoo

2015-06-01

Angular momentum of the body is a highly controlled quantity signifying stability, therefore, it is essential to understand its regulation during stair descent. The purpose of this study was to investigate how older adults use gravity and ground reaction force to regulate the angular momentum of the body during stair descent. A total of 28 participants (12 male and 16 female; 68.5 years and 69.0 years of mean age respectively) performed stair descent from a level walk in a step-over-step manner at a self-selected speed over a custom made three-step staircase with embedded force plates. Kinematic and force data were used to calculate angular momentum, gravitational moment, and ground reaction force moment about the stance foot center of pressure. Women show a significantly greater change in normalized angular momentum (0.92Nms/Kgm; p=.004) as compared to men (0.45Nms/Kgm). Women produce higher normalized GRF (p=.031) during the double support phase. The angular momentum changes show largest backward regulation for Step 0 and forward regulation for Step 2. This greater difference in overall change in the angular momentum in women may explain their increased risk of fall over the stairs. Copyright © 2015 Elsevier B.V. All rights reserved.

PP32 and SET/TAF-Iβ proteins regulate the acetylation of newly synthesized histone H4

PubMed Central

Saavedra, Francisco; Rivera, Carlos; Rivas, Elizabeth; Merino, Paola; Garrido, Daniel; Hernández, Sergio; Forné, Ignasi; Vassias, Isabelle; Gurard-Levin, Zachary A.; Alfaro, Iván E.; Imhof, Axel; Almouzni, Geneviève

2017-01-01

Abstract Newly synthesized histones H3 and H4 undergo a cascade of maturation steps to achieve proper folding and to establish post-translational modifications prior to chromatin deposition. Acetylation of H4 on lysines 5 and 12 by the HAT1 acetyltransferase is observed late in the histone maturation cascade. A key question is to understand how to establish and regulate the distinct timing of sequential modifications and their biological significance. Here, we perform proteomic analysis of the newly synthesized histone H4 complex at the earliest time point in the cascade. In addition to known binding partners Hsp90 and Hsp70, we also identify for the first time two subunits of the histone acetyltransferase inhibitor complex (INHAT): PP32 and SET/TAF-Iβ. We show that both proteins function to prevent HAT1-mediated H4 acetylation in vitro. When PP32 and SET/TAF-Iβ protein levels are down-regulated in vivo, we detect hyperacetylation on lysines 5 and 12 and other H4 lysine residues. Notably, aberrantly acetylated H4 is less stable and this reduces the interaction with Hsp90. As a consequence, PP32 and SET/TAF-Iβ depleted cells show an S-phase arrest. Our data demonstrate a novel function of PP32 and SET/TAF-Iβ and provide new insight into the mechanisms regulating acetylation of newly synthesized histone H4. PMID:28977641

PP32 and SET/TAF-Iβ proteins regulate the acetylation of newly synthesized histone H4.

PubMed

Saavedra, Francisco; Rivera, Carlos; Rivas, Elizabeth; Merino, Paola; Garrido, Daniel; Hernández, Sergio; Forné, Ignasi; Vassias, Isabelle; Gurard-Levin, Zachary A; Alfaro, Iván E; Imhof, Axel; Almouzni, Geneviève; Loyola, Alejandra

2017-11-16

Newly synthesized histones H3 and H4 undergo a cascade of maturation steps to achieve proper folding and to establish post-translational modifications prior to chromatin deposition. Acetylation of H4 on lysines 5 and 12 by the HAT1 acetyltransferase is observed late in the histone maturation cascade. A key question is to understand how to establish and regulate the distinct timing of sequential modifications and their biological significance. Here, we perform proteomic analysis of the newly synthesized histone H4 complex at the earliest time point in the cascade. In addition to known binding partners Hsp90 and Hsp70, we also identify for the first time two subunits of the histone acetyltransferase inhibitor complex (INHAT): PP32 and SET/TAF-Iβ. We show that both proteins function to prevent HAT1-mediated H4 acetylation in vitro. When PP32 and SET/TAF-Iβ protein levels are down-regulated in vivo, we detect hyperacetylation on lysines 5 and 12 and other H4 lysine residues. Notably, aberrantly acetylated H4 is less stable and this reduces the interaction with Hsp90. As a consequence, PP32 and SET/TAF-Iβ depleted cells show an S-phase arrest. Our data demonstrate a novel function of PP32 and SET/TAF-Iβ and provide new insight into the mechanisms regulating acetylation of newly synthesized histone H4. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Distinct roles for Arp2/3 regulators in actin assembly and endocytosis.

PubMed

Galletta, Brian J; Chuang, Dennis Y; Cooper, John A

2008-01-01

The Arp2/3 complex is essential for actin assembly and motility in many cell processes, and a large number of proteins have been found to bind and regulate it in vitro. A critical challenge is to understand the actions of these proteins in cells, especially in settings where multiple regulators are present. In a systematic study of the sequential multicomponent actin assembly processes that accompany endocytosis in yeast, we examined and compared the roles of WASp, two type-I myosins, and two other Arp2/3 activators, along with that of coronin, which is a proposed inhibitor of Arp2/3. Quantitative analysis of high-speed fluorescence imaging revealed individual functions for the regulators, manifested in part by novel phenotypes. We conclude that Arp2/3 regulators have distinct and overlapping roles in the processes of actin assembly that drive endocytosis in yeast. The formation of the endocytic actin patch, the creation of the endocytic vesicle, and the movement of the vesicle into the cytoplasm display distinct dependencies on different Arp2/3 regulators. Knowledge of these roles provides insight into the in vivo relevance of the dendritic nucleation model for actin assembly.

Indicator Expansion with Analysis Pipeline

DTIC Science & Technology

2015-01-13

INTERNAL FILTER trackInfectedHosts FILTER badTraffic SIP infectedHosts 1 DAY END INTERNAL FILTER 11 Step 3 watch where infected hosts go FILTER...nonWhiteListPostInfected SIP IN LIST infectedHosts DIP NOT IN LIST safePopularIPs.set END FILTER 12 Step 4 & 5: Count Hosts Per IP and Alert EVALUATION...CHECK THRESHOLD DISTINCT SIP > 50 TIME WINDOW 36 HOURS END CHECK END EVALUATION 13 Step 6: Report Expanded Indicators LIST CONFIGURATION secondLevelIPs

Distinct neural processes are engaged in the modulation of mimicry by social group-membership and emotional expressions.

PubMed

Rauchbauer, Birgit; Majdandžić, Jasminka; Hummer, Allan; Windischberger, Christian; Lamm, Claus

2015-09-01

People often spontaneously engage in copying each other's postures and mannerisms, a phenomenon referred to as behavioral mimicry. Social psychology experiments indicate that mimicry denotes an implicit affiliative signal flexibly regulated in response to social requirements. Yet, the mediating processes and neural underpinnings of such regulation are largely unexplored. The present functional magnetic resonance imaging (fMRI) study examined mimicry regulation by combining an automatic imitation task with facial stimuli, varied on two social-affective dimensions: emotional expression (angry vs happy) and ethnic group membership (in- vs out-group). Behavioral data revealed increased mimicry when happy and when out-group faces were shown. Imaging results revealed that mimicry regulation in response to happy faces was associated with increased activation in the right temporo-parietal junction (TPJ), right dorsal premotor cortex (dPMC), and right superior parietal lobule (SPL). Mimicry regulation in response to out-group faces was related to increased activation in the left ventral premotor cortex (vPMC) and inferior parietal lobule (IPL), bilateral anterior insula, and mid-cingulate cortex (MCC). We suggest that mimicry in response to happy and to out-group faces is driven by distinct affiliative goals, and that mimicry regulation to attain these goals is mediated by distinct neuro-cognitive processes. Higher mimicry in response to happy faces seems to denote reciprocation of an affiliative signal. Higher mimicry in response to out-group faces, reflects an appeasement attempt towards an interaction partner perceived as threatening (an interpretation supported by implicit measures showing that out-group members are more strongly associated with threat). Our findings show that subtle social cues can result in the implicit regulation of mimicry. This regulation serves to achieve distinct affiliative goals, is mediated by different regulatory processes, and relies on distinct parts of an overarching network of task-related brain areas. Our findings shed new light on the neural mechanisms underlying the interplay between implicit action control and social cognition. Copyright © 2015 Elsevier Ltd. All rights reserved.

Atomic friction at exposed and buried graphite step edges: Experiments and simulations

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

Ye, Zhijiang; Martini, Ashlie, E-mail: amartini@ucmerced.edu

2015-06-08

The surfaces of layered materials such as graphite exhibit step edges that affect friction. Step edges can be exposed, where the step occurs at the outmost layer, or buried, where the step is underneath another layer of material. Here, we study friction at exposed and buried step edges on graphite using an atomic force microscope (AFM) and complementary molecular dynamics simulations of the AFM tip apex. Exposed and buried steps exhibit distinct friction behavior, and the friction on either step is affected by the direction of sliding, i.e., moving up or down the step, and the bluntness of the tip.more » These trends are analyzing in terms of the trajectory of the AFM tip as it moves over the step, which is a convolution of the topography of the surface and the tip shape.« less

48 CFR 3036.104-90 - Authority for one-step turn-key design-build contracting for the United States Coast Guard (USCG).

Code of Federal Regulations, 2014 CFR

2014-10-01

... 48 Federal Acquisition Regulations System 7 2014-10-01 2014-10-01 false Authority for one-step turn-key design-build contracting for the United States Coast Guard (USCG). 3036.104-90 Section 3036.104-90 Federal Acquisition Regulations System DEPARTMENT OF HOMELAND SECURITY, HOMELAND SECURITY ACQUISITION REGULATION (HSAR) SPECIAL CATEGORIES OF...

Endochondral bone formation in embryonic mouse pre-metatarsals

NASA Technical Reports Server (NTRS)

Klement, B. J.; Spooner, B. S.

1992-01-01

Long term exposure to a reduced gravitational environment has a deleterious effect on bone. The developmental events which occur prior to initial bone deposition will provide insight into the regulation of mature bone physiology. We have characterized a system in which the events preceding bone formation take place in an isolated in vitro organ culture environment. We show that cultured pre-metatarsal tissue parallels development of pre-metatarsal tissue in the embryo. Both undergo mesenchyme differentiation and morphogenesis to form a cartilage rod, which resembles the future bone, followed by terminal chondrocyte differentiation in a definite morphogenetic pattern. These sequential steps occur prior to osteoblast maturation and bone matrix deposition in the developing organism. Alkaline phosphatase (ALP) activity is a distinctive enzymatic marker for mineralizing tissues. We have measured this activity throughout pre-metatarsal development and show (a) where in the tissue it is predominantly found, and (b) that this is indeed the mineralizing isoform of the enzyme.

Body self. Development, psychopathologies, and psychoanalytic significance.

PubMed

Krueger, D W

2001-01-01

Ego development or, more broadly, the sense of self has at its core a cohesive, distinct, and accurate body self. Compromise of body self development as a result of early overstimulation, empathic unavailability or nonresponse of the caretaker, and inconsistency or selectivity of response can lead to specific developmental arrests, including body-image distortions, nonintegration of body self and psychological self, and difficulties in the regulation of tension states and affect. The individual may then attempt to repair those disrupted developmental needs by such symptomatic expressions as eating disorders, compulsive exercise, substance abuse, and the creation of physical danger, as a step toward integration of mind and body as well as a defensive antidote to painful affect. In the psychoanalytic treatment of these patients, the need for the analyst's attunement to the patient's development of body self as well as psychological self development is illustrated by clinical vignettes of the enactments and attempted restitution of specific developmental trauma.

Hormonal control of implantation.

PubMed

Sandra, Olivier

2016-06-01

In mammals, implantation represents a key step of pregnancy and its progression conditions not only the success of pregnancy but health of the offspring. Implantation requires a complex and specific uterine tissue, the endometrium, whose biological functions are tightly regulated by numerous signals, including steroids and polypeptide hormones. Endometrial tissue is endowed with dynamic properties that associate its ability to control the developmental trajectory of the embryo (driver property) and its ability to react to embryos displaying distinct capacities to develop to term (sensor property). Since dynamical properties of the endometrium can be affected by pre- and post-conceptional environment, determining how maternal hormonal signals and their biological actions are affected by environmental factors (e.g. nutrition, stress, infections) is mandatory to reduce or even to prevent their detrimental effects on endometrial physiology in order to preserve the optimal functionality of this tissue. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Neutrophils suppress intraluminal NK-mediated tumor cell clearance and enhance extravasation of disseminated carcinoma cells

PubMed Central

Spiegel, Asaf; Brooks, Mary W.; Houshyar, Samin; Reinhardt, Ferenc; Ardolino, Michele; Fessler, Evelyn; Chen, Michelle B.; Krall, Jordan A.; DeCock, Jasmine; Zervantonakis, Ioannis K.; Iannello, Alexandre; Iwamoto, Yoshiko; Cortez-Retamozo, Virna; Kamm, Roger D.; Pittet, Mikael J.; Raulet, David H.; Weinberg, Robert A.

2016-01-01

Immune cells promote the initial metastatic dissemination of carcinoma cells from primary tumors. In contrast to their well-studied functions in the initial stages of metastasis, the specific roles of immunocytes in facilitating progression through the critical later steps of the invasion-metastasis cascade remain poorly understood. Here we define novel functions of neutrophils in promoting intraluminal survival and extravasation at sites of metastatic dissemination. We show that CD11b+/Ly6G+ neutrophils enhance metastasis formation via two distinct mechanisms. First, neutrophils inhibit natural killer cell function, which leads to a significant increase in the intraluminal survival time of tumor cells. Thereafter, neutrophils operate to facilitate extravasation of tumor cells through the secretion of IL-1β and matrix metalloproteinases. These results identify neutrophils as key regulators of intraluminal survival and extravasation through their crosstalk with host cells and disseminating carcinoma cells. PMID:27072748

Growth-dependent regulation of rRNA synthesis is mediated by a transcription initiation factor (TIF-IA).

PubMed

Buttgereit, D; Pflugfelder, G; Grummt, I

1985-11-25

Mouse RNA polymerase I requires at least two chromatographically distinct transcription factors (designated TIF-IA and TIF-IB) to initiate transcription accurately and efficiently in vitro. In this paper we describe the partial purification of TIF-IA by a four-step fractionation procedure. The amount or activity of TIF-IA fluctuates in response to the physiological state of the cells. Extracts from quiescent cells are incapable of specific transcription and do not contain detectable levels of TIF-IA. Transcriptionally inactive extracts can be restored by the addition of TIF-IA preparations that have been highly purified from exponentially growing cells. During the fractionating procedure TIF-IA co-purifies with RNA polymerase I, suggesting that it is functionally associated with the transcribing enzyme. We suggest that only those enzyme molecules that are associated with TIF-IA are capable to interact with TIF-IB and to initiate transcription.

Spatial and temporal controls target pal-1 blastomere-specification activity to a single blastomere lineage in C. elegans embryos.

PubMed

Hunter, C P; Kenyon, C

1996-10-18

The early asymmetric cleavages of Caenorhabditis elegans embryos produce blastomeres with distinct developmental potentials. Here, we show that the caudal-like homeodomain protein PAL-1 is required to specify the somatic identity of one posterior blastomere in the 4 cell embryo. We find that pal-1 activity is sequentially restricted to this blastomere. First, at the 4 cell stage, it is translated only in the two posterior blastomeres. Then, its function is restricted to one of these blastomeres. This second targeting step is dependent on the activities of the posteriorly localized SKN-1 and asymmetrically segregated PIE-1 proteins. We propose that the segregation of PIE-1, combined with the temporal decay of SKN-1, targets pal-1 activity to this posterior lineage, thus coupling the regulation of this conserved posterior patterning gene to asymmetric cell cleavages.

Targeted knockout in Physcomitrella reveals direct actions of phytochrome in the cytoplasm.

PubMed

Mittmann, Franz; Brücker, Gerhard; Zeidler, Mathias; Repp, Alexander; Abts, Thomas; Hartmann, Elmar; Hughes, Jon

2004-09-21

The plant photoreceptor phytochrome plays an important role in the nucleus as a regulator of transcription. Numerous studies imply, however, that phytochromes in both higher and lower plants mediate physiological reactions within the cytoplasm. In particular, the tip cells of moss protonemal filaments use phytochrome to sense light direction, requiring a signaling system that transmits the directional information directly to the microfilaments that direct tip growth. In this work we describe four canonical phytochrome genes in the model moss species Physcomitrella patens, each of which was successfully targeted via homologous recombination and the distinct physiological functions of each gene product thereby identified. One homolog in particular mediates positive phototropism, polarotropism, and chloroplast movement in polarized light. This photoreceptor thus interacts with a cytoplasmic signal/response system. This is our first step in elucidating the cytoplasmic signaling function of phytochrome at the molecular level.

Tailored logistics: the next advantage.

PubMed

Fuller, J B; O'Conor, J; Rawlinson, R

1993-01-01

How many top executives have ever visited with managers who move materials from the factory to the store? How many still reduce the costs of logistics to the rent of warehouses and the fees charged by common carriers? To judge by hours of senior management attention, logistics problems do not rank high. But logistics have the potential to become the next governing element of strategy. Whether they know it or not, senior managers of every retail store and diversified manufacturing company compete in logistically distinct businesses. Customer needs vary, and companies can tailor their logistics systems to serve their customers better and more profitably. Companies do not create value for customers and sustainable advantage for themselves merely by offering varieties of goods. Rather, they offer goods in distinct ways. A particular can of Coca-Cola, for example, might be a can of Coca-Cola going to a vending machine, or a can of Coca-Cola that comes with billing services. There is a fortune buried in this distinction. The goal of logistics strategy is building distinct approaches to distinct groups of customers. The first step is organizing a cross-functional team to proceed through the following steps: segmenting customers according to purchase criteria, establishing different standards of service for different customer segments, tailoring logistics pipelines to support each segment, and creating economics of scale to determine which assets can be shared among various pipelines. The goal of establishing logistically distinct businesses is familiar: improved knowledge of customers and improved means of satisfying them.

Preschool Teachers' Skills in Teaching Music: Two Steps Forward One Step Back

ERIC Educational Resources Information Center

Ehrlin, Anna; Wallerstedt, Cecilia

2014-01-01

This study investigates through observations and interviews what importance further education has for preschool teachers' practice in two music-profiled preschool and their way of conceptualising it. A distinction between music as a method for teaching, on the one hand, and as a content of knowledge, on the other, is used in the analysis. The…

48 CFR 52.214-25 - Step Two of Two-Step Sealed Bidding.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 2 2010-10-01 2010-10-01 false Step Two of Two-Step... Clauses 52.214-25 Step Two of Two-Step Sealed Bidding. As prescribed in 14.201-6(t), insert the following provision: Step Two of Two-Step Sealed Bidding (APR 1985) (a) This invitation for bids is issued to initiate...

14 CFR 1252.411 - Age distinctions.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 14 Aeronautics and Space 5 2013-01-01 2013-01-01 false Age distinctions. 1252.411 Section 1252.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION NONDISCRIMINATION ON THE BASIS OF AGE IN... Procedures § 1252.411 Age distinctions. There are no Federal statutes or regulations containing age...

14 CFR 1252.411 - Age distinctions.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 14 Aeronautics and Space 5 2011-01-01 2010-01-01 true Age distinctions. 1252.411 Section 1252.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION NONDISCRIMINATION ON THE BASIS OF AGE IN... Procedures § 1252.411 Age distinctions. There are no Federal statutes or regulations containing age...

14 CFR 1252.411 - Age distinctions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 14 Aeronautics and Space 5 2010-01-01 2010-01-01 false Age distinctions. 1252.411 Section 1252.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION NONDISCRIMINATION ON THE BASIS OF AGE IN... Procedures § 1252.411 Age distinctions. There are no Federal statutes or regulations containing age...

14 CFR 1252.411 - Age distinctions.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 14 Aeronautics and Space 5 2012-01-01 2012-01-01 false Age distinctions. 1252.411 Section 1252.411 Aeronautics and Space NATIONAL AERONAUTICS AND SPACE ADMINISTRATION NONDISCRIMINATION ON THE BASIS OF AGE IN... Procedures § 1252.411 Age distinctions. There are no Federal statutes or regulations containing age...

Polyamines: naturally occurring small molecule modulators of electrostatic protein-protein interactions.

PubMed

Berwanger, Anja; Eyrisch, Susanne; Schuster, Inge; Helms, Volkhard; Bernhardt, Rita

2010-02-01

Modulations of protein-protein interactions are a key step in regulating protein function, especially in networks. Modulators of these interactions are supposed to be candidates for the development of novel drugs. Here, we describe the role of the small, polycationic and highly abundant natural polyamines that could efficiently bind to charged spots at protein interfaces as modulators of such protein-protein interactions. Using the mitochondrial cytochrome P45011A1 (CYP11A1) electron transfer system as a model, we have analyzed the capability of putrescine, spermidine, and spermine at physiologically relevant concentrations to affect the protein-protein interactions between adrenodoxin reductase (AdR), adrenodoxin (Adx), and CYP11A1. The actions of polyamines on the individual components, on their association/dissociation, on electron transfer, and on substrate conversion were examined. These studies revealed modulating effects of polyamines on distinct interactions and on the entire system in a complex way. Modulation via changed protein-protein interactions appeared plausible from docking experiments that suggested favourable high-affinity binding sites of polyamines (spermine>spermidine>putrescine) at the AdR-Adx interface. Our findings imply for the first time that small endogenous compounds are capable of interfering with distinct components of transient protein complexes and might control protein functions by modulating electrostatic protein-protein interactions.

Identification of Ser-543 as the major regulatory phosphorylation site in spinach leaf nitrate reductase

NASA Technical Reports Server (NTRS)

Bachmann, M.; Shiraishi, N.; Campbell, W. H.; Yoo, B. C.; Harmon, A. C.; Huber, S. C.; Davies, E. (Principal Investigator)

1996-01-01

Spinach leaf NADH:nitrate reductase (NR) responds to light/dark signals and photosynthetic activity in part as a result of rapid regulation by reversible protein phosphorylation. We have identified the major regulatory phosphorylation site as Ser-543, which is located in the hinge 1 region connecting the cytochrome b domain with the molybdenum-pterin cofactor binding domain of NR, using recombinant NR fragments containing or lacking the phosphorylation site sequence. Studies with NR partial reactions indicated that the block in electron flow caused by phosphorylation also could be localized to the hinge 1 region. A synthetic peptide (NR6) based on the phosphorylation site sequence was phosphorylated readily by NR kinase (NRk) in vitro. NR6 kinase activity tracked the ATP-dependent inactivation of NR during several chromatographic steps and completely inhibited inactivation/phosphorylation of native NR in vitro. Two forms of NRk were resolved by using anion exchange chromatography. Studies with synthetic peptide analogs indicated that both forms of NRk had similar specificity determinants, requiring a basic residue at P-3 (i.e., three amino acids N-terminal to the phosphorylated serine) and a hydrophobic residue at P-5. Both forms are strictly calcium dependent but belong to distinct families of protein kinases because they are distinct immunochemically.

Small G proteins in insulin action: Rab and Rho families at the crossroads of signal transduction and GLUT4 vesicle traffic.

PubMed

Ishikura, S; Koshkina, A; Klip, A

2008-01-01

Insulin stimulates glucose uptake into muscle and adipose tissues through glucose transporter 4 (GLUT4). GLUT4 cycles between the intracellular compartments and the plasma membrane. GLUT4 traffic-regulating insulin signals are largely within the insulin receptor-insulin receptor substrate-phosphatidylinositol 3-kinase (IR-IRS-PI3K) axis. In muscle cells, insulin signal bifurcates downstream of the PI3K into one arm leading to the activation of the Ser/Thr kinases Akt and atypical protein kinase C, and another leading to the activation of Rho family protein Rac1 leading to actin remodelling. Activated Akt inactivates AS160, a GTPase-activating protein for Rab family small G proteins. Here we review the roles of Rab and Rho proteins, particularly Rab substrates of AS160 and Rac1, in insulin-stimulated GLUT4 traffic. We discuss: (1) how distinct steps in GLUT4 traffic may be regulated by discrete Rab proteins, and (2) the importance of Rac1 activation in insulin-induced actin remodelling in muscle cells, a key element for the net gain in surface GLUT4.

Crosstalk Regulates the Capacity for Robust Collective Decision Making in Heterogeneous Microbial Communities

NASA Astrophysics Data System (ADS)

Yusufaly, Tahir; Boedicker, James

Microbial communities frequently communicate via quorum sensing (QS), where cells produce, secrete, and respond to a threshold level of an autoinducer (AI) molecule, thereby modulating density-dependent gene expression. However, the biology of QS remains incompletely understood in heterogeneous communities, where crosstalk between distinct QS systems leads to novel effects. Such knowledge is necessary both for understanding signaling in real microbial communities, and for the rational design of synthetic communities with designer properties. As a step towards this goal, we investigate the effects of crosstalk between Gram-negative bacteria communicating via LuxI/LuxR-type QS systems, with acyl-homoserine lactone (AHL) AI molecules. After mapping QS in a heterogeneous community onto an artificial neural network model, we systematically analyze how heterogeneity regulates the community's capability for stable yet flexible decision making. We find that there are preferred distributions of interactions which provide optimal tradeoffs between capacity, or the number of different decisions a population can make, and robustness, or the tolerance of the community to disturbances. We compare our results to inferences made from experimental data, and critically discuss implications for the biological significance of crosstalk.

Multiple Phosphatidylinositol 3-Kinases Regulate Vaccinia Virus Morphogenesis

PubMed Central

McNulty, Shannon; Bornmann, William; Schriewer, Jill; Werner, Chas; Smith, Scott K.; Olson, Victoria A.; Damon, Inger K.; Buller, R. Mark; Heuser, John; Kalman, Daniel

2010-01-01

Poxvirus morphogenesis is a complex process that involves the successive wrapping of the virus in host cell membranes. We screened by plaque assay a focused library of kinase inhibitors for those that caused a reduction in viral growth and identified several compounds that selectively inhibit phosphatidylinositol 3-kinase (PI3K). Previous studies demonstrated that PI3Ks mediate poxviral entry. Using growth curves and electron microscopy in conjunction with inhibitors, we show that that PI3Ks additionally regulate morphogenesis at two distinct steps: immature to mature virion (IMV) transition, and IMV envelopment to form intracellular enveloped virions (IEV). Cells derived from animals lacking the p85 regulatory subunit of Type I PI3Ks (p85α−/−β−/−) presented phenotypes similar to those observed with PI3K inhibitors. In addition, VV appear to redundantly use PI3Ks, as PI3K inhibitors further reduce plaque size and number in p85α−/−β−/− cells. Together, these data provide evidence for a novel regulatory mechanism for virion morphogenesis involving phosphatidylinositol dynamics and may represent a new therapeutic target to contain poxviruses. PMID:20526370

The Recycling Endosome of Madin-Darby Canine Kidney Cells Is a Mildly Acidic Compartment Rich in Raft Components

PubMed Central

Gagescu, Raluca; Demaurex, Nicolas; Parton, Robert G.; Hunziker, Walter; Huber, Lukas A.; Gruenberg, Jean

2000-01-01

We present a biochemical and morphological characterization of recycling endosomes containing the transferrin receptor in the epithelial Madin-Darby canine kidney cell line. We find that recycling endosomes are enriched in molecules known to regulate transferrin recycling but lack proteins involved in early endosome membrane dynamics, indicating that recycling endosomes are distinct from conventional early endosomes. We also find that recycling endosomes are less acidic than early endosomes because they lack a functional vacuolar ATPase. Furthermore, we show that recycling endosomes can be reached by apically internalized tracers, confirming that the apical endocytic pathway intersects the transferrin pathway. Strikingly, recycling endosomes are enriched in the raft lipids sphingomyelin and cholesterol as well as in the raft-associated proteins caveolin-1 and flotillin-1. These observations may suggest that a lipid-based sorting mechanism operates along the Madin-Darby canine kidney recycling pathway, contributing to the maintenance of cell polarity. Altogether, our data indicate that recycling endosomes and early endosomes differ functionally and biochemically and thus that different molecular mechanisms regulate protein sorting and membrane traffic at each step of the receptor recycling pathway. PMID:10930469

Defining Hsp70 Subnetworks in Dengue Virus Replication Reveals Key Vulnerability in Flavivirus Infection

PubMed Central

Taguwa, Shuhei; Maringer, Kevin; Li, Xiaokai; Bernal-Rubio, Dabeiba; Rauch, Jennifer N.; Gestwicki, Jason E.; Andino, Raul; Fernandez-Sesma, Ana; Frydman, Judith

2015-01-01

Summary Viral protein homeostasis depends entirely on the machinery of the infected cell. Accordingly, viruses can illuminate the interplay between cellular proteostasis components and their distinct substrates. Here we define how the Hsp70 chaperone network mediates the dengue virus life cycle. Cytosolic Hsp70 isoforms are required at distinct steps of the viral cycle, including entry, RNA replication and virion biogenesis. Hsp70 function at each step is specified by nine distinct DNAJ cofactors. Of these, DnaJB11 relocalizes to virus-induced replication complexes to promote RNA synthesis, while DnaJB6 associates with capsid protein and facilitates virion biogenesis. Importantly, an allosteric Hsp70 inhibitor, JG40, potently blocks infection of different dengue serotypes in human primary blood cells without eliciting viral resistance or exerting toxicity to the host cells. JG40 also blocks replication of other medically-important flaviviruses including yellow fever, West Nile and Japanese encephalitis viruses. Thus, targeting host Hsp70 subnetworks provides a path for broad-spectrum antivirals. PMID:26582131

Neuronal somata and extrasomal compartments play distinct roles during synapse formation between Lymnaea neurons.

PubMed

Xu, Fenglian; Luk, Collin C; Wiersma-Meems, Ryanne; Baehre, Kelly; Herman, Cameron; Zaidi, Wali; Wong, Noelle; Syed, Naweed I

2014-08-20

Proper synapse formation is pivotal for all nervous system functions. However, the precise mechanisms remain elusive. Moreover, compared with the neuromuscular junction, steps regulating the synaptogenic program at central cholinergic synapses remain poorly defined. In this study, we identified different roles of neuronal compartments (somal vs extrasomal) in chemical and electrical synaptogenesis. Specifically, the electrically synapsed Lymnaea pedal dorsal A cluster neurons were used to study electrical synapses, whereas chemical synaptic partners, visceral dorsal 4 (presynaptic, cholinergic), and left pedal dorsal 1 (LPeD1; postsynaptic) were explored for chemical synapse formation. Neurons were cultured in a soma-soma or soma-axon configuration and synapses explored electrophysiologically. We provide the first direct evidence that electrical synapses develop in a soma-soma, but not soma-axon (removal of soma) configuration, indicating the requirement of gene transcription regulation in the somata of both synaptic partners. In addition, the soma-soma electrical coupling was contingent upon trophic factors present in Lymnaea brain-conditioned medium. Further, we demonstrate that chemical (cholinergic) synapses between soma-soma and soma-axon pairs were indistinguishable, with both exhibiting a high degree of contact site and target cell type specificity. We also provide direct evidence that presynaptic cell contact-mediated, clustering of postsynaptic cholinergic receptors at the synaptic site requires transmitter-receptor interaction, receptor internalization, and a protein kinase C-dependent lateral migration toward the contact site. This study provides novel insights into synaptogenesis between central neurons revealing both distinct and synergistic roles of cell-cell signaling and extrinsic trophic factors in executing the synaptogenic program. Copyright © 2014 the authors 0270-6474/14/3411304-12$15.00/0.

Nucleosome occupancy as a novel chromatin parameter for replication origin functions

PubMed Central

Rodriguez, Jairo; Lee, Laura; Lynch, Bryony; Tsukiyama, Toshio

2017-01-01

Eukaryotic DNA replication initiates from multiple discrete sites in the genome, termed origins of replication (origins). Prior to S phase, multiple origins are poised to initiate replication by recruitment of the pre-replicative complex (pre-RC). For proper replication to occur, origin activation must be tightly regulated. At the population level, each origin has a distinct firing time and frequency of activation within S phase. Many studies have shown that chromatin can strongly influence initiation of DNA replication. However, the chromatin parameters that affect properties of origins have not been thoroughly established. We found that nucleosome occupancy in G1 varies greatly around origins across the S. cerevisiae genome, and nucleosome occupancy around origins significantly correlates with the activation time and efficiency of origins, as well as pre-RC formation. We further demonstrate that nucleosome occupancy around origins in G1 is established during transition from G2/M to G1 in a pre-RC-dependent manner. Importantly, the diminished cell-cycle changes in nucleosome occupancy around origins in the orc1-161 mutant are associated with an abnormal global origin usage profile, suggesting that proper establishment of nucleosome occupancy around origins is a critical step for regulation of global origin activities. Our work thus establishes nucleosome occupancy as a novel and key chromatin parameter for proper origin regulation. PMID:27895110

44 CFR 7.926 - Age distinctions contained in FEMA regulations.

Code of Federal Regulations, 2014 CFR

2014-10-01

... 44 Emergency Management and Assistance 1 2014-10-01 2014-10-01 false Age distinctions contained in... REG. 5) Nondiscrimination on the Basis of Age in Programs or Activities Receiving Federal Financial Assistance From FEMA Standards for Determining Age Discrimination § 7.926 Age distinctions contained in FEMA...

44 CFR 7.926 - Age distinctions contained in FEMA regulations.

Code of Federal Regulations, 2012 CFR

2012-10-01

... 44 Emergency Management and Assistance 1 2012-10-01 2011-10-01 true Age distinctions contained in... REG. 5) Nondiscrimination on the Basis of Age in Programs or Activities Receiving Federal Financial Assistance From FEMA Standards for Determining Age Discrimination § 7.926 Age distinctions contained in FEMA...

44 CFR 7.926 - Age distinctions contained in FEMA regulations.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 44 Emergency Management and Assistance 1 2011-10-01 2011-10-01 false Age distinctions contained in... REG. 5) Nondiscrimination on the Basis of Age in Programs or Activities Receiving Federal Financial Assistance From FEMA Standards for Determining Age Discrimination § 7.926 Age distinctions contained in FEMA...

15 CFR 732.4 - Steps regarding License Exceptions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Steps regarding License Exceptions... (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS STEPS FOR USING THE EAR § 732.4 Steps regarding License Exceptions. (a) Introduction to Steps for License...

Executive function is necessary for the regulation of the stepping activity when stepping in place in older adults.

PubMed

Dalton, Christopher; Sciadas, Ria; Nantel, Julie

2016-10-01

To determine the effect of age on stepping performance and to compare the cognitive demand required to regulate repetitive stepping between older and younger adults while performing a stepping in place task (SIP). Fourteen younger (25.4 ± 6.5) and 15 older adults (71.0 ± 9.0) participated in this study. They performed a seated category fluency task and Stroop test, followed by a 60 s SIP task. Following this, both the cognitive and motor tasks were performed simultaneously. We assessed cognitive performance, SIP cycle duration, asymmetry, and arrhythmicity. Compared to younger adults, older adults had larger SIP arrhythmicity both as a single task and when combined with the Category (p < 0.001) and Stroop (p < 0.01) tasks. Older adults also had larger arrhythmicity when dual tasking compared to SIP alone (p < 0.001). Older adults showed greater SIP asymmetry when combined with Category (p = 0.006) and Stroop (p = 0.06) tasks. Finally, they had lower cognitive performance than younger adults in both single and dual tasks (p < 0.01). Age and type of cognitive task performed with the motor task affected different components of stepping. While SIP arrhythmicity was larger for all conditions in older compared to younger adults, cycle duration was not different, and asymmetry tended to be larger during SIP when paired with a verbal fluency task. SIP does not require a high level of control for dynamic stability, therefore demonstrating that higher-level executive function is necessary for the regulation of stepping activity independently of the regulation of postural balance. Furthermore, older adults may lack the cognitive resources needed to adequately regulate stepping activity while performing a cognitive task relying on the executive function.

Preparation of next-generation sequencing libraries using Nextera™ technology: simultaneous DNA fragmentation and adaptor tagging by in vitro transposition.

PubMed

Caruccio, Nicholas

2011-01-01

DNA library preparation is a common entry point and bottleneck for next-generation sequencing. Current methods generally consist of distinct steps that often involve significant sample loss and hands-on time: DNA fragmentation, end-polishing, and adaptor-ligation. In vitro transposition with Nextera™ Transposomes simultaneously fragments and covalently tags the target DNA, thereby combining these three distinct steps into a single reaction. Platform-specific sequencing adaptors can be added, and the sample can be enriched and bar-coded using limited-cycle PCR to prepare di-tagged DNA fragment libraries. Nextera technology offers a streamlined, efficient, and high-throughput method for generating bar-coded libraries compatible with multiple next-generation sequencing platforms.

5 CFR 531.508 - Evaluation of quality step increase authority.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 5 Administrative Personnel 1 2010-01-01 2010-01-01 false Evaluation of quality step increase... REGULATIONS PAY UNDER THE GENERAL SCHEDULE Quality Step Increases § 531.508 Evaluation of quality step... grant quality step increases. The agency shall take any corrective action required by the Office. [60 FR...

48 CFR 14.503-1 - Step one.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Step one. 14.503-1 Section... AND CONTRACT TYPES SEALED BIDDING Two-Step Sealed Bidding 14.503-1 Step one. (a) Requests for... use the two step method. (3) The requirements of the technical proposal. (4) The evaluation criteria...

Comprehensive Expression Map of Transcription Regulators in the Adult Zebrafish Telencephalon Reveals Distinct Neurogenic Niches

PubMed Central

Diotel, Nicolas; Rodriguez Viales, Rebecca; Armant, Olivier; März, Martin; Ferg, Marco; Rastegar, Sepand; Strähle, Uwe

2015-01-01

The zebrafish has become a model to study adult vertebrate neurogenesis. In particular, the adult telencephalon has been an intensely studied structure in the zebrafish brain. Differential expression of transcriptional regulators (TRs) is a key feature of development and tissue homeostasis. Here we report an expression map of 1,202 TR genes in the telencephalon of adult zebrafish. Our results are summarized in a database with search and clustering functions to identify genes expressed in particular regions of the telencephalon. We classified 562 genes into 13 distinct patterns, including genes expressed in the proliferative zone. The remaining 640 genes displayed unique and complex patterns of expression and could thus not be grouped into distinct classes. The neurogenic ventricular regions express overlapping but distinct sets of TR genes, suggesting regional differences in the neurogenic niches in the telencephalon. In summary, the small telencephalon of the zebrafish shows a remarkable complexity in TR gene expression. The adult zebrafish telencephalon has become a model to study neurogenesis. We established the expression pattern of more than 1200 transcription regulators (TR) in the adult telencephalon. The neurogenic regions express overlapping but distinct sets of TR genes suggesting regional differences in the neurogenic potential. J. Comp. Neurol. 523:1202–1221, 2015. © 2015 Wiley Periodicals, Inc. PMID:25556858

Comprehensive expression map of transcription regulators in the adult zebrafish telencephalon reveals distinct neurogenic niches.

PubMed

Diotel, Nicolas; Rodriguez Viales, Rebecca; Armant, Olivier; März, Martin; Ferg, Marco; Rastegar, Sepand; Strähle, Uwe

2015-06-01

The zebrafish has become a model to study adult vertebrate neurogenesis. In particular, the adult telencephalon has been an intensely studied structure in the zebrafish brain. Differential expression of transcriptional regulators (TRs) is a key feature of development and tissue homeostasis. Here we report an expression map of 1,202 TR genes in the telencephalon of adult zebrafish. Our results are summarized in a database with search and clustering functions to identify genes expressed in particular regions of the telencephalon. We classified 562 genes into 13 distinct patterns, including genes expressed in the proliferative zone. The remaining 640 genes displayed unique and complex patterns of expression and could thus not be grouped into distinct classes. The neurogenic ventricular regions express overlapping but distinct sets of TR genes, suggesting regional differences in the neurogenic niches in the telencephalon. In summary, the small telencephalon of the zebrafish shows a remarkable complexity in TR gene expression. The adult zebrafish telencephalon has become a model to study neurogenesis. We established the expression pattern of more than 1200 transcription regulators (TR) in the adult telencephalon. The neurogenic regions express overlapping but distinct sets of TR genes suggesting regional differences in the neurogenic potential. © 2015 Wiley Periodicals, Inc.

Proteomics of Dense Core Secretory Vesicles Reveal Distinct Protein Categories for Secretion of Neuroeffectors for Cell-Cell Communication

PubMed Central

Wegrzyn, Jill L.; Bark, Steven J.; Funkelstein, Lydiane; Mosier, Charles; Yap, Angel; Kazemi-Esfarjani, Parasa; La Spada, Albert; Sigurdson, Christina; O’Connor, Daniel T.; Hook, Vivian

2010-01-01

Regulated secretion of neurotransmitters and neurohumoural factors from dense core secretory vesicles provides essential neuroeffectors for cell-cell communication in the nervous and endocrine systems. This study provides comprehensive proteomic characterization of the categories of proteins in chromaffin dense core secretory vesicles that participate in cell-cell communication from the adrenal medulla. Proteomic studies were conducted by nano-HPLC Chip MS/MS tandem mass spectrometry. Results demonstrate that these secretory vesicles contain proteins of distinct functional categories consisting of neuropeptides and neurohumoural factors, protease systems, neurotransmitter enzymes and transporters, receptors, enzymes for biochemical processes, reduction/oxidation regulation, ATPases, protein folding, lipid biochemistry, signal transduction, exocytosis, calcium regulation, as well as structural and cell adhesion proteins. The secretory vesicle proteomic data identified 371 distinct proteins in the soluble fraction and 384 distinct membrane proteins, for a total of 686 distinct secretory vesicle proteins. Notably, these proteomic analyses illustrate the presence of several neurological disease-related proteins in these secretory vesicles, including huntingtin interacting protein, cystatin C, ataxin 7, and prion protein. Overall, these findings demonstrate that multiple protein categories participate in dense core secretory vesicles for production, storage, and secretion of bioactive neuroeffectors for cell-cell communication in health and disease. PMID:20695487

Hemolytic activity in Flavobacterium psychrophilum is a contact-dependent, two-step mechanism and differently expressed in smooth and rough phenotypes.

PubMed

Högfors-Rönnholm, Eva; Wiklund, Tom

2010-12-01

The hemolytic activity of cells of smooth and rough phenotypic variants of the Gram-negative fish pathogen Flavobacterium psychrophilum was investigated in two different assays, a microplate and an agarose hemolysis assay, using rainbow trout erythrocytes. The smooth cells showed a high and the rough cells a negligible, concentration dependent, hemolytic activity in the microplate assay. Both smooth and rough cells showed a rather weak hemolytic activity, with two distinct hemolytic patterns, in the agarose assay. The hemolytic activity of the cells was not regulated by iron availability and cell-free extracellular products did not show any hemolytic activity. The smooth cells, in contrast to the rough cells, showed a high ability to agglutinate erythrocytes and both hemagglutination and hemolytic activity was impaired by treatment of the cells with sialic acid. The hemolytic activity was furthermore reduced after proteolytic and heat treatment of the cells. The results from the present study suggest that the hemolytic activity in F. psychrophilum is highly expressed in the smooth phenotype, and that it is a contact-dependent and two-step mechanism that is initiated by the binding of the bacterial cells to the erythrocytes through sialic acid-binding lectins and then executed by thermolabile proteinaceous hemolysins. Copyright © 2010 Elsevier Ltd. All rights reserved.

Sequential steps in DNA replication are inhibited to ensure reduction of ploidy in meiosis

PubMed Central

Hua, Hui; Namdar, Mandana; Ganier, Olivier; Gregan, Juraj; Méchali, Marcel; Kearsey, Stephen E.

2013-01-01

Meiosis involves two successive rounds of chromosome segregation without an intervening S phase. Exit from meiosis I is distinct from mitotic exit, in that replication origins are not licensed by Mcm2-7 chromatin binding, but spindle disassembly occurs during a transient interphase-like state before meiosis II. The absence of licensing is assumed to explain the block to DNA replication, but this has not been formally tested. Here we attempt to subvert this block by expressing the licensing control factors Cdc18 and Cdt1 during the interval between meiotic nuclear divisions. Surprisingly, this leads only to a partial round of DNA replication, even when these factors are overexpressed and effect clear Mcm2-7 chromatin binding. Combining Cdc18 and Cdt1 expression with modulation of cyclin-dependent kinase activity, activation of Dbf4-dependent kinase, or deletion of the Spd1 inhibitor of ribonucleotide reductase has little additional effect on the extent of DNA replication. Single-molecule analysis indicates this partial round of replication results from inefficient progression of replication forks, and thus both initiation and elongation replication steps may be inhibited in late meiosis. In addition, DNA replication or damage during the meiosis I–II interval fails to arrest meiotic progress, suggesting absence of checkpoint regulation of meiosis II entry. PMID:23303250

Conformational heterogeneity and bubble dynamics in single bacterial transcription initiation complexes

PubMed Central

Duchi, Diego; Gryte, Kristofer; Robb, Nicole C; Morichaud, Zakia; Sheppard, Carol; Wigneshweraraj, Sivaramesh

2018-01-01

Abstract Transcription initiation is a major step in gene regulation for all organisms. In bacteria, the promoter DNA is first recognized by RNA polymerase (RNAP) to yield an initial closed complex. This complex subsequently undergoes conformational changes resulting in DNA strand separation to form a transcription bubble and an RNAP-promoter open complex; however, the series and sequence of conformational changes, and the factors that influence them are unclear. To address the conformational landscape and transitions in transcription initiation, we applied single-molecule Förster resonance energy transfer (smFRET) on immobilized Escherichia coli transcription open complexes. Our results revealed the existence of two stable states within RNAP–DNA complexes in which the promoter DNA appears to adopt closed and partially open conformations, and we observed large-scale transitions in which the transcription bubble fluctuated between open and closed states; these transitions, which occur roughly on the 0.1 s timescale, are distinct from the millisecond-timescale dynamics previously observed within diffusing open complexes. Mutational studies indicated that the σ70 region 3.2 of the RNAP significantly affected the bubble dynamics. Our results have implications for many steps of transcription initiation, and support a bend-load-open model for the sequence of transitions leading to bubble opening during open complex formation. PMID:29177430

The differential effects of intrinsic and identified motivation on well-being and performance: prospective, experimental, and implicit approaches to self-determination theory.

PubMed

Burton, Kimberly D; Lydon, John E; D'Alessandro, David U; Koestner, Richard

2006-10-01

Self-determination theory research has demonstrated that intrinsic and identified self-regulations are associated with successful adaptation. However, few distinctions are typically made between these regulations and their outcomes. In the present studies, the associations between intrinsic and identified motivations and outcomes of psychological well-being and academic performance are compared in educational settings. In Study 1, intrinsic self-regulation predicted psychological well-being, independent of academic performance. In contrast, identified regulation predicted academic performance. Additionally, the more that students demonstrated an identified academic regulation, the more that their psychological well-being was contingent on performance. In Study 2a, priming intrinsic self-regulation led to greater psychological well-being 10 days later. In Study 2b, an implicit measure of identified regulation predicted academic performance 6 weeks later. Results indicate the need to address important distinctions between intrinsic and identified regulations. 2006 APA, all rights reserved

New clinical grading scales and objective measurement for conjunctival injection.

PubMed

Park, In Ki; Chun, Yeoun Sook; Kim, Kwang Gi; Yang, Hee Kyung; Hwang, Jeong-Min

2013-08-05

To establish a new clinical grading scale and objective measurement method to evaluate conjunctival injection. Photographs of conjunctival injection with variable ocular diseases in 429 eyes were reviewed. Seventy-three images with concordance by three ophthalmologists were classified into a 4-step and 10-step subjective grading scale, and used as standard photographs. Each image was quantified in four ways: the relative magnitude of the redness component of each red-green-blue (RGB) pixel; two different algorithms based on the occupied area by blood vessels (K-means clustering with LAB color model and contrast-limited adaptive histogram equalization [CLAHE] algorithm); and the presence of blood vessel edges, based on the Canny edge-detection algorithm. Area under the receiver operating characteristic curves (AUCs) were calculated to summarize diagnostic accuracies of the four algorithms. The RGB color model, K-means clustering with LAB color model, and CLAHE algorithm showed good correlation with the clinical 10-step grading scale (R = 0.741, 0.784, 0.919, respectively) and with the clinical 4-step grading scale (R = 0.645, 0.702, 0.838, respectively). The CLAHE method showed the largest AUC, best distinction power (P < 0.001, ANOVA, Bonferroni multiple comparison test), and high reproducibility (R = 0.996). CLAHE algorithm showed the best correlation with the 10-step and 4-step subjective clinical grading scales together with high distinction power and reproducibility. CLAHE algorithm can be a useful for method for assessment of conjunctival injection.

7 CFR 65.230 - Production step.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 3 2010-01-01 2010-01-01 false Production step. 65.230 Section 65.230 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Standards..., PEANUTS, AND GINSENG General Provisions Definitions § 65.230 Production step. Production step means, in...

Traction force microscopy in rapidly moving cells reveals separate roles for ROCK and MLCK in the mechanics of retraction.

PubMed

Morin, Timothy R; Ghassem-Zadeh, Sean A; Lee, Juliet

2014-08-15

Retraction is a major rate-limiting step in cell motility, particularly in slow moving cell types that form large stable adhesions. Myosin II dependent contractile forces are thought to facilitate detachment by physically pulling up the rear edge. However, retraction can occur in the absence of myosin II activity in cell types that form small labile adhesions. To investigate the role of contractile force generation in retraction, we performed traction force microscopy during the movement of fish epithelial keratocytes. By correlating changes in local traction stress at the rear with the area retracted, we identified four distinct modes of retraction. "Recoil" retractions are preceded by a rise in local traction stress, while rear edge is temporarily stuck, followed by a sharp drop in traction stress upon detachment. This retraction type was most common in cells generating high average traction stress. In "pull" type retractions local traction stress and area retracted increase concomitantly. This was the predominant type of retraction in keratocytes and was observed mostly in cells generating low average traction stress. "Continuous" type retractions occur without any detectable change in traction stress, and are seen in cells generating low average traction stress. In contrast, to many other cell types, "release" type retractions occur in keratocytes following a decrease in local traction stress. Our identification of distinct modes of retraction suggests that contractile forces may play different roles in detachment that are related to rear adhesion strength. To determine how the regulation of contractility via MLCK or Rho kinase contributes to the mechanics of detachment, inhibitors were used to block or augment these pathways. Modulation of MLCK activity led to the most rapid change in local traction stress suggesting its importance in regulating attachment strength. Surprisingly, Rho kinase was not required for detachment, but was essential for localizing retraction to the rear. We suggest that in keratocytes MLCK and Rho kinase play distinct, complementary roles in the respective temporal and spatial control of rear detachment that is essential for maintaining rapid motility. Copyright © 2014 Elsevier Inc. All rights reserved.

Supporting Collaboration with Technology: Does Shared Cognition Lead to Co-Regulation in Medicine?

ERIC Educational Resources Information Center

Lajoie, Susanne P.; Lu, Jingyan

2012-01-01

The theoretical distinctions between metacognition, self-regulation and self-regulated learning are often blurred which makes the definition of co-regulation in group learning situations even more difficult. We have started to explore co-regulation in the context of decision making in simulated emergencies where medical teams work together to…

5 CFR 531.504 - Level of performance required for quality step increase.

Code of Federal Regulations, 2010 CFR

2010-01-01

... step increase. 531.504 Section 531.504 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS PAY UNDER THE GENERAL SCHEDULE Quality Step Increases § 531.504 Level of performance required for quality step increase. A quality step increase shall not be required but may be granted only...

5 CFR 531.505 - Restrictions on granting quality step increases.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 5 Administrative Personnel 1 2010-01-01 2010-01-01 false Restrictions on granting quality step... REGULATIONS PAY UNDER THE GENERAL SCHEDULE Quality Step Increases § 531.505 Restrictions on granting quality step increases. As provided by 5 U.S.C. 5336, a quality step increase may not be granted to an employee...

48 CFR 14.503-2 - Step two.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Step two. 14.503-2 Section... AND CONTRACT TYPES SEALED BIDDING Two-Step Sealed Bidding 14.503-2 Step two. (a) Sealed bidding... submitting acceptable technical proposals in step one; (2) Include the provision prescribed in 14.201-6(t...

A Common STEP in the Synaptic Pathology of Diverse Neuropsychiatric Disorders

PubMed Central

Johnson, Micah A.; Lombroso, Paul J.

2012-01-01

Synaptic function is critical for proper cognition, and synaptopathologies have been implicated in diverse neuropsychiatric disorders. STriatal-Enriched protein tyrosine Phosphatase (STEP) is a brain-enriched tyrosine phosphatase that normally opposes synaptic strengthening by dephosphorylating key neuronal signaling molecules. STEP targets include N-methyl D-aspartate receptors (NMDARs) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), as well as extracellular signal-regulated kinase (ERK) and the tyrosine kinase Fyn. STEP-mediated dephosphorylation promotes the internalization of NMDARs and AMPARs and the inactivation of ERK and Fyn. Regulation of STEP is complex, and recent work has implicated STEP dysregulation in the pathophysiology of several neuropsychiatric disorders. Both high levels and low levels of STEP are found in a diverse group of illnesses. This review focuses on the role of STEP in three disorders in which STEP levels are elevated: Alzheimer’s disease, fragile X syndrome, and schizophrenia. The presence of elevated STEP in all three of these disorders raises the intriguing possibility that cognitive deficits resulting from diverse etiologies may share a common molecular pathway. PMID:23239949

Synergistic Allosteric Mechanism of Fructose-1,6-bisphosphate and Serine for Pyruvate Kinase M2 via Dynamics Fluctuation Network Analysis.

PubMed

Yang, Jingxu; Liu, Hao; Liu, Xiaorui; Gu, Chengbo; Luo, Ray; Chen, Hai-Feng

2016-06-27

Pyruvate kinase M2 (PKM2) plays a key role in tumor metabolism and regulates the rate-limiting final step of glycolysis. In tumor cells, there are two allosteric effectors for PKM2: fructose-1,6-bisphosphate (FBP) and serine. However, the relationship between FBP and serine for allosteric regulation of PKM2 is unknown. Here we constructed residue/residue fluctuation correlation network based on all-atom molecular dynamics simulations to reveal the regulation mechanism. The results suggest that the correlation network in bound PKM2 is distinctly different from that in the free state, FBP/PKM2, or Ser/PKM2. The community network analysis indicates that the information can freely transfer from the allosteric sites of FBP and serine to the substrate site in bound PKM2, while there exists a bottleneck for information transfer in the network of the free state. Furthermore, the binding free energy between the substrate and PKM2 for bound PKM2 is significantly lower than either of FBP/PKM2 or Ser/PKM2. Thus, a hypothesis of "synergistic allosteric mechanism" is proposed for the allosteric regulation of FBP and serine. This hypothesis was further confirmed by the perturbational and mutational analyses of community networks and binding free energies. Finally, two possible synergistic allosteric pathways of FBP-K433-T459-R461-A109-V71-R73-MG2-OXL and Ser-I47-C49-R73-MG2-OXL were identified based on the shortest path algorithm and were confirmed by the network perturbation analysis. Interestingly, no similar pathways could be found in the free state. The process targeting on the allosteric pathways can better regulate the glycolysis of PKM2 and significantly inhibit the progression of tumor.

Beclin 1 Is Required for Neuron Viability and Regulates Endosome Pathways via the UVRAG-VPS34 Complex

PubMed Central

Wold, Mitchell S.; Gong, Shiaoching; Phillips, Greg R.; Dou, Zhixun; Zhao, Yanxiang; Heintz, Nathaniel; Zong, Wei-Xing; Yue, Zhenyu

2014-01-01

Deficiency of autophagy protein beclin 1 is implicated in tumorigenesis and neurodegenerative diseases, but the molecular mechanism remains elusive. Previous studies showed that Beclin 1 coordinates the assembly of multiple VPS34 complexes whose distinct phosphatidylinositol 3-kinase III (PI3K-III) lipid kinase activities regulate autophagy at different steps. Recent evidence suggests a function of beclin 1 in regulating multiple VPS34-mediated trafficking pathways beyond autophagy; however, the precise role of beclin 1 in autophagy-independent cellular functions remains poorly understood. Herein we report that beclin 1 regulates endocytosis, in addition to autophagy, and is required for neuron viability in vivo. We find that neuronal beclin 1 associates with endosomes and regulates EEA1/early endosome localization and late endosome formation. Beclin 1 maintains proper cellular phosphatidylinositol 3-phosphate (PI(3)P) distribution and total levels, and loss of beclin 1 causes a disruption of active Rab5 GTPase-associated endosome formation and impairment of endosome maturation, likely due to a failure of Rab5 to recruit VPS34. Furthermore, we find that Beclin 1 deficiency causes complete loss of the UVRAG-VPS34 complex and associated lipid kinase activity. Interestingly, beclin 1 deficiency impairs p40phox-linked endosome formation, which is rescued by overexpressed UVRAG or beclin 1, but not by a coiled-coil domain-truncated beclin 1 (a UVRAG-binding mutant), Atg14L or RUBICON. Thus, our study reveals the essential role for beclin 1 in neuron survival involving multiple membrane trafficking pathways including endocytosis and autophagy, and suggests that the UVRAG-beclin 1 interaction underlies beclin 1's function in endocytosis. PMID:25275521

Quantitative Analysis of Pac1/LIS1-mediated Dynein Targeting: Implications for Regulation of Dynein Activity in Budding Yeast

PubMed Central

Markus, Steven M.; Plevock, Karen M.; St. Germain, Bryan J.; Punch, Jesse J.; Meaden, Christopher W.; Lee, Wei-Lih

2011-01-01

LIS1 is a critical regulator of dynein function during mitosis and organelle transport. Here, we investigated how Pac1, the budding yeast LIS1 homologue, regulates dynein targeting and activity during nuclear migration. We show that Pac1 and Dyn1 (dynein heavy chain) are dependent upon each other and upon Bik1 (budding yeast CLIP-170 homologue) for plus end localization, whereas Bik1 is independent of either. Dyn1, Pac1 and Bik1 interact in vivo at the plus ends, where an excess amount of Bik1 recruits approximately equal amounts of Pac1 and Dyn1. Overexpression of Pac1 enhanced plus end targeting of Dyn1 and vice versa, while affinity-purification of Dyn1 revealed that it exists in a complex with Pac1 in the absence of Bik1, leading us to conclude that the Pac1-Dyn1 complex preassembles in the cytoplasm prior to loading onto Bik1-decorated plus ends. Strikingly, we found that Pac1-overexpression augments cortical dynein activity through a mechanism distinct from loss of She1, a negative regulator of dynein-dynactin association. While Pac1-overexpression enhances the frequency of cortical targeting for dynein and dynactin, the stoichiometry of these complexes remains relatively unchanged at the plus ends compared to that in wild-type cells (~3 dynein to 1 dynactin). Loss of She1, however, enhances dynein-dynactin association at the plus ends and the cell cortex, resulting in an apparent 1:1 stoichiometry. Our results reveal differential regulation of cortical dynein activity by She1 and Pac1, and provide a potentially new regulatory step in the off-loading model for dynein function. PMID:21294277

Inattention and Impulsivity: Differential Impact on School Readiness Capacities

ERIC Educational Resources Information Center

Sasser, Tyler; Bierman, Karen

2011-01-01

Despite the conceptual link between self-regulation skills and school readiness capacities, questions remain regarding how distinct but related facets of self-regulation (i.e., attention regulation, behavior regulation) differentially impact the development of school readiness capacities during early childhood. Additionally, little is known about…

Regulation of Epithelial Sodium Transport via Epithelial Na+ Channel

PubMed Central

Marunaka, Yoshinori; Niisato, Naomi; Taruno, Akiyuki; Ohta, Mariko; Miyazaki, Hiroaki; Hosogi, Shigekuni; Nakajima, Ken-ichi; Kusuzaki, Katsuyuki; Ashihara, Eishi; Nishio, Kyosuke; Iwasaki, Yoshinobu; Nakahari, Takashi; Kubota, Takahiro

2011-01-01

Renal epithelial Na+ transport plays an important role in homeostasis of our body fluid content and blood pressure. Further, the Na+ transport in alveolar epithelial cells essentially controls the amount of alveolar fluid that should be kept at an appropriate level for normal gas exchange. The epithelial Na+ transport is generally mediated through two steps: (1) the entry step of Na+ via epithelial Na+ channel (ENaC) at the apical membrane and (2) the extrusion step of Na+ via the Na+, K+-ATPase at the basolateral membrane. In general, the Na+ entry via ENaC is the rate-limiting step. Therefore, the regulation of ENaC plays an essential role in control of blood pressure and normal gas exchange. In this paper, we discuss two major factors in ENaC regulation: (1) activity of individual ENaC and (2) number of ENaC located at the apical membrane. PMID:22028593

Distinct functional outputs of PTEN signalling are controlled by dynamic association with β-arrestins

PubMed Central

Lima-Fernandes, Evelyne; Enslen, Hervé; Camand, Emeline; Kotelevets, Larissa; Boularan, Cédric; Achour, Lamia; Benmerah, Alexandre; Gibson, Lucien C D; Baillie, George S; Pitcher, Julie A; Chastre, Eric; Etienne-Manneville, Sandrine; Marullo, Stefano; Scott, Mark G H

2011-01-01

The tumour suppressor PTEN (phosphatase and tensin deleted on chromosome 10) regulates major cellular functions via lipid phosphatase-dependent and -independent mechanisms. Despite its fundamental pathophysiological importance, how PTEN's cellular activity is regulated has only been partially elucidated. We report that the scaffolding proteins β-arrestins (β-arrs) are important regulators of PTEN. Downstream of receptor-activated RhoA/ROCK signalling, β-arrs activate the lipid phosphatase activity of PTEN to negatively regulate Akt and cell proliferation. In contrast, following wound-induced RhoA activation, β-arrs inhibit the lipid phosphatase-independent anti-migratory effects of PTEN. β-arrs can thus differentially control distinct functional outputs of PTEN important for cell proliferation and migration. PMID:21642958

Regulated traffic of anion transporters in mammalian Brunner's glands: a role for water and fluid transport.

PubMed

Collaco, Anne M; Jakab, Robert L; Hoekstra, Nadia E; Mitchell, Kisha A; Brooks, Amos; Ameen, Nadia A

2013-08-01

The Brunner's glands of the proximal duodenum exert barrier functions through secretion of glycoproteins and antimicrobial peptides. However, ion transporter localization, function, and regulation in the glands are less clear. Mapping the subcellular distribution of transporters is an important step toward elucidating trafficking mechanisms of fluid transport in the gland. The present study examined 1) changes in the distribution of intestinal anion transporters and the aquaporin 5 (AQP5) water channel in rat Brunner's glands following second messenger activation and 2) anion transporter distribution in Brunner's glands from healthy and disease-affected human tissues. Cystic fibrosis transmembrane conductance regulator (CFTR), AQP5, sodium-potassium-coupled chloride cotransporter 1 (NKCC1), sodium-bicarbonate cotransporter (NBCe1), and the proton pump vacuolar ATPase (V-ATPase) were localized to distinct membrane domains and in endosomes at steady state. Carbachol and cAMP redistributed CFTR to the apical membrane. cAMP-dependent recruitment of CFTR to the apical membrane was accompanied by recruitment of AQP5 that was reversed by a PKA inhibitor. cAMP also induced apical trafficking of V-ATPase and redistribution of NKCC1 and NBCe1 to the basolateral membranes. The steady-state distribution of AQP5, CFTR, NBCe1, NKCC1, and V-ATPase in human Brunner's glands from healthy controls, cystic fibrosis, and celiac disease resembled that of rat; however, the distribution profiles were markedly attenuated in the disease-affected duodenum. These data support functional transport of chloride, bicarbonate, water, and protons by second messenger-regulated traffic in mammalian Brunner's glands under physiological and pathophysiological conditions.

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria†

PubMed Central

Deutscher, Josef; Francke, Christof; Postma, Pieter W.

2006-01-01

The phosphoenolpyruvate(PEP):carbohydrate phosphotransferase system (PTS) is found only in bacteria, where it catalyzes the transport and phosphorylation of numerous monosaccharides, disaccharides, amino sugars, polyols, and other sugar derivatives. To carry out its catalytic function in sugar transport and phosphorylation, the PTS uses PEP as an energy source and phosphoryl donor. The phosphoryl group of PEP is usually transferred via four distinct proteins (domains) to the transported sugar bound to the respective membrane component(s) (EIIC and EIID) of the PTS. The organization of the PTS as a four-step phosphoryl transfer system, in which all P derivatives exhibit similar energy (phosphorylation occurs at histidyl or cysteyl residues), is surprising, as a single protein (or domain) coupling energy transfer and sugar phosphorylation would be sufficient for PTS function. A possible explanation for the complexity of the PTS was provided by the discovery that the PTS also carries out numerous regulatory functions. Depending on their phosphorylation state, the four proteins (domains) forming the PTS phosphorylation cascade (EI, HPr, EIIA, and EIIB) can phosphorylate or interact with numerous non-PTS proteins and thereby regulate their activity. In addition, in certain bacteria, one of the PTS components (HPr) is phosphorylated by ATP at a seryl residue, which increases the complexity of PTS-mediated regulation. In this review, we try to summarize the known protein phosphorylation-related regulatory functions of the PTS. As we shall see, the PTS regulation network not only controls carbohydrate uptake and metabolism but also interferes with the utilization of nitrogen and phosphorus and the virulence of certain pathogens. PMID:17158705

Perceptual-motor regulation in locomotor pointing while approaching a curb.

PubMed

Andel, Steven van; Cole, Michael H; Pepping, Gert-Jan

2018-02-01

Locomotor pointing is a task that has been the focus of research in the context of sport (e.g. long jumping and cricket) as well as normal walking. Collectively, these studies have produced a broad understanding of locomotor pointing, but generalizability has been limited to laboratory type tasks and/or tasks with high spatial demands. The current study aimed to generalize previous findings in locomotor pointing to the common daily task of approaching and stepping on to a curb. Sixteen people completed 33 repetitions of a task that required them to walk up to and step onto a curb. Information about their foot placement was collected using a combination of measures derived from a pressure-sensitive walkway and video data. Variables related to perceptual-motor regulation were analyzed on an inter-trial, intra-step and inter-step level. Similar to previous studies, analysis of the foot placements showed that, variability in foot placement decreased as the participants drew closer to the curb. Regulation seemed to be initiated earlier in this study compared to previous studies, as shown by a decreasing variability in foot placement as early as eight steps before reaching the curb. Furthermore, it was shown that when walking up to the curb, most people regulated their walk in a way so as to achieve minimal variability in the foot placement on top of the curb, rather than a placement in front of the curb. Combined, these results showed a strong perceptual-motor coupling in the task of approaching and stepping up a curb, rendering this task a suitable test for perceptual-motor regulation in walking. Copyright © 2017 Elsevier B.V. All rights reserved.

5 CFR 531.506 - Effective date of a quality step increase.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 5 Administrative Personnel 1 2010-01-01 2010-01-01 false Effective date of a quality step increase... REGULATIONS PAY UNDER THE GENERAL SCHEDULE Quality Step Increases § 531.506 Effective date of a quality step increase. The quality step increase should be made effective as soon as practicable after it is approved...

15 CFR 732.3 - Steps regarding the ten general prohibitions.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Steps regarding the ten general... (Continued) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS STEPS FOR USING THE EAR § 732.3 Steps regarding the ten general prohibitions. (a) Introduction. If your item...

15 CFR 732.1 - Steps overview.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Steps overview. 732.1 Section 732.1... INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS STEPS FOR USING THE EAR § 732.1 Steps overview. (a)(1) Introduction. In this part, references to the EAR are references to 15...

48 CFR 15.202 - Advisory multi-step process.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Advisory multi-step... Information 15.202 Advisory multi-step process. (a) The agency may publish a presolicitation notice (see 5.204... participate in the acquisition. This process should not be used for multi-step acquisitions where it would...

Distinct phosphodiesterase 5A-containing compartments allow selective regulation of cGMP-dependent signalling in human arterial smooth muscle cells.

PubMed

Wilson, Lindsay S; Guo, Manhong; Umana, M Bibiana; Maurice, Donald H

2017-08-01

Cyclic GMP (cGMP) translates and integrates much of the information encoded by nitric oxide (NO · ) and several natriuretic peptides, including the atrial natriuretic peptide (ANP). Previously, we reported that integration of a cGMP-specific cyclic nucleotide phosphodiesterase, namely phosphodiesterase 5A (PDE5A), into a protein kinase G (PKG)- and inositol-1,4,5-trisphosphate receptor (IP 3 R)-containing endoplasmic reticulum (ER) signalosome allows localized control of PDE5A activity and of PKG-dependent inhibition of IP 3 -mediated release of ER Ca 2+ in human platelets. Herein, we report that PDE5A integrates into an analogous signalosome in human arterial smooth muscle cells (HASMC), wherein it regulates muscarinic agonist-dependent Ca 2+ release and is activated selectively by PKG-dependent phosphorylation. In addition, we report that PDE5A also regulates HASMC functions via events independent of PKG, but rather through actions coordinated by competitive cGMP-mediated inhibition of cAMP hydrolysis by the so-called cGMP-inhibited cAMP PDE, namely phosphodiesterase 3A (PDE3A). Indeed, we show that ANP increases both cGMP and cAMP levels in HASMC and promotes phosphorylation of vasodilator-stimulated phospho-protein (VASP) at each the PKG and PKA phospho-acceptor sites. Since selective inhibition of PDE5 decreased DNA synthesis and chemotaxis of HASMC, and that PDE3A knockdown obviated these effects, our findings are consistent with a role for a PDE5A-PDE3A-PKA axis in their regulation. Our findings provide insight into the existence of distinct "pools" of PDE5A in HASMC and support the idea that these discrete compartments regulate distinct cGMP-dependent events. As a corollary, we suggest that it may be possible to target these distinct PDE5A-regulated pools and in so-doing differentially impact selected cGMP-regulated functions in these cells. Copyright © 2017. Published by Elsevier Inc.

Regulative Loops, Step Loops and Task Loops

ERIC Educational Resources Information Center

VanLehn, Kurt

2016-01-01

This commentary suggests a generalization of the conception of the behavior of tutoring systems, which the target article characterized as having an outer loop that was executed once per task and an inner loop that was executed once per step of the task. A more general conception sees these two loops as instances of regulative loops, which…

Massive Analysis of Rice Small RNAs: Mechanistic Implications of Regulated MicroRNAs and Variants for Differential Target RNA Cleavage[W][OA

PubMed Central

Jeong, Dong-Hoon; Park, Sunhee; Zhai, Jixian; Gurazada, Sai Guna Ranjan; De Paoli, Emanuele; Meyers, Blake C.; Green, Pamela J.

2011-01-01

Small RNAs have a variety of important roles in plant development, stress responses, and other processes. They exert their influence by guiding mRNA cleavage, translational repression, and chromatin modification. To identify previously unknown rice (Oryza sativa) microRNAs (miRNAs) and those regulated by environmental stress, 62 small RNA libraries were constructed from rice plants and used for deep sequencing with Illumina technology. The libraries represent several tissues from control plants and plants subjected to different environmental stress treatments. More than 94 million genome-matched reads were obtained, resulting in more than 16 million distinct small RNA sequences. This allowed an evaluation of ~400 annotated miRNAs with current criteria and the finding that among these, ~150 had small interfering RNA–like characteristics. Seventy-six new miRNAs were found, and miRNAs regulated in response to water stress, nutrient stress, or temperature stress were identified. Among the new examples of miRNA regulation were members of the same miRNA family that were differentially regulated in different organs and had distinct sequences Some of these distinct family members result in differential target cleavage and provide new insight about how an agriculturally important rice phenotype could be regulated in the panicle. This high-resolution analysis of rice miRNAs should be relevant to plant miRNAs in general, particularly in the Poaceae. PMID:22158467

Differential regulation of microtubule severing by APC underlies distinct patterns of projection neuron and interneuron migration

PubMed Central

Eom, Tae-Yeon; Stanco, Amelia; Guo, Jiami; Wilkins, Gary; Deslauriers, Danielle; Yan, Jessica; Monckton, Chase; Blair, Josh; Oon, Eesim; Perez, Abby; Salas, Eduardo; Oh, Adrianna; Ghukasyan, Vladimir; Snider, William D.; Rubenstein, John L. R.; Anton, E. S.

2014-01-01

Coordinated migration of distinct classes of neurons to appropriate positions leads to the formation of functional neuronal circuitry in the cerebral cortex. Two major classes of cortical neurons, interneurons and projection neurons, utilize distinctly different modes (radial vs. tangential) and routes of migration to arrive at their final positions in the cerebral cortex. Here, we show that adenomatous polyposis coli (APC) modulates microtubule (MT) severing in interneurons to facilitate tangential mode of interneuron migration, but not the glial-guided, radial migration of projection neurons. APC regulates the stability and activity of the MT severing protein p60-katanin in interneurons to promote the rapid remodeling of neuronal processes necessary for interneuron migration. These findings reveal how severing and restructuring of MTs facilitate distinct modes of neuronal migration necessary for laminar organization of neurons in the developing cerebral cortex. PMID:25535916

Defining Hsp70 Subnetworks in Dengue Virus Replication Reveals Key Vulnerability in Flavivirus Infection.

PubMed

Taguwa, Shuhei; Maringer, Kevin; Li, Xiaokai; Bernal-Rubio, Dabeiba; Rauch, Jennifer N; Gestwicki, Jason E; Andino, Raul; Fernandez-Sesma, Ana; Frydman, Judith

2015-11-19

Viral protein homeostasis depends entirely on the machinery of the infected cell. Accordingly, viruses can illuminate the interplay between cellular proteostasis components and their distinct substrates. Here, we define how the Hsp70 chaperone network mediates the dengue virus life cycle. Cytosolic Hsp70 isoforms are required at distinct steps of the viral cycle, including entry, RNA replication, and virion biogenesis. Hsp70 function at each step is specified by nine distinct DNAJ cofactors. Of these, DnaJB11 relocalizes to virus-induced replication complexes to promote RNA synthesis, while DnaJB6 associates with capsid protein and facilitates virion biogenesis. Importantly, an allosteric Hsp70 inhibitor, JG40, potently blocks infection of different dengue serotypes in human primary blood cells without eliciting viral resistance or exerting toxicity to the host cells. JG40 also blocks replication of other medically-important flaviviruses including yellow fever, West Nile and Japanese encephalitis viruses. Thus, targeting host Hsp70 subnetworks provides a path for broad-spectrum antivirals. Copyright © 2015 Elsevier Inc. All rights reserved.

Impact analysis of tap switch out of step for converter transformer

NASA Astrophysics Data System (ADS)

Hong-yue, ZHANG; Zhen-hua, ZHANG; Zhang-xue, XIONG; Gao-wang, YU

2017-06-01

AC transformer load regulation is mainly used to adjust the load side voltage level, improve the quality of power supply, the voltage range is relatively narrow. In DC system, converter transformer is the core equipment of AC and DC power converter and inverter. converter transformer tap adjustment can maintain the normal operation of the converter in small angle range control, the absorption of reactive power, economic operation, valve less stress, valve damping circuit loss, AC / DC harmonic component is also smaller. In this way, the tap switch action is more frequent, and a large range of the tap switch adjustment is required. Converter transformer with a more load voltage regulation switch, the voltage regulation range of the switch is generally 20~30%, the adjustment of each file is 1%~2%. Recently it is often found that the tap switch of Converter Transformers is out of step in Converter station. In this paper, it is analyzed in detail the impact of tap switch out of step for differential protection, overexcitation protection and zero sequence over current protection. Analysis results show that: the tap switch out of step has no effect on the differential protection and the overexcitation protection including the tap switch. But the tap switch out of step has effect on zero sequence overcurrent protection of out of step star-angle converter transformer. The zero sequence overcurrent protection will trip when the tap switch out of step is greater than 3 for out of step star-angle converter transformer.

Unlocking the coral calcification process: Insights from boron isotope measurements and a skeletal growth model

NASA Astrophysics Data System (ADS)

Mollica, N. R.; Guo, W.; Cohen, A. L.; Huang, K. F.; Foster, G. L.; Donald, H.; Solow, A.

2017-12-01

Carbonate skeletons of scleractinian corals are important archives of ocean climate and environmental change. However, corals don't accrete their skeletons directly from ambient seawater, but from a calcifying fluid whose composition is strongly regulated. There is mounting evidence that the carbonate chemistry of this calcifying fluid significantly impacts the amount of carbonate the coral can precipitate, which in turn affects the geochemical composition of the skeleton produced. However the mechanistic link between calcifying fluid (cf) chemistry, particularly the up-regulation of pHcf and thereby aragonite saturation state (Ωcf), and coral calcification is not well understood. We explored this link by combining boron isotope measurements with in situ measurements of seawater temperature, salinity, and DIC to estimate Ωcf of nine Porites corals from four Pacific reefs. Associated calcification rates were quantified for each core via CT scanning. We do not observe a relationship between calcification rates and Ωcf or Ωsw. Instead, when we deconvolve calcification into linear extension and skeletal density, a significant correlation is observed between density and Ωcf, and also Ωsw while extension does not correlate with either. These observations are consistent with the two-step model of coral calcification, in which skeleton is secreted in two distinct phases: vertical extension creating new skeletal elements, followed by lateral thickening of existing elements that are covered by living tissue. We developed a numerical model of Porites skeletal growth that builds on this two-step model and links skeletal density with the external seawater environment via its influence on the chemistry of coral calcifying fluid. We validated the model using existing coral skeletal datasets from six Porites species collected across five reef sites, and quantified the effects of each seawater parameter (e.g. temperature, pH, DIC) on skeletal density. Our findings illustrate the sensitivity of the second phase of coral calcification to the carbonate chemistry of the calcifying fluid, and support previous coral proxy system modelling efforts by validating the two-step growth model on annual and seasonal scales.

Distinct domains within the NITROGEN LIMITATION ADAPTATION protein mediate its subcellular localization and function in the nitrate-dependent phosphate homeostasis pathway

USDA-ARS?s Scientific Manuscript database

The NITROGEN LIMITATION ADAPTATION (NLA) protein is a RING-type E3 ubiquitin ligase that plays an essential role in the regulation of nitrogen and phosphate homeostasis. NLA is localized to two distinct subcellular sites, the plasma membrane and nucleus, and contains four distinct domains: i) a RING...

Modeling-independent elucidation of inactivation pathways in recombinant and native A-type Kv channels.

PubMed

Fineberg, Jeffrey D; Ritter, David M; Covarrubias, Manuel

2012-11-01

A-type voltage-gated K(+) (Kv) channels self-regulate their activity by inactivating directly from the open state (open-state inactivation [OSI]) or by inactivating before they open (closed-state inactivation [CSI]). To determine the inactivation pathways, it is often necessary to apply several pulse protocols, pore blockers, single-channel recording, and kinetic modeling. However, intrinsic hurdles may preclude the standardized application of these methods. Here, we implemented a simple method inspired by earlier studies of Na(+) channels to analyze macroscopic inactivation and conclusively deduce the pathways of inactivation of recombinant and native A-type Kv channels. We investigated two distinct A-type Kv channels expressed heterologously (Kv3.4 and Kv4.2 with accessory subunits) and their native counterparts in dorsal root ganglion and cerebellar granule neurons. This approach applies two conventional pulse protocols to examine inactivation induced by (a) a simple step (single-pulse inactivation) and (b) a conditioning step (double-pulse inactivation). Consistent with OSI, the rate of Kv3.4 inactivation (i.e., the negative first derivative of double-pulse inactivation) precisely superimposes on the profile of the Kv3.4 current evoked by a single pulse because the channels must open to inactivate. In contrast, the rate of Kv4.2 inactivation is asynchronous, already changing at earlier times relative to the profile of the Kv4.2 current evoked by a single pulse. Thus, Kv4.2 inactivation occurs uncoupled from channel opening, indicating CSI. Furthermore, the inactivation time constant versus voltage relation of Kv3.4 decreases monotonically with depolarization and levels off, whereas that of Kv4.2 exhibits a J-shape profile. We also manipulated the inactivation phenotype by changing the subunit composition and show how CSI and CSI combined with OSI might affect spiking properties in a full computational model of the hippocampal CA1 neuron. This work unambiguously elucidates contrasting inactivation pathways in neuronal A-type Kv channels and demonstrates how distinct pathways might impact neurophysiological activity.

Redox and Reactive Oxygen Species Regulation of Mitochondrial Cytochrome c Oxidase Biogenesis

PubMed Central

Bourens, Myriam; Fontanesi, Flavia; Soto, Iliana C.; Liu, Jingjing

2013-01-01

Abstract Significance: Cytochrome c oxidase (COX), the last enzyme of the mitochondrial respiratory chain, is the major oxygen consumer enzyme in the cell. COX biogenesis involves several redox-regulated steps. The process is highly regulated to prevent the formation of pro-oxidant intermediates. Recent Advances: Regulation of COX assembly involves several reactive oxygen species and redox-regulated steps. These include: (i) Intricate redox-controlled machineries coordinate the expression of COX isoenzymes depending on the environmental oxygen concentration. (ii) COX is a heme A-copper metalloenzyme. COX copper metallation involves the copper chaperone Cox17 and several other recently described cysteine-rich proteins, which are oxidatively folded in the mitochondrial intermembrane space. Copper transfer to COX subunits 1 and 2 requires concomitant transfer of redox power. (iii) To avoid the accumulation of reactive assembly intermediates, COX is regulated at the translational level to minimize synthesis of the heme A-containing Cox1 subunit when assembly is impaired. Critical Issues: An increasing number of regulatory pathways converge to facilitate efficient COX assembly, thus preventing oxidative stress. Future Directions: Here we will review on the redox-regulated COX biogenesis steps and will discuss their physiological relevance. Forthcoming insights into the precise regulation of mitochondrial COX biogenesis in normal and stress conditions will likely open future perspectives for understanding mitochondrial redox regulation and prevention of oxidative stress. Antioxid. Redox Signal. 19, 1940–1952. PMID:22937827

Direction Finding With Mutually Orthogonal Antennas

DTIC Science & Technology

2011-03-24

information of these waves must be estimated. The third step, referred to as geolocation , attempts to use the bearing information from step two...DF is distinct from, but related to, the geolocation problem. In DF we seek to answer, “where did that signal come from?” The geolocation problem...Sensor Technology Division) contracted a major aeronautical systems development company to research a DF and geolocation system for UAVs. The system

Striatal-enriched Protein-tyrosine Phosphatase (STEP) Regulates Pyk2 Kinase Activity*

PubMed Central

Xu, Jian; Kurup, Pradeep; Bartos, Jason A.; Patriarchi, Tommaso; Hell, Johannes W.; Lombroso, Paul J.

2012-01-01

Proline-rich tyrosine kinase 2 (Pyk2) is a member of the focal adhesion kinase family and is highly expressed in brain and hematopoietic cells. Pyk2 plays diverse functions in cells, including the regulation of cell adhesion, migration, and cytoskeletal reorganization. In the brain, it is involved in the induction of long term potentiation through regulation of N-methyl-d-aspartate receptor trafficking. This occurs through the phosphorylation and activation of Src family tyrosine kinase members, such as Fyn, that phosphorylate GluN2B at Tyr1472. Phosphorylation at this site leads to exocytosis of GluN1-GluN2B receptors to synaptic membranes. Pyk2 activity is modulated by phosphorylation at several critical tyrosine sites, including Tyr402. In this study, we report that Pyk2 is a substrate of striatal-enriched protein-tyrosine phosphatase (STEP). STEP binds to and dephosphorylates Pyk2 at Tyr402. STEP KO mice showed enhanced phosphorylation of Pyk2 at Tyr402 and of the Pyk2 substrates paxillin and ASAP1. Functional studies indicated that STEP opposes Pyk2 activation after KCl depolarization of cortical slices and blocks Pyk2 translocation to postsynaptic densities, a key step required for Pyk2 activation and function. This is the first study to identify Pyk2 as a substrate for STEP. PMID:22544749

E2f1–3 Are Critical for Myeloid Development*

PubMed Central

Trikha, Prashant; Sharma, Nidhi; Opavsky, Rene; Reyes, Andres; Pena, Clarissa; Ostrowski, Michael C.; Roussel, Martine F.; Leone, Gustavo

2011-01-01

Hematopoietic development involves the coordinated activity of differentiation and cell cycle regulators. In current models of mammalian cell cycle control, E2f activators (E2f1, E2f2, and E2f3) are portrayed as the ultimate transcriptional effectors that commit cells to enter and progress through S phase. Using conditional gene knock-out strategies, we show that E2f1–3 are not required for the proliferation of early myeloid progenitors. Rather, these E2fs are critical for cell survival and proliferation at two distinct steps of myeloid development. First, E2f1–3 are required as transcriptional repressors for the survival of CD11b+ myeloid progenitors, and then they are required as activators for the proliferation of CD11b+ macrophages. In bone marrow macrophages, we show that E2f1–3 respond to CSF1-Myc mitogenic signals and serve to activate E2f target genes and promote their proliferation. Together, these findings expose dual functions for E2f1–3 at distinct stages of myeloid development in vivo, first as repressors in cell survival and then as activators in cell proliferation. In summary, this work places E2f1–3 in a specific signaling cascade that is critical for myeloid development in vivo. PMID:21115501

GABA Signaling Promotes Synapse Elimination and Axon Pruning in Developing Cortical Inhibitory Interneurons

PubMed Central

Wu, Xiaoyun; Fu, Yu; Knott, Graham; Lu, Jiangteng; Di Cristo, Graziella

2012-01-01

Accumulating evidence indicates that GABA acts beyond inhibitory synaptic transmission and regulates the development of inhibitory synapses in the vertebrate brain, but the underlying cellular mechanism is not well understood. We have combined live imaging of cortical GABAergic axons across time scales from minutes to days with single-cell genetic manipulation of GABA release to examine its role in distinct steps of inhibitory synapse formation in the mouse neocortex. We have shown previously, by genetic knockdown of GABA synthesis in developing interneurons, that GABA signaling promotes the maturation of inhibitory synapses and axons. Here we found that a complete blockade of GABA release in basket interneurons resulted in an opposite effect, a cell-autonomous increase in axon and bouton density with apparently normal synapse structures. These results not only demonstrate that GABA is unnecessary for synapse formation per se but also uncover a novel facet of GABA in regulating synapse elimination and axon pruning. Live imaging revealed that developing GABAergic axons form a large number of transient boutons, but only a subset was stabilized. Release blockade led to significantly increased bouton stability and filopodia density, increased axon branch extension, and decreased branch retraction. Our results suggest that a major component of GABA function in synapse development is transmission-mediated elimination of subsets of nascent contacts. Therefore, GABA may regulate activity-dependent inhibitory synapse formation by coordinately eliminating certain nascent contacts while promoting the maturation of other nascent synapses. PMID:22219294

Arabidopsis DRB4, AGO1, AGO7, and RDR6 participate in a DCL4-initiated antiviral RNA silencing pathway negatively regulated by DCL1.

PubMed

Qu, Feng; Ye, Xiaohong; Morris, T Jack

2008-09-23

Plant RNA silencing machinery enlists four primary classes of proteins to achieve sequence-specific regulation of gene expression and mount an antiviral defense. These include Dicer-like ribonucleases (DCLs), Argonaute proteins (AGOs), dsRNA-binding proteins (DRBs), and RNA-dependent RNA polymerases (RDRs). Although at least four distinct endogenous RNA silencing pathways have been thoroughly characterized, a detailed understanding of the antiviral RNA silencing pathway is just emerging. In this report, we have examined the role of four DCLs, two AGOs, one DRB, and one RDR in controlling viral RNA accumulation in infected Arabidopsis plants by using a mutant virus lacking its silencing suppressor. Our results show that all four DCLs contribute to antiviral RNA silencing. We confirm previous reports implicating both DCL4 and DCL2 in this process and establish a minor role for DCL3. Surprisingly, we found that DCL1 represses antiviral RNA silencing through negatively regulating the expression of DCL4 and DCL3. We also implicate DRB4 in antiviral RNA silencing. Finally, we show that both AGO1 and AGO7 function to ensure efficient clearance of viral RNAs and establish that AGO1 is capable of targeting viral RNAs with more compact structures, whereas AGO7 and RDR6 favor less structured RNA targets. Our results resolve several key steps in the antiviral RNA silencing pathway and provide a basis for further in-depth analysis.

Integration of anteroposterior and dorsoventral regulation of Phox2b transcription in cranial motoneuron progenitors by homeodomain proteins.

PubMed

Samad, Omar Abdel; Geisen, Marc J; Caronia, Giuliana; Varlet, Isabelle; Zappavigna, Vincenzo; Ericson, Johan; Goridis, Christo; Rijli, Filippo M

2004-08-01

Little is known about the molecular mechanisms that integrate anteroposterior (AP) and dorsoventral (DV) positional information in neural progenitors that specify distinct neuronal types within the vertebrate neural tube. We have previously shown that in ventral rhombomere (r)4 of Hoxb1 and Hoxb2 mutant mouse embryos, Phox2b expression is not properly maintained in the visceral motoneuron progenitor domain (pMNv), resulting in a switch to serotonergic fate. Here, we show that Phox2b is a direct target of Hoxb1 and Hoxb2. We found a highly conserved Phox2b proximal enhancer that mediates rhombomere-restricted expression and contains separate Pbx-Hox (PH) and Prep/Meis (P/M) binding sites. We further show that both the PH and P/M sites are essential for Hox-Pbx-Prep ternary complex formation and regulation of the Phox2b enhancer activity in ventral r4. Moreover, the DV factor Nkx2.2 enhances Hox-mediated transactivation via a derepression mechanism. Finally, we show that induction of ectopic Phox2b-expressing visceral motoneurons in the chick hindbrain requires the combined activities of Hox and Nkx2 homeodomain proteins. This study takes an important first step to understand how activators and repressors, induced along the AP and DV axes in response to signaling pathways, interact to regulate specific target gene promoters, leading to neuronal fate specification in the appropriate developmental context.

Identification of Isn1 and Sdt1 as Glucose- and Vitamin-regulated Nicotinamide Mononucleotide and Nicotinic Acid Mononucleotide 5′-Nucleotidases Responsible for Production of Nicotinamide Riboside and Nicotinic Acid Riboside*

PubMed Central

Bogan, Katrina L.; Evans, Charles; Belenky, Peter; Song, Peng; Burant, Charles F.; Kennedy, Robert; Brenner, Charles

2009-01-01

Recently, we discovered that nicotinamide riboside and nicotinic acid riboside are biosynthetic precursors of NAD+, which are utilized through two pathways consisting of distinct enzymes. In addition, we have shown that exogenously supplied nicotinamide riboside is imported into yeast cells by a dedicated transporter, and it extends replicative lifespan on high glucose medium. Here, we show that nicotinamide riboside and nicotinic acid riboside are authentic intracellular metabolites in yeast. Secreted nicotinamide riboside was detected with a biological assay, and intracellular levels of nicotinamide riboside, nicotinic acid riboside, and other NAD+ metabolites were determined by a liquid chromatography-mass spectrometry method. A biochemical genomic screen indicated that three yeast enzymes possess nicotinamide mononucleotide 5′-nucleotidase activity in vitro. Metabolic profiling of knock-out mutants established that Isn1 and Sdt1 are responsible for production of nicotinamide riboside and nicotinic acid riboside in cells. Isn1, initially classified as an IMP-specific 5′-nucleotidase, and Sdt1, initially classified as a pyrimidine 5′-nucleotidase, are additionally responsible for dephosphorylation of pyridine mononucleotides. Sdt1 overexpression is growth-inhibitory to cells in a manner that depends on its active site and correlates with reduced cellular NAD+. Expression of Isn1 protein is positively regulated by the availability of nicotinic acid and glucose. These results reveal unanticipated and highly regulated steps in NAD+ metabolism. PMID:19846558

Identification of Isn1 and Sdt1 as glucose- and vitamin-regulated nicotinamide mononucleotide and nicotinic acid mononucleotide [corrected] 5'-nucleotidases responsible for production of nicotinamide riboside and nicotinic acid riboside.

PubMed

Bogan, Katrina L; Evans, Charles; Belenky, Peter; Song, Peng; Burant, Charles F; Kennedy, Robert; Brenner, Charles

2009-12-11

Recently, we discovered that nicotinamide riboside and nicotinic acid riboside are biosynthetic precursors of NAD(+), which are utilized through two pathways consisting of distinct enzymes. In addition, we have shown that exogenously supplied nicotinamide riboside is imported into yeast cells by a dedicated transporter, and it extends replicative lifespan on high glucose medium. Here, we show that nicotinamide riboside and nicotinic acid riboside are authentic intracellular metabolites in yeast. Secreted nicotinamide riboside was detected with a biological assay, and intracellular levels of nicotinamide riboside, nicotinic acid riboside, and other NAD(+) metabolites were determined by a liquid chromatography-mass spectrometry method. A biochemical genomic screen indicated that three yeast enzymes possess nicotinamide mononucleotide 5'-nucleotidase activity in vitro. Metabolic profiling of knock-out mutants established that Isn1 and Sdt1 are responsible for production of nicotinamide riboside and nicotinic acid riboside in cells. Isn1, initially classified as an IMP-specific 5'-nucleotidase, and Sdt1, initially classified as a pyrimidine 5'-nucleotidase, are additionally responsible for dephosphorylation of pyridine mononucleotides. Sdt1 overexpression is growth-inhibitory to cells in a manner that depends on its active site and correlates with reduced cellular NAD(+). Expression of Isn1 protein is positively regulated by the availability of nicotinic acid and glucose. These results reveal unanticipated and highly regulated steps in NAD(+) metabolism.

Seed maturation associated transcriptional programs and regulatory networks underlying genotypic difference in seed dormancy and size/weight in wheat (Triticum aestivum L.).

PubMed

Yamasaki, Yuji; Gao, Feng; Jordan, Mark C; Ayele, Belay T

2017-09-16

Maturation forms one of the critical seed developmental phases and it is characterized mainly by programmed cell death, dormancy and desiccation, however, the transcriptional programs and regulatory networks underlying acquisition of dormancy and deposition of storage reserves during the maturation phase of seed development are poorly understood in wheat. The present study performed comparative spatiotemporal transcriptomic analysis of seed maturation in two wheat genotypes with contrasting seed weight/size and dormancy phenotype. The embryo and endosperm tissues of maturing seeds appeared to exhibit genotype-specific temporal shifts in gene expression profile that might contribute to the seed phenotypic variations. Functional annotations of gene clusters suggest that the two tissues exhibit distinct but genotypically overlapping molecular functions. Motif enrichment predicts genotypically distinct abscisic acid (ABA) and gibberellin (GA) regulated transcriptional networks contribute to the contrasting seed weight/size and dormancy phenotypes between the two genotypes. While other ABA responsive element (ABRE) motifs are enriched in both genotypes, the prevalence of G-box-like motif specifically in tissues of the dormant genotype suggests distinct ABA mediated transcriptional mechanisms control the establishment of dormancy during seed maturation. In agreement with this, the bZIP transcription factors that co-express with ABRE enriched embryonic genes differ with genotype. The enrichment of SITEIIATCYTC motif specifically in embryo clusters of maturing seeds irrespective of genotype predicts a tissue specific role for the respective TCP transcription factors with no or minimal contribution to the variations in seed dormancy. The results of this study advance our understanding of the seed maturation associated molecular mechanisms underlying variation in dormancy and weight/size in wheat seeds, which is a critical step towards the designing of molecular strategies for enhancing seed yield and quality.

15 CFR 732.6 - Steps for other requirements.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Steps for other requirements. 732.6...) BUREAU OF INDUSTRY AND SECURITY, DEPARTMENT OF COMMERCE EXPORT ADMINISTRATION REGULATIONS STEPS FOR USING THE EAR § 732.6 Steps for other requirements. Sections 732.1 through 732.4 of this part are useful in...

45 CFR 1172.1 - Purpose.

Code of Federal Regulations, 2014 CFR

2014-10-01

... Regulations Relating to Public Welfare (Continued) NATIONAL FOUNDATION ON THE ARTS AND THE HUMANITIES NATIONAL... distinctions and factors other than age which meet the requirements of the Act and the regulations in this part. The regulations in this part are based upon the general, government-wide age discrimination...

Self-Regulated Strategy Development Instruction for Teaching Multi-Step Equations to Middle School Students Struggling in Math

ERIC Educational Resources Information Center

Cuenca-Carlino, Yojanna; Freeman-Green, Shaqwana; Stephenson, Grant W.; Hauth, Clara

2016-01-01

Six middle school students identified as having a specific learning disability or at risk for mathematical difficulties were taught how to solve multi-step equations by using the self-regulated strategy development (SRSD) model of instruction. A multiple-probe-across-pairs design was used to evaluate instructional effects. Instruction was provided…

Regulatory effects of cotranscriptional RNA structure formation and transitions.

PubMed

Liu, Sheng-Rui; Hu, Chun-Gen; Zhang, Jin-Zhi

2016-09-01

RNAs, which play significant roles in many fundamental biological processes of life, fold into sophisticated and precise structures. RNA folding is a dynamic and intricate process, which conformation transition of coding and noncoding RNAs form the primary elements of genetic regulation. The cellular environment contains various intrinsic and extrinsic factors that potentially affect RNA folding in vivo, and experimental and theoretical evidence increasingly indicates that the highly flexible features of the RNA structure are affected by these factors, which include the flanking sequence context, physiochemical conditions, cis RNA-RNA interactions, and RNA interactions with other molecules. Furthermore, distinct RNA structures have been identified that govern almost all steps of biological processes in cells, including transcriptional activation and termination, transcriptional mutagenesis, 5'-capping, splicing, 3'-polyadenylation, mRNA export and localization, and translation. Here, we briefly summarize the dynamic and complex features of RNA folding along with a wide variety of intrinsic and extrinsic factors that affect RNA folding. We then provide several examples to elaborate RNA structure-mediated regulation at the transcriptional and posttranscriptional levels. Finally, we illustrate the regulatory roles of RNA structure and discuss advances pertaining to RNA structure in plants. WIREs RNA 2016, 7:562-574. doi: 10.1002/wrna.1350 For further resources related to this article, please visit the WIREs website. © 2016 Wiley Periodicals, Inc.

Phosphopeptidomics Reveals Differential Phosphorylation States and Novel SxE Phosphosite Motifs of Neuropeptides in Dense Core Secretory Vesicles

NASA Astrophysics Data System (ADS)

Lietz, Christopher B.; Toneff, Thomas; Mosier, Charles; Podvin, Sonia; O'Donoghue, Anthony J.; Hook, Vivian

2018-05-01

Neuropeptides are vital for cell-cell communication and function in the regulation of the nervous and endocrine systems. They are generated by post-translational modification (PTM) steps resulting in small active peptides generated from prohormone precursors. Phosphorylation is a significant PTM for the bioactivity of neuropeptides. From the known diversity of distinct neuropeptide functions, it is hypothesized that the extent of phosphorylation varies among different neuropeptides. To assess this hypothesis, neuropeptide-containing dense core secretory vesicles from bovine adrenal medullary chromaffin cells were subjected to global phosphopeptidomics analyses by liquid chromatography (LC)-mass spectrometry (MS/MS). Phosphopeptides were identified directly by LC-MS/MS and indirectly by phosphatase treatment followed by LC-MS/MS. The data identified numerous phosphorylated peptides derived from neuropeptide precursors such as chromogranins, secretogranins, proenkephalin and pro-NPY. Phosphosite occupancies were observed at high and low levels among identified peptides and many of the high occupancy phosphopeptides represent prohormone-derived peptides with currently unknown bioactivities. Peptide sequence analyses demonstrated SxE as the most prevalent phosphorylation site motif, corresponding to phosphorylation sites of the Fam20C protein kinase known to be present in the secretory pathway. The range of high to low phosphosite occupancies for neuropeptides demonstrates cellular regulation of neuropeptide phosphorylation. [Figure not available: see fulltext.

Antagonism between the transcription factors NANOG and OTX2 specifies rostral or caudal cell fate during neural patterning transition.

PubMed

Su, Zhenghui; Zhang, Yanqi; Liao, Baojian; Zhong, Xiaofen; Chen, Xin; Wang, Haitao; Guo, Yiping; Shan, Yongli; Wang, Lihui; Pan, Guangjin

2018-03-23

During neurogenesis, neural patterning is a critical step during which neural progenitor cells differentiate into neurons with distinct functions. However, the molecular determinants that regulate neural patterning remain poorly understood. Here we optimized the "dual SMAD inhibition" method to specifically promote differentiation of human pluripotent stem cells (hPSCs) into forebrain and hindbrain neural progenitor cells along the rostral-caudal axis. We report that neural patterning determination occurs at the very early stage in this differentiation. Undifferentiated hPSCs expressed basal levels of the transcription factor orthodenticle homeobox 2 (OTX2) that dominantly drove hPSCs into the "default" rostral fate at the beginning of differentiation. Inhibition of glycogen synthase kinase 3β (GSK3β) through CHIR99021 application sustained transient expression of the transcription factor NANOG at early differentiation stages through Wnt signaling. Wnt signaling and NANOG antagonized OTX2 and, in the later stages of differentiation, switched the default rostral cell fate to the caudal one. Our findings have uncovered a mutual antagonism between NANOG and OTX2 underlying cell fate decisions during neural patterning, critical for the regulation of early neural development in humans. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Phosphopeptidomics Reveals Differential Phosphorylation States and Novel SxE Phosphosite Motifs of Neuropeptides in Dense Core Secretory Vesicles

NASA Astrophysics Data System (ADS)

Lietz, Christopher B.; Toneff, Thomas; Mosier, Charles; Podvin, Sonia; O'Donoghue, Anthony J.; Hook, Vivian

2018-03-01

Neuropeptides are vital for cell-cell communication and function in the regulation of the nervous and endocrine systems. They are generated by post-translational modification (PTM) steps resulting in small active peptides generated from prohormone precursors. Phosphorylation is a significant PTM for the bioactivity of neuropeptides. From the known diversity of distinct neuropeptide functions, it is hypothesized that the extent of phosphorylation varies among different neuropeptides. To assess this hypothesis, neuropeptide-containing dense core secretory vesicles from bovine adrenal medullary chromaffin cells were subjected to global phosphopeptidomics analyses by liquid chromatography (LC)-mass spectrometry (MS/MS). Phosphopeptides were identified directly by LC-MS/MS and indirectly by phosphatase treatment followed by LC-MS/MS. The data identified numerous phosphorylated peptides derived from neuropeptide precursors such as chromogranins, secretogranins, proenkephalin and pro-NPY. Phosphosite occupancies were observed at high and low levels among identified peptides and many of the high occupancy phosphopeptides represent prohormone-derived peptides with currently unknown bioactivities. Peptide sequence analyses demonstrated SxE as the most prevalent phosphorylation site motif, corresponding to phosphorylation sites of the Fam20C protein kinase known to be present in the secretory pathway. The range of high to low phosphosite occupancies for neuropeptides demonstrates cellular regulation of neuropeptide phosphorylation. [Figure not available: see fulltext.

Modular Ligation Extension of Guide RNA Operons (LEGO) for Multiplexed dCas9 Regulation of Metabolic Pathways in Saccharomyces cerevisiae.

PubMed

Deaner, Matthew; Holzman, Allison; Alper, Hal S

2018-04-16

Metabolic engineering typically utilizes a suboptimal step-wise gene target optimization approach to parse a highly connected and regulated cellular metabolism. While the endonuclease-null CRISPR/Cas system has enabled gene expression perturbations without genetic modification, it has been mostly limited to small sets of gene targets in eukaryotes due to inefficient methods to assemble and express large sgRNA operons. In this work, we develop a TEF1p-tRNA expression system and demonstrate that the use of tRNAs as splicing elements flanking sgRNAs provides higher efficiency than both Pol III and ribozyme-based expression across a variety of single sgRNA and multiplexed contexts. Next, we devise and validate a scheme to allow modular construction of tRNA-sgRNA (TST) operons using an iterative Type IIs digestion/ligation extension approach, termed CRISPR-Ligation Extension of sgRNA Operons (LEGO). This approach enables facile construction of large TST operons. We demonstrate this utility by constructing a metabolic rewiring prototype for 2,3-butanediol production in 2 distinct yeast strain backgrounds. These results demonstrate that our approach can act as a surrogate for traditional genetic modification on a much shorter design-cycle timescale. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Different cucumber CsYUC genes regulate response to abiotic stresses and flower development.

PubMed

Yan, Shuangshuang; Che, Gen; Ding, Lian; Chen, Zijing; Liu, Xiaofeng; Wang, Hongyin; Zhao, Wensheng; Ning, Kang; Zhao, Jianyu; Tesfamichael, Kiflom; Wang, Qian; Zhang, Xiaolan

2016-02-09

The phytohormone auxin is essential for plant growth and development, and YUCCA (YUC) proteins catalyze a rate-limiting step for endogenous auxin biosynthesis. Despite YUC family genes have been isolated from several species, systematic expression analyses of YUCs in response to abiotic stress are lacking, and little is known about the function of YUC homologs in agricultural crops. Cucumber (Cucumis sativus L.) is a world cultivated vegetable crop with great economical and nutritional value. In this study, we isolated 10 YUC family genes (CsYUCs) from cucumber and explored their expression pattern under four types of stress treatments. Our data showed that CsYUC8 and CsYUC9 were specifically upregulated to elevate the auxin level under high temperature. CsYUC10b was dramatically increased but CsYUC4 was repressed in response to low temperature. CsYUC10a and CsYUC11 act against the upregulation of CsYUC10b under salinity stress, suggesting that distinct YUC members participate in different stress response, and may even antagonize each other to maintain the proper auxin levels in cucumber. Further, CsYUC11 was specifically expressed in the male flower in cucumber, and enhanced tolerance to salinity stress and regulated pedicel and stamen development through auxin biosynthesis in Arabidopsis.

Nucleosome regulatory dynamics in response to TGFβ

PubMed Central

Enroth, Stefan; Andersson, Robin; Bysani, Madhusudhan; Wallerman, Ola; Termén, Stefan; Tuch, Brian B.; De La Vega, Francisco M.; Heldin, Carl-Henrik; Moustakas, Aristidis; Komorowski, Jan; Wadelius, Claes

2014-01-01

Nucleosomes play important roles in a cell beyond their basal functionality in chromatin compaction. Their placement affects all steps in transcriptional regulation, from transcription factor (TF) binding to messenger ribonucleic acid (mRNA) synthesis. Careful profiling of their locations and dynamics in response to stimuli is important to further our understanding of transcriptional regulation by the state of chromatin. We measured nucleosome occupancy in human hepatic cells before and after treatment with transforming growth factor beta 1 (TGFβ1), using massively parallel sequencing. With a newly developed method, SuMMIt, for precise positioning of nucleosomes we inferred dynamics of the nucleosomal landscape. Distinct nucleosome positioning has previously been described at transcription start site and flanking TF binding sites. We found that the average pattern is present at very few sites and, in case of TF binding, the double peak surrounding the sites is just an artifact of averaging over many loci. We systematically searched for depleted nucleosomes in stimulated cells compared to unstimulated cells and identified 24 318 loci. Depending on genomic annotation, 44–78% of them were over-represented in binding motifs for TFs. Changes in binding affinity were verified for HNF4α by qPCR. Strikingly many of these loci were associated with expression changes, as measured by RNA sequencing. PMID:24771338

Extensive Use of RNA-Binding Proteins in Drosophila Sensory Neuron Dendrite Morphogenesis

PubMed Central

Olesnicky, Eugenia C.; Killian, Darrell J.; Garcia, Evelyn; Morton, Mary C.; Rathjen, Alan R.; Sola, Ismail E.; Gavis, Elizabeth R.

2013-01-01

The large number of RNA-binding proteins and translation factors encoded in the Drosophila and other metazoan genomes predicts widespread use of post-transcriptional regulation in cellular and developmental processes. Previous studies identified roles for several RNA-binding proteins in dendrite branching morphogenesis of Drosophila larval sensory neurons. To determine the larger contribution of post-transcriptional gene regulation to neuronal morphogenesis, we conducted an RNA interference screen to identify additional Drosophila proteins annotated as either RNA-binding proteins or translation factors that function in producing the complex dendritic trees of larval class IV dendritic arborization neurons. We identified 88 genes encoding such proteins whose knockdown resulted in aberrant dendritic morphology, including alterations in dendritic branch number, branch length, field size, and patterning of the dendritic tree. In particular, splicing and translation initiation factors were associated with distinct and characteristic phenotypes, suggesting that different morphogenetic events are best controlled at specific steps in post-transcriptional messenger RNA metabolism. Many of the factors identified in the screen have been implicated in controlling the subcellular distributions and translation of maternal messenger RNAs; thus, common post-transcriptional regulatory strategies may be used in neurogenesis and in the generation of asymmetry in the female germline and embryo. PMID:24347626

Deposition of an Ultraflat Graphene Oxide Nanosheet on Atomically Flat Substrates

NASA Astrophysics Data System (ADS)

Khan, M. Z. H.; Shahed, S. M. F.; Yuta, N.; Komeda, T.

2017-07-01

In this study, graphene oxide (GO) sheets produced in the form of stable aqueous dispersions were deposited on Au (111), freshly cleaved mica, and highly oriented pyrolytic graphite (HOPG) substrates. Atomic force microscopy (AFM) and scanning electron microscopy (SEM) were used to study the presence and distinct contact of GO sheets on the substrates. It was revealed from the topography images that high-quality ultraflat GO monolayer sheets formed on the substrates without distinct cracking/wrinkling or folding. GO sheets with apparent height variation observed by microscopy also indicate ultraflat deposition with clear underlying steps. It was observed that ultrasonication and centrifuge steps prior to deposition were very effective for getting oxidation debris (OD)-free ultraflat single monolayer GO nanosheets onto substrates and that the process depends on the concentration of supplied GO solutions.

PF-4/CXCL4 and CXCL4L1 exhibit distinct subcellular localization and a differentially regulated mechanism of secretion.

PubMed

Lasagni, Laura; Grepin, Renaud; Mazzinghi, Benedetta; Lazzeri, Elena; Meini, Claudia; Sagrinati, Costanza; Liotta, Francesco; Frosali, Francesca; Ronconi, Elisa; Alain-Courtois, Nathalie; Ballerini, Lara; Netti, Giuseppe Stefano; Maggi, Enrico; Annunziato, Francesco; Serio, Mario; Romagnani, Sergio; Bikfalvi, Andreas; Romagnani, Paola

2007-05-15

PF-4/CXCL4 is a member of the CXC chemokine family, which is mainly produced by platelets and known for its pleiotropic biological functions. Recently, the proteic product of a nonallelic variant gene of CXCL4 was isolated from human platelets and named as CXCL4L1. CXCL4L1 shows only 4.3% amino acid divergence in the mature protein, but exhibits a 38% amino acid divergence in the signal peptide region. We hypothesized that this may imply a difference in the cell type in which CXCL4L1 is expressed or a difference in its mode of secretion. In different types of transfected cells, CXCL4 and CXCL4L1 exhibited a distinct subcellular localization and a differential regulation of secretion, CXCL4 being stored in secretory granules and released in response to protein kinase C activation, whereas CXCL4L1 was continuously synthesized and secreted through a constitutive pathway. A protein kinase C-regulated CXCL4 secretion was observed also in lymphocytes, a cell type expressing mainly CXCL4 mRNA, whereas smooth muscle cells, which preferentially expressed CXCL4L1, exhibited a constitutive pathway of secretion. These results demonstrate that CXCL4 and CXCL4L1 exhibit a distinct subcellular localization and are secreted in a differentially regulated manner, suggesting distinct roles in inflammatory or homeostatic processes.

The Situated Dynamics of Purposes of Engagement and Self-Regulation Strategies: A Mixed-Methods Case Study of Writing

ERIC Educational Resources Information Center

Kaplan, Avi; Lichtinger, Einat; Margulis, Michal

2011-01-01

Background: Common conceptions of motivation and self-regulation view them as related but distinct entities. Most research on motivation and self-regulation investigates quantitative relations between level (e.g., self-efficacy) or type of motivation (e.g., mastery goals) and level of self-regulation. Purpose: Alternatively, the current study…

Identity-specific motivation: How distinct identities direct self-regulation across distinct situations.

PubMed

Browman, Alexander S; Destin, Mesmin; Molden, Daniel C

2017-12-01

Research on self-regulation has traditionally emphasized that people's thoughts and actions are guided by either (a) domain-general motivations that emerge from a cumulative history of life experiences, or (b) situation-specific motivations that emerge in immediate response to the incentives present in a particular context. However, more recent studies have illustrated the importance of understanding the interplay between such domain-general and situation-specific motivations across the types of contexts people regularly encounter. The present research, therefore, expands existing perspectives on self-regulation by investigating how people's identities -the internalized roles, relationships, and social group memberships that define who they are-systemically guide when and how different domain-general motivations are activated within specific types of situations. Using the motivational framework described by regulatory focus theory (Higgins, 1997), Studies 1 and 2 demonstrate that people indeed have distinct, identity-specific motivations that uniquely influence their current self-regulation when such identities are active. Studies 3-5 then begin to explore how identity-specific motivations are situated within people's larger self-concept. Studies 3a and 3b demonstrate that the less compatible people's specific identities, the more distinct are the motivations connected to those identities. Studies 4-5 then provide some initial, suggestive evidence that identity-specific motivations are not a separate, superordinate feature of people's identities that then alter how they pursue any subordinate, identity-relevant traits, but instead that such motivations emerge from the cumulative motivational significance of the subordinate traits to which the identities themselves become attached. Implications for understanding the role of the self-concept in self-regulation are discussed. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

Therapeutic Implications for Striatal-Enriched Protein Tyrosine Phosphatase (STEP) in Neuropsychiatric Disorders

PubMed Central

Goebel-Goody, Susan M.; Baum, Matthew; Paspalas, Constantinos D.; Fernandez, Stephanie M.; Carty, Niki C.; Kurup, Pradeep

2012-01-01

Striatal-enriched protein tyrosine phosphatase (STEP) is a brain-specific phosphatase that modulates key signaling molecules involved in synaptic plasticity and neuronal function. Targets include extracellular-regulated kinase 1 and 2 (ERK1/2), stress-activated protein kinase p38 (p38), the Src family tyrosine kinase Fyn, N-methyl-d-aspartate receptors (NMDARs), and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). STEP-mediated dephosphorylation of ERK1/2, p38, and Fyn leads to inactivation of these enzymes, whereas STEP-mediated dephosphorylation of surface NMDARs and AMPARs promotes their endocytosis. Accordingly, the current model of STEP function posits that it opposes long-term potentiation and promotes long-term depression. Phosphorylation, cleavage, dimerization, ubiquitination, and local translation all converge to maintain an appropriate balance of STEP in the central nervous system. Accumulating evidence over the past decade indicates that STEP dysregulation contributes to the pathophysiology of several neuropsychiatric disorders, including Alzheimer's disease, schizophrenia, fragile X syndrome, epileptogenesis, alcohol-induced memory loss, Huntington's disease, drug abuse, stroke/ischemia, and inflammatory pain. This comprehensive review discusses STEP expression and regulation and highlights how disrupted STEP function contributes to the pathophysiology of diverse neuropsychiatric disorders. PMID:22090472

A Latent Profile Analysis of University Students' Self-Regulated Learning Strategies

ERIC Educational Resources Information Center

Ning, Hoi Kwan; Downing, Kevin

2015-01-01

Based on self-reported cognitive, metacognitive, and behavioural strategy measures obtained from 828 final-year students from a university in Hong Kong, latent profile analysis (LPA) identified four distinct types of students with differential self-regulated learning strategy orientations: "competent self-regulated learners",…

Proteomic Identification of Differentially Expressed Proteins during Alfalfa (Medicago sativa L.) Flower Development.

PubMed

Chen, Lingling; Chen, Quanzhu; Zhu, Yanqiao; Hou, Longyu; Mao, Peisheng

2016-01-01

Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa ( Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase, and NADPH: quinone oxidoreductase-like protein. Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and contributes to the understanding of the basic molecular mechanisms during the alfalfa flowering process. These results may offer insight into potential strategies for improving seed yield, quality, and stress tolerance in alfalfa.

Proteomic Identification of Differentially Expressed Proteins during Alfalfa (Medicago sativa L.) Flower Development

PubMed Central

Chen, Lingling; Chen, Quanzhu; Zhu, Yanqiao; Hou, Longyu; Mao, Peisheng

2016-01-01

Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa (Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase, and NADPH: quinone oxidoreductase-like protein. Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and contributes to the understanding of the basic molecular mechanisms during the alfalfa flowering process. These results may offer insight into potential strategies for improving seed yield, quality, and stress tolerance in alfalfa. PMID:27757120

Early steps in protein synthesis and their regulation: a background study related to the biological effects of radiation. Progress report, July 1, 1975--June 30, 1976

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

Zamecnik, P.C.

This is a continuing study of protein synthesis, involving a search for the role of Ap/sub 4/A and other unusual nucleotides in growth regulation; studies of the mechanism of action of aminoacyl-tRNA ligases and the effect thereof on protein synthesis; a search for new regulators of the translation step, in cell-free systems; and an effort to improve the sensitivity and quantitation of chemical sequencing at the 3'-end of messenger RNA.

Understanding pain and coping in women with interstitial cystitis/bladder pain syndrome.

PubMed

Katz, Laura; Tripp, Dean A; Carr, Lesley K; Mayer, Robert; Moldwin, Robert M; Nickel, J Curtis

2017-08-01

To examine a self-regulation and coping model for interstitial cystitis/bladder pain syndrome (IC/BPS) that may help us understand the pain experience of patients with chronic IC/BPS. The model tested illness perceptions, illness-focused coping, emotional regulation, mental health and disability in a stepwise method using factor analysis and structural equation modelling. Step 1, explored the underlying constructs. Step 2, confirmed the measurement models to determine the structure/composition of the main constructs. Step 3, evaluated the model fit and specified pathways in the proposed IC/BPS self-regulation model. In all, 217 female patients with urologist diagnosed IC/BPS were recruited and diagnosed across tertiary care centres in North America. The data were collected through self-report questionnaires. An IC/BPS self-regulation model was supported. Physical disability was worsened by patient's negative perception of their illness, attempts to cope using illness-focused coping and poorer emotional regulation. Mental health was supported by perceptions that individuals could do something about their illness, using wellness-focused behavioural strategies and adaptive emotion regulation. The results clarify the complex and unique process of self-regulation in women with IC/BPS, implicating cognitive and coping targets, and highlighting emotional regulation. This knowledge should help clinicians understand and manage these patients' distress and disability. © 2017 The Authors BJU International © 2017 BJU International Published by John Wiley & Sons Ltd.

Program risk analysis handbook

NASA Technical Reports Server (NTRS)

Batson, R. G.

1987-01-01

NASA regulations specify that formal risk analysis be performed on a program at each of several milestones. Program risk analysis is discussed as a systems analysis approach, an iterative process (identification, assessment, management), and a collection of techniques. These techniques, which range from extremely simple to complex network-based simulation, are described in this handbook in order to provide both analyst and manager with a guide for selection of the most appropriate technique. All program risk assessment techniques are shown to be based on elicitation and encoding of subjective probability estimates from the various area experts on a program. Techniques to encode the five most common distribution types are given. Then, a total of twelve distinct approaches to risk assessment are given. Steps involved, good and bad points, time involved, and degree of computer support needed are listed. Why risk analysis should be used by all NASA program managers is discussed. Tools available at NASA-MSFC are identified, along with commercially available software. Bibliography (150 entries) and a program risk analysis check-list are provided.

CLIP-related methodologies and their application to retrovirology.

PubMed

Bieniasz, Paul D; Kutluay, Sebla B

2018-05-02

Virtually every step of HIV-1 replication and numerous cellular antiviral defense mechanisms are regulated by the binding of a viral or cellular RNA-binding protein (RBP) to distinct sequence or structural elements on HIV-1 RNAs. Until recently, these protein-RNA interactions were studied largely by in vitro binding assays complemented with genetics approaches. However, these methods are highly limited in the identification of the relevant targets of RBPs in physiologically relevant settings. Development of crosslinking-immunoprecipitation sequencing (CLIP) methodology has revolutionized the analysis of protein-nucleic acid complexes. CLIP combines immunoprecipitation of covalently crosslinked protein-RNA complexes with high-throughput sequencing, providing a global account of RNA sequences bound by a RBP of interest in cells (or virions) at near-nucleotide resolution. Numerous variants of the CLIP protocol have recently been developed, some with major improvements over the original. Herein, we briefly review these methodologies and give examples of how CLIP has been successfully applied to retrovirology research.

Some Lipid Droplets Are More Equal Than Others: Different Metabolic Lipid Droplet Pools in Hepatic Stellate Cells.

PubMed

Molenaar, Martijn R; Vaandrager, Arie B; Helms, J Bernd

2017-01-01

Hepatic stellate cells (HSCs) are professional lipid-storing cells and are unique in their property to store most of the retinol (vitamin A) as retinyl esters in large-sized lipid droplets. Hepatic stellate cell activation is a critical step in the development of chronic liver disease, as activated HSCs cause fibrosis. During activation, HSCs lose their lipid droplets containing triacylglycerols, cholesteryl esters, and retinyl esters. Lipidomic analysis revealed that the dynamics of disappearance of these different classes of neutral lipids are, however, very different from each other. Although retinyl esters steadily decrease during HSC activation, triacylglycerols have multiple pools one of which becomes transiently enriched in polyunsaturated fatty acids before disappearing. These observations are consistent with the existence of preexisting "original" lipid droplets with relatively slow turnover and rapidly recycling lipid droplets that transiently appear during activation of HSCs. Elucidation of the molecular machinery involved in the regulation of these distinct lipid droplet pools may open new avenues for the treatment of liver fibrosis.

Corynebacterium glutamicum promoters: a practical approach

PubMed Central

Pátek, Miroslav; Holátko, Jiří; Busche, Tobias; Kalinowski, Jörn; Nešvera, Jan

2013-01-01

Summary Transcription initiation is the key step in gene expression in bacteria, and it is therefore studied for both theoretical and practical reasons. Promoters, the traffic lights of transcription initiation, are used as construction elements in biotechnological efforts to coordinate ‘green waves’ in the metabolic pathways leading to the desired metabolites. Detailed analyses of Corynebacterium glutamicum promoters have already provided large amounts of data on their structures, regulatory mechanisms and practical capabilities in metabolic engineering. In this minireview the main aspects of promoter studies, the methods developed for their analysis and their practical use in C. glutamicum are discussed. These include definitions of the consensus sequences of the distinct promoter classes, promoter localization and characterization, activity measurements, the functions of transcriptional regulators and examples of practical uses of constitutive, inducible and modified promoters in biotechnology. The implications of the introduction of novel techniques, such as in vitro transcription and RNA sequencing, to C. glutamicum promoter studies are outlined. PMID:23305350

A dual affinity-tag strategy for the expression and purification of human linker histone H1.4 in Escherichia coli.

PubMed

Ryan, Daniel P; Tremethick, David J

2016-04-01

Linker histones are an abundant and critical component of the eukaryotic chromatin landscape. They play key roles in regulating the higher order structure of chromatin and many genetic processes. Higher eukaryotes possess a number of different linker histone subtypes and new data are consistently emerging that indicate these subtypes are functionally distinct. We were interested in studying one of the most abundant human linker histone subtypes, H1.4. We have produced recombinant full-length H1.4 in Escherichia coli. An N-terminal Glutathione-S-Transferase tag was used to promote soluble expression and was combined with a C-terminal hexahistidine tag to facilitate a simple non-denaturing two-step affinity chromatography procedure that results in highly pure full-length H1.4. The purified H1.4 was shown to be functional via in vitro chromatin assembly experiments and remains active after extended storage at -80 °C. Copyright © 2015 Elsevier Inc. All rights reserved.

The glucagon-miniglucagon interplay: a new level in the metabolic regulation.

PubMed

Bataille, Dominique; Fontés, Ghislaine; Costes, Safia; Longuet, Christine; Dalle, Stéphane

2006-07-01

Miniglucagon (glucagon 19-29) is the ultimate processing product of proglucagon, present in the glucagon-secreting granules of the alpha cells, at a close vicinity of the insulin-secreting beta cells. Co-released with glucagon and thanks to its original mode of action and its huge potency, it suppresses, inside the islet of Langerhans, the detrimental effect of glucagon on insulin secretion, while it leaves untouched the beneficial effect of glucagon on glucose competence of the beta cell. At the periphery, miniglucagon is processed at the surface of glucagon- and insulin-sensitive cells from circulating glucagon. At that level, it acts via a cellular pathway which uses initial molecular steps distinct from that of insulin which, when impaired, are involved in insulin resistence. This bypass allows miniglucagon to act as an insulin-like component, a characteristic which makes this peptide of particular interest from a pathophysiological and pharmacological point of views in understanding and treating metabolic diseases, such as the type 2 diabetes.

Bacterial Sigma Factors and Anti-Sigma Factors: Structure, Function and Distribution

PubMed Central

Paget, Mark S.

2015-01-01

Sigma factors are multi-domain subunits of bacterial RNA polymerase (RNAP) that play critical roles in transcription initiation, including the recognition and opening of promoters as well as the initial steps in RNA synthesis. This review focuses on the structure and function of the major sigma-70 class that includes the housekeeping sigma factor (Group 1) that directs the bulk of transcription during active growth, and structurally-related alternative sigma factors (Groups 2–4) that control a wide variety of adaptive responses such as morphological development and the management of stress. A recurring theme in sigma factor control is their sequestration by anti-sigma factors that occlude their RNAP-binding determinants. Sigma factors are then released through a wide variety of mechanisms, often involving branched signal transduction pathways that allow the integration of distinct signals. Three major strategies for sigma release are discussed: regulated proteolysis, partner-switching, and direct sensing by the anti-sigma factor. PMID:26131973

Allosteric substrate switching in a voltage-sensing lipid phosphatase.

PubMed

Grimm, Sasha S; Isacoff, Ehud Y

2016-04-01

Allostery provides a critical control over enzyme activity, biasing the catalytic site between inactive and active states. We found that the Ciona intestinalis voltage-sensing phosphatase (Ci-VSP), which modifies phosphoinositide signaling lipids (PIPs), has not one but two sequential active states with distinct substrate specificities, whose occupancy is allosterically controlled by sequential conformations of the voltage-sensing domain (VSD). Using fast fluorescence resonance energy transfer (FRET) reporters of PIPs to monitor enzyme activity and voltage-clamp fluorometry to monitor conformational changes in the VSD, we found that Ci-VSP switches from inactive to a PIP3-preferring active state when the VSD undergoes an initial voltage-sensing motion and then into a second PIP2-preferring active state when the VSD activates fully. This two-step allosteric control over a dual-specificity enzyme enables voltage to shape PIP concentrations in time, and provides a mechanism for the complex modulation of PIP-regulated ion channels, transporters, cell motility, endocytosis and exocytosis.

Allosteric substrate switching in a voltage sensing lipid phosphatase

PubMed Central

Grimm, Sasha S.; Isacoff, Ehud Y.

2016-01-01

Allostery provides a critical control over enzyme activity, biasing the catalytic site between inactive and active states. We find the Ciona intestinalis voltage-sensing phosphatase (Ci-VSP), which modifies phosphoinositide signaling lipids (PIPs), to have not one but two sequential active states with distinct substrate specificities, whose occupancy is allosterically controlled by sequential conformations of the voltage sensing domain (VSD). Using fast FRET reporters of PIPs to monitor enzyme activity and voltage clamp fluorometry to monitor conformational changes in the VSD, we find that Ci-VSP switches from inactive to a PIP3-preferring active state when the VSD undergoes an initial voltage sensing motion and then into a second PIP2-preferring active state when the VSD activates fully. This novel 2-step allosteric control over a dual specificity enzyme enables voltage to shape PIP concentrations in time, and provides a mechanism for the complex modulation of PIP-regulated ion channels, transporters, cell motility and endo/exocytosis. PMID:26878552

Renshaw cell interneuron specialization is controlled by a temporally restricted transcription factor program

PubMed Central

Stam, Floor J.; Hendricks, Timothy J.; Zhang, Jingming; Geiman, Eric J.; Francius, Cedric; Labosky, Patricia A.; Clotman, Frederic; Goulding, Martyn

2012-01-01

The spinal cord contains a diverse array of physiologically distinct interneuron cell types that subserve specialized roles in somatosensory perception and motor control. The mechanisms that generate these specialized interneuronal cell types from multipotential spinal progenitors are not known. In this study, we describe a temporally regulated transcriptional program that controls the differentiation of Renshaw cells (RCs), an anatomically and functionally discrete spinal interneuron subtype. We show that the selective activation of the Onecut transcription factors Oc1 and Oc2 during the first wave of V1 interneuron neurogenesis is a key step in the RC differentiation program. The development of RCs is additionally dependent on the forkhead transcription factor Foxd3, which is more broadly expressed in postmitotic V1 interneurons. Our demonstration that RCs are born, and activate Oc1 and Oc2 expression, in a narrow temporal window leads us to posit that neuronal diversity in the developing spinal cord is established by the composite actions of early spatial and temporal determinants. PMID:22115757

Uncovering the Atomistic Mechanism for Calcite Step Growth

DOE PAGES

De La Pierre, Marco; Raiteri, Paolo; Stack, Andrew G.; ...

2017-04-13

Determining a complete atomic-level picture of how minerals grow from aqueous solution remains a challenge as macroscopic rates can be a convolution of many reactions. For the case of calcite (CaCO 3) in water, computer simulations have been used in this paper to map the complex thermodynamic landscape leading to growth of the two distinct steps, acute and obtuse, on the basal surface. The carbonate ion is found to preferentially adsorb at the upper edge of acute steps while Ca 2+ only adsorbs after CO 3 2-. In contrast to the conventional picture, ion pairs prefer to bind at themore » upper edge of the step with only one ion, at most, coordinated to the step and lower terrace. Finally, migration of the first carbonate ion to a growth site is found to be rate-limiting for kink nucleation, with this process having a lower activation energy on the obtuse step.« less

Bipartite functions of the CREB co-activators selectively direct alternative splicing or transcriptional activation

PubMed Central

Amelio, Antonio L; Caputi, Massimo; Conkright, Michael D

2009-01-01

The CREB regulated transcription co-activators (CRTCs) regulate many biological processes by integrating and converting environmental inputs into transcriptional responses. Although the mechanisms by which CRTCs sense cellular signals are characterized, little is known regarding how CRTCs contribute to the regulation of cAMP inducible genes. Here we show that these dynamic regulators, unlike other co-activators, independently direct either pre-mRNA splice-site selection or transcriptional activation depending on the cell type or promoter context. Moreover, in other scenarios, the CRTC co-activators coordinately regulate transcription and splicing. Mutational analyses showed that CRTCs possess distinct functional domains responsible for regulating either pre-mRNA splicing or transcriptional activation. Interestingly, the CRTC1–MAML2 oncoprotein lacks the splicing domain and is incapable of altering splice-site selection despite robustly activating transcription. The differential usage of these distinct domains allows CRTCs to selectively mediate multiple facets of gene regulation, indicating that co-activators are not solely restricted to coordinating alternative splicing with increase in transcriptional activity. PMID:19644446

Distinct intracellular sAC-cAMP domains regulate ER Ca2+ signaling and OXPHOS function.

PubMed

Valsecchi, Federica; Konrad, Csaba; D'Aurelio, Marilena; Ramos-Espiritu, Lavoisier S; Stepanova, Anna; Burstein, Suzanne R; Galkin, Alexander; Magranè, Jordi; Starkov, Anatoly; Buck, Jochen; Levin, Lonny R; Manfredi, Giovanni

2017-11-01

cAMP regulates a wide variety of physiological functions in mammals. This single second messenger can regulate multiple, seemingly disparate functions within independently regulated cell compartments. We have previously identified one such compartment inside the matrix of the mitochondria, where soluble adenylyl cyclase (sAC) regulates oxidative phosphorylation (OXPHOS). We now show that sAC knockout fibroblasts have a defect in OXPHOS activity and attempt to compensate for this defect by increasing OXPHOS proteins. Importantly, sAC knockout cells also exhibit decreased probability of endoplasmic reticulum (ER) Ca 2+ release associated with diminished phosphorylation of the inositol 3-phosphate receptor. Restoring sAC expression exclusively in the mitochondrial matrix rescues OXPHOS activity and reduces mitochondrial biogenesis, indicating that these phenotypes are regulated by intramitochondrial sAC. In contrast, Ca 2+ release from the ER is only rescued when sAC expression is restored throughout the cell. Thus, we show that functionally distinct, sAC-defined, intracellular cAMP signaling domains regulate metabolism and Ca 2+ signaling. © 2017. Published by The Company of Biologists Ltd.

Multiple degradation pathways regulate versatile CIP/KIP CDK inhibitors.

PubMed

Starostina, Natalia G; Kipreos, Edward T

2012-01-01

The mammalian CIP/KIP family of cyclin-dependent kinase (CDK) inhibitors (CKIs) comprises three proteins--p21(Cip1/WAF1), p27(Kip1), and p57(Kip2)--that bind and inhibit cyclin-CDK complexes, which are key regulators of the cell cycle. CIP/KIP CKIs have additional independent functions in regulating transcription, apoptosis and actin cytoskeletal dynamics. These divergent functions are performed in distinct cellular compartments and contribute to the seemingly contradictory observation that the CKIs can both suppress and promote cancer. Multiple ubiquitin ligases (E3s) direct the proteasome-mediated degradation of p21, p27 and p57. This review analyzes recent data highlighting our current understanding of how distinct E3 pathways regulate subpopulations of the CKIs to control their diverse functions. Copyright © 2011 Elsevier Ltd. All rights reserved.

Proton electrochemical gradient: Driving and regulating neurotransmitter uptake.

PubMed

Farsi, Zohreh; Jahn, Reinhard; Woehler, Andrew

2017-05-01

Accumulation of neurotransmitters in the lumen of synaptic vesicles (SVs) relies on the activity of the vacuolar-type H + -ATPase. This pump drives protons into the lumen, generating a proton electrochemical gradient (Δμ H+ ) across the membrane. Recent work has demonstrated that the balance between the chemical (ΔpH) and electrical (ΔΨ) components of Δμ H+ is regulated differently by some distinct vesicle types. As different neurotransmitter transporters use ΔpH and ΔΨ with different relative efficiencies, regulation of this gradient balance has the potential to influence neurotransmitter uptake. Nevertheless, the underlying mechanisms responsible for this regulation remain poorly understood. In this review, we provide an overview of current neurotransmitter uptake models, with a particular emphasis on the distinct roles of the electrical and chemical gradients and current hypotheses for regulatory mechanisms. © 2017 WILEY Periodicals, Inc.

Behavior-specific changes in transcriptional modules lead to distinct and predictable neurogenomic states

PubMed Central

Chandrasekaran, Sriram; Ament, Seth A.; Eddy, James A.; Rodriguez-Zas, Sandra L.; Schatz, Bruce R.; Price, Nathan D.; Robinson, Gene E.

2011-01-01

Using brain transcriptomic profiles from 853 individual honey bees exhibiting 48 distinct behavioral phenotypes in naturalistic contexts, we report that behavior-specific neurogenomic states can be inferred from the coordinated action of transcription factors (TFs) and their predicted target genes. Unsupervised hierarchical clustering of these transcriptomic profiles showed three clusters that correspond to three ecologically important behavioral categories: aggression, maturation, and foraging. To explore the genetic influences potentially regulating these behavior-specific neurogenomic states, we reconstructed a brain transcriptional regulatory network (TRN) model. This brain TRN quantitatively predicts with high accuracy gene expression changes of more than 2,000 genes involved in behavior, even for behavioral phenotypes on which it was not trained, suggesting that there is a core set of TFs that regulates behavior-specific gene expression in the bee brain, and other TFs more specific to particular categories. TFs playing key roles in the TRN include well-known regulators of neural and behavioral plasticity, e.g., Creb, as well as TFs better known in other biological contexts, e.g., NF-κB (immunity). Our results reveal three insights concerning the relationship between genes and behavior. First, distinct behaviors are subserved by distinct neurogenomic states in the brain. Second, the neurogenomic states underlying different behaviors rely upon both shared and distinct transcriptional modules. Third, despite the complexity of the brain, simple linear relationships between TFs and their putative target genes are a surprisingly prominent feature of the networks underlying behavior. PMID:21960440

Subcellular localization, interactions and dynamics of the phage-shock protein-like Lia response in Bacillus subtilis.

PubMed

Domínguez-Escobar, Julia; Wolf, Diana; Fritz, Georg; Höfler, Carolin; Wedlich-Söldner, Roland; Mascher, Thorsten

2014-05-01

The liaIH operon of Bacillus subtilis is the main target of the envelope stress-inducible two-component system LiaRS. Here, we studied the localization, interaction and cellular dynamics of Lia proteins to gain insights into the physiological role of the Lia response. We demonstrate that LiaI serves as the membrane anchor for the phage-shock protein A homologue LiaH. Under non-inducing conditions, LiaI locates in highly motile membrane-associated foci, while LiaH is dispersed throughout the cytoplasm. Under stress conditions, both proteins are strongly induced and colocalize in numerous distinct static spots at the cytoplasmic membrane. This behaviour is independent of MreB and does also not correlate with the stalling of the cell wall biosynthesis machinery upon antibiotic inhibition. It can be induced by antibiotics that interfere with the membrane-anchored steps of cell wall biosynthesis, while compounds that inhibit the cytoplasmic or extracytoplasmic steps do not trigger this response. Taken together, our data are consistent with a model in which the Lia system scans the cytoplasmic membrane for envelope perturbations. Upon their detection, LiaS activates the cognate response regulator LiaR, which in turn strongly induces the liaIH operon. Simultaneously, LiaI recruits LiaH to the membrane, presumably to protect the envelope and counteract the antibiotic-induced damage. © 2014 John Wiley & Sons Ltd.

To eliminate the sporting purposes distinction in the gun laws.

THOMAS, 113th Congress

Rep. Collins, Doug [R-GA-9

2013-06-06

House - 07/15/2013 Referred to the Subcommittee on Crime, Terrorism, Homeland Security, and Investigations. (All Actions) Tracker: This bill has the status IntroducedHere are the steps for Status of Legislation:

Regulation of step frequency in transtibial amputee endurance athletes using a running-specific prosthesis.

PubMed

Oudenhoven, Laura M; Boes, Judith M; Hak, Laura; Faber, Gert S; Houdijk, Han

2017-01-25

Running specific prostheses (RSP) are designed to replicate the spring-like behaviour of the human leg during running, by incorporating a real physical spring in the prosthesis. Leg stiffness is an important parameter in running as it is strongly related to step frequency and running economy. To be able to select a prosthesis that contributes to the required leg stiffness of the athlete, it needs to be known to what extent the behaviour of the prosthetic leg during running is dominated by the stiffness of the prosthesis or whether it can be regulated by adaptations of the residual joints. The aim of this study was to investigate whether and how athletes with an RSP could regulate leg stiffness during distance running at different step frequencies. Seven endurance runners with an unilateral transtibial amputation performed five running trials on a treadmill at a fixed speed, while different step frequencies were imposed (preferred step frequency (PSF) and -15%, -7.5%, +7.5% and +15% of PSF). Among others, step time, ground contact time, flight time, leg stiffness and joint kinetics were measured for both legs. In the intact leg, increasing step frequency was accompanied by a decrease in both contact and flight time, while in the prosthetic leg contact time remained constant and only flight time decreased. In accordance, leg stiffness increased in the intact leg, but not in the prosthetic leg. Although a substantial contribution of the residual leg to total leg stiffness was observed, this contribution did not change considerably with changing step frequency. Amputee athletes do not seem to be able to alter prosthetic leg stiffness to regulate step frequency during running. This invariant behaviour indicates that RSP stiffness has a large effect on total leg stiffness and therefore can have an important influence on running performance. Nevertheless, since prosthetic leg stiffness was considerably lower than stiffness of the RSP, compliance of the residual leg should not be ignored when selecting RSP stiffness. Copyright © 2016 Elsevier Ltd. All rights reserved.

ERK/MAPK Regulates Hippocampal Histone Phosphorylation Following Contextual Fear Conditioning

ERIC Educational Resources Information Center

Levenson, Jonathan M.; Sweatt, J. David; Chwang, Wilson B.; O'Riordan, Kenneth J.

2006-01-01

Long-term memory formation is regulated by many distinct molecular mechanisms that control gene expression. An emerging model for effecting a stable, coordinated pattern of gene transcription involves epigenetic tagging through modifications of histones or DNA. In this study, we investigated the regulation of histone phosphorylation in the…

Seizure-Induced Regulations of Amyloid-β, STEP61, and STEP61 Substrates Involved in Hippocampal Synaptic Plasticity

PubMed Central

Jang, Sung-Soo; Royston, Sara E.; Lee, Gunhee; Wang, Shuwei; Chung, Hee Jung

2016-01-01

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive cognitive decline. Pathologic accumulation of soluble amyloid-β (Aβ) oligomers impairs synaptic plasticity and causes epileptic seizures, both of which contribute to cognitive dysfunction in AD. However, whether seizures could regulate Aβ-induced synaptic weakening remains unclear. Here we show that a single episode of electroconvulsive seizures (ECS) increased protein expression of membrane-associated STriatal-Enriched protein tyrosine Phosphatase (STEP61) and decreased tyrosine-phosphorylation of its substrates N-methyl D-aspartate receptor (NMDAR) subunit GluN2B and extracellular signal regulated kinase 1/2 (ERK1/2) in the rat hippocampus at 2 days following a single ECS. Interestingly, a significant decrease in ERK1/2 expression and an increase in APP and Aβ levels were observed at 3-4 days following a single ECS when STEP61 level returned to the baseline. Given that pathologic levels of Aβ increase STEP61 activity and STEP61-mediated dephosphorylation of GluN2B and ERK1/2 leads to NMDAR internalization and ERK1/2 inactivation, we propose that upregulation of STEP61 and downregulation of GluN2B and ERK1/2 phosphorylation mediate compensatory weakening of synaptic strength in response to acute enhancement of hippocampal network activity, whereas delayed decrease in ERK1/2 expression and increase in APP and Aβ expression may contribute to the maintenance of this synaptic weakening. PMID:27127657

NCKX3 was compensated by calcium transporting genes and bone resorption in a NCKX3 KO mouse model.

PubMed

Yang, Hyun; Ahn, Changhwan; Shin, Eun-Kyeong; Lee, Ji-Sun; An, Beum-Soo; Jeung, Eui-Bae

2017-10-15

Gene knockout is the most powerful tool for determination of gene function or permanent modification of the phenotypic characteristics of an animal. Existing methods for gene disruption are limited by their efficiency, time required for completion and potential for confounding off-target effects. In this study, a rapid single-step approach to knockout of a targeted gene in mice using zinc-finger nucleases (ZFNs) was demonstrated for generation of mutant (knockout; KO) alleles. Specifically, ZFNs to target the sodium/calcium/potassium exchanger3 (NCKX3) gene in C57bl/6j were designed using the concept of this approach. NCKX3 KO mice were generated and the phenotypic characterization and molecular regulation of active calcium transporting genes was assessed when mice were fed different calcium diets during growth. General phenotypes such as body weight and plasma ion level showed no distinct abnormalities. Thus, the potassium/sodium/calcium exchanger of NCKX3 KO mice proceeded normally in this study. As a result, the compensatory molecular regulation of this mechanism was elucidated. Renal TRPV5 mRNA of NCKX3 KO mice increased in both male and female mice. Expression of TRPV6 mRNA was only down-regulated in the duodenum of male KO mice. Renal- and duodenal expression of PTHR and VDR were not changed; however, GR mRNA expression was increased in the kidney of NCKX3 KO mice. Depletion of the NCKX3 gene in a KO mouse model showed loss of bone mineral contents and increased plasma parathyroid hormone, suggesting that NCKX3 may play a role in regulating calcium homeostasis. Copyright © 2017 Elsevier B.V. All rights reserved.

Regulation of bone mass through pineal-derived melatonin-MT2 receptor pathway.

PubMed

Sharan, Kunal; Lewis, Kirsty; Furukawa, Takahisa; Yadav, Vijay K

2017-09-01

Tryptophan, an essential amino acid through a series of enzymatic reactions gives rise to various metabolites, viz. serotonin and melatonin, that regulate distinct biological functions. We show here that tryptophan metabolism in the pineal gland favors bone mass accrual through production of melatonin, a pineal-derived neurohormone. Pineal gland-specific deletion of Tph1, the enzyme that catalyzes the first step in the melatonin biosynthesis lead to a decrease in melatonin levels and a low bone mass due to an isolated decrease in bone formation while bone resorption parameters remained unaffected. Skeletal analysis of the mice deficient in MT1 or MT2 melatonin receptors showed a low bone mass in MT2-/- mice while MT1-/- mice had a normal bone mass compared to the WT mice. This low bone mass in the MT2-/- mice was due to an isolated decrease in osteoblast numbers and bone formation. In vitro assays of the osteoblast cultures derived from the MT1-/- and MT2-/- mice showed a cell intrinsic defect in the proliferation, differentiation and mineralization abilities of MT2-/- osteoblasts compared to WT counterparts, and the mutant cells did not respond to melatonin addition. Finally, we demonstrate that daily oral administration of melatonin can increase bone accrual during growth and can cure ovariectomy-induced structural and functional degeneration of bone by specifically increasing bone formation. By identifying pineal-derived melatonin as a regulator of bone mass through MT2 receptors, this study expands the role played by tryptophan derivatives in the regulation of bone mass and underscores its therapeutic relevance in postmenopausal osteoporosis. © 2017 The Authors. Journal of Pineal Research Published by John Wiley & Sons Ltd.

Fusel Alcohols Regulate Translation Initiation by Inhibiting eIF2B to Reduce Ternary Complex in a Mechanism That May Involve Altering the Integrity and Dynamics of the eIF2B Body

PubMed Central

Taylor, Eleanor J.; Campbell, Susan G.; Griffiths, Christian D.; Reid, Peter J.; Slaven, John W.; Harrison, Richard J.; Sims, Paul F.G.; Pavitt, Graham D.; Delneri, Daniela

2010-01-01

Recycling of eIF2-GDP to the GTP-bound form constitutes a core essential, regulated step in eukaryotic translation. This reaction is mediated by eIF2B, a heteropentameric factor with important links to human disease. eIF2 in the GTP-bound form binds to methionyl initiator tRNA to form a ternary complex, and the levels of this ternary complex can be a critical determinant of the rate of protein synthesis. Here we show that eIF2B serves as the target for translation inhibition by various fusel alcohols in yeast. Fusel alcohols are endpoint metabolites from amino acid catabolism, which signal nitrogen scarcity. We show that the inhibition of eIF2B leads to reduced ternary complex levels and that different eIF2B subunit mutants alter fusel alcohol sensitivity. A DNA tiling array strategy was developed that overcame difficulties in the identification of these mutants where the phenotypic distinctions were too subtle for classical complementation cloning. Fusel alcohols also lead to eIF2α dephosphorylation in a Sit4p-dependent manner. In yeast, eIF2B occupies a large cytoplasmic body where guanine nucleotide exchange on eIF2 can occur and be regulated. Fusel alcohols impact on both the movement and dynamics of this 2B body. Overall, these results confirm that the guanine nucleotide exchange factor, eIF2B, is targeted by fusel alcohols. Moreover, they highlight a potential connection between the movement or integrity of the 2B body and eIF2B regulation. PMID:20444979

Conceptual framework of the eco-physiological phases of insect diapause development justified by transcriptomic profiling

PubMed Central

Štětina, Tomáš; Poupardin, Rodolphe; Korbelová, Jaroslava; Bruce, Alexander William

2017-01-01

Insects often overcome unfavorable seasons in a hormonally regulated state of diapause during which their activity ceases, development is arrested, metabolic rate is suppressed, and tolerance of environmental stress is bolstered. Diapausing insects pass through a stereotypic succession of eco-physiological phases termed “diapause development.” The phasing is varied in the literature, and the whole concept is sometimes criticized as being too artificial. Here we present the results of transcriptional profiling using custom microarrays representing 1,042 genes in the drosophilid fly, Chymomyza costata. Fully grown, third-instar larvae programmed for diapause by a photoperiodic (short-day) signal were assayed as they traversed the diapause developmental program. When analyzing the gradual dynamics in the transcriptomic profile, we could readily distinguish distinct diapause developmental phases associated with induction/initiation, maintenance, cold acclimation, and termination by cold or by photoperiodic signal. Accordingly, each phase is characterized by a specific pattern of gene expression, supporting the physiological relevance of the concept of diapause phasing. Further, we have dissected in greater detail the changes in transcript levels of elements of several signaling pathways considered critical for diapause regulation. The phase of diapause termination is associated with enhanced transcript levels in several positive elements stimulating direct development (the 20-hydroxyecdysone pathway: Ecr, Shd, Broad; the Wnt pathway: basket, c-jun) that are countered by up-regulation in some negative elements (the insulin-signaling pathway: Ilp8, PI3k, Akt; the target of rapamycin pathway: Tsc2 and 4EBP; the Wnt pathway: shaggy). We speculate such up-regulations may represent the early steps linked to termination of diapause programming. PMID:28720705

29 CFR 1952.384 - Completed developmental steps.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 29 Labor 9 2010-07-01 2010-07-01 false Completed developmental steps. 1952.384 Section 1952.384 Labor Regulations Relating to Labor (Continued) OCCUPATIONAL SAFETY AND HEALTH ADMINISTRATION....384 Completed developmental steps. (a) In accordance with the requirements of § 1952.10, Puerto Rico's...

When the Brain Takes 'BOLD' Steps: Real-Time fMRI Neurofeedback Can Further Enhance the Ability to Gradually Self-regulate Regional Brain Activation.

PubMed

Sorger, Bettina; Kamp, Tabea; Weiskopf, Nikolaus; Peters, Judith Caroline; Goebel, Rainer

2018-05-15

Brain-computer interfaces (BCIs) based on real-time functional magnetic resonance imaging (rtfMRI) are currently explored in the context of developing alternative (motor-independent) communication and control means for the severely disabled. In such BCI systems, the user encodes a particular intention (e.g., an answer to a question or an intended action) by evoking specific mental activity resulting in a distinct brain state that can be decoded from fMRI activation. One goal in this context is to increase the degrees of freedom in encoding different intentions, i.e., to allow the BCI user to choose from as many options as possible. Recently, the ability to voluntarily modulate spatial and/or temporal blood oxygenation level-dependent (BOLD)-signal features has been explored implementing different mental tasks and/or different encoding time intervals, respectively. Our two-session fMRI feasibility study systematically investigated for the first time the possibility of using magnitudinal BOLD-signal features for intention encoding. Particularly, in our novel paradigm, participants (n=10) were asked to alternately self-regulate their regional brain-activation level to 30%, 60% or 90% of their maximal capacity by applying a selected activation strategy (i.e., performing a mental task, e.g., inner speech) and modulation strategies (e.g., using different speech rates) suggested by the experimenters. In a second step, we tested the hypothesis that the additional availability of feedback information on the current BOLD-signal level within a region of interest improves the gradual-self regulation performance. Therefore, participants were provided with neurofeedback in one of the two fMRI sessions. Our results show that the majority of the participants were able to gradually self-regulate regional brain activation to at least two different target levels even in the absence of neurofeedback. When provided with continuous feedback on their current BOLD-signal level, most participants further enhanced their gradual self-regulation ability. Our findings were observed across a wide variety of mental tasks and across clinical MR field strengths (i.e., at 1.5T and 3T), indicating that these findings are robust and can be generalized across mental tasks and scanner types. The suggested novel parametric activation paradigm enriches the spectrum of current rtfMRI-neurofeedback and BCI methodology and has considerable potential for fundamental and clinical neuroscience applications. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

The role of self-regulated learning in explaining examination performance of college students in first-semester general chemistry

NASA Astrophysics Data System (ADS)

Beckley, Scott

Many college students struggle with first-semester general chemistry. Prior studies have shown that a student's prior knowledge of chemistry, a cognitive factor, does not account for the total variance when measured by examination scores. This study explored the role of self-regulated learning (SRL) to identify the degree of success or failure of students with two outcome variables (i.e., American Chemical Society Comprehensive First-Term General Chemistry Examination (Form 2009) and hour-examination averages). The SRL construct consists of three interrelated components (i.e., cognitive, metacognitive, and motivational). SRL theory focuses on the idea of reciprocal determinism, in which the impact of one component of self-regulation affects the other two components. In the quantitative portion of this mixed methods study, eight measures of SRL were used to determine the `level' of self-regulation for each student. SRL variables were used in regression analysis and provided additional and unique variances. Cluster analysis techniques identified two distinct groups of students (i.e., adaptive and maladaptive). Generally, adaptive learners were associated with higher levels of SRL and success in the course; maladaptive learners had lower levels of SRL and struggled with the course demands. For the qualitative portion of the study, student volunteers (n = 8) were interviewed to gauge their views on the role of instruction in influencing their examination performances. The findings indicated that perceptions of teaching methods, demands of the course, course structure, feedback, and assessments were associated with the students' levels of self-regulation. Interviews revealed four SRL styles. Rote memorizers tended to fragment instruction and then memorize each fragment, while algorithmic memorizers tended to imitate the step-by-step problem-solving strategies of the instructor or the textbook. Globalizers were intrinsically motivated to learn the material but tended to focus on learning topics that were of practical interest to them, while conceptualizers tended to use flexible problem-solving approaches. The interviewees' actual outcome scores were compared to their predicted scores as generated from the regression models. In general, the actual scores of rote and algorithmic memorizers were lower than their predicted scores, while the actual scores for globalizers and conceptualizers were higher than predicted. From a teaching perspective, this study provided direct relationships between SRL and academic performance. This study also identified SRL styles and found evidence to support that differences in these styles influence students' performances.

Distinct regulation of alternative polyadenylation and gene expression by nuclear poly(A) polymerases

PubMed Central

Li, Wencheng; Laishram, Rakesh S.; Hoque, Mainul; Ji, Zhe

2017-01-01

Abstract Polyadenylation of nascent RNA by poly(A) polymerase (PAP) is important for 3′ end maturation of almost all eukaryotic mRNAs. Most mammalian genes harbor multiple polyadenylation sites (PASs), leading to expression of alternative polyadenylation (APA) isoforms with distinct functions. How poly(A) polymerases may regulate PAS usage and hence gene expression is poorly understood. Here, we show that the nuclear canonical (PAPα and PAPγ) and non-canonical (Star-PAP) PAPs play diverse roles in PAS selection and gene expression. Deficiencies in the PAPs resulted in perturbations of gene expression, with Star-PAP impacting lowly expressed mRNAs and long-noncoding RNAs to the greatest extent. Importantly, different PASs of a gene are distinctly regulated by different PAPs, leading to widespread relative expression changes of APA isoforms. The location and surrounding sequence motifs of a PAS appear to differentiate its regulation by the PAPs. We show Star-PAP-specific PAS usage regulates the expression of the eukaryotic translation initiation factor EIF4A1, the tumor suppressor gene PTEN and the long non-coding RNA NEAT1. The Star-PAP-mediated APA of PTEN is essential for DNA damage-induced increase of PTEN protein levels. Together, our results reveal a PAS-guided and PAP-mediated paradigm for gene expression in response to cellular signaling cues. PMID:28911096

Functional dissection of the paired domain of Pax6 reveals molecular mechanisms of coordinating neurogenesis and proliferation

PubMed Central

Walcher, Tessa; Xie, Qing; Sun, Jian; Irmler, Martin; Beckers, Johannes; Öztürk, Timucin; Niessing, Dierk; Stoykova, Anastassia; Cvekl, Ales; Ninkovic, Jovica; Götz, Magdalena

2013-01-01

To achieve adequate organ development and size, cell proliferation and differentiation have to be tightly regulated and coordinated. The transcription factor Pax6 regulates patterning, neurogenesis and proliferation in forebrain development. The molecular basis of this regulation is not well understood. As the bipartite DNA-binding paired domain of Pax6 regulates forebrain development, we examined mice with point mutations in its individual DNA-binding subdomains PAI (Pax6Leca4, N50K) and RED (Pax6Leca2, R128C). This revealed distinct roles in regulating proliferation in the developing cerebral cortex, with the PAI and RED subdomain mutations reducing and increasing, respectively, the number of mitoses. Conversely, neurogenesis was affected only by the PAI subdomain mutation, phenocopying the neurogenic defects observed in full Pax6 mutants. Genome-wide expression profiling identified molecularly discrete signatures of Pax6Leca4 and Pax6Leca2 mutations. Comparison to Pax6 targets identified by chromatin immunoprecipitation led to the identification and functional characterization of distinct DNA motifs in the promoters of target genes dysregulated in the Pax6Leca2 or Pax6Leca4 mutants, further supporting the distinct regulatory functions of the DNA-binding subdomains. Thus, Pax6 achieves its key roles in the developing forebrain by utilizing particular subdomains to coordinate patterning, neurogenesis and proliferation simultaneously. PMID:23404109

Acetylation within the N- and C-Terminal Domains of Src Regulates Distinct Roles of STAT3-Mediated Tumorigenesis.

PubMed

Huang, Chao; Zhang, Zhe; Chen, Lihan; Lee, Hank W; Ayrapetov, Marina K; Zhao, Ting C; Hao, Yimei; Gao, Jinsong; Yang, Chunzhang; Mehta, Gautam U; Zhuang, Zhengping; Zhang, Xiaoren; Hu, Guohong; Chin, Y Eugene

2018-06-01

Posttranslational modifications of mammalian c-Src N-terminal and C-terminal domains regulate distinct functions. Myristoylation of G 2 controls its cell membrane association and phosphorylation of Y419/Y527 controls its activation or inactivation, respectively. We provide evidence that Src-cell membrane association-dissociation and catalytic activation-inactivation are both regulated by acetylation. In EGF-treated cells, CREB binding protein (CBP) acetylates an N-terminal lysine cluster (K5, K7, and K9) of c-Src to promote dissociation from the cell membrane. CBP also acetylates the C-terminal K401, K423, and K427 of c-Src to activate intrinsic kinase activity for STAT3 recruitment and activation. N-terminal domain phosphorylation (Y14, Y45, and Y68) of STAT3 by c-Src activates transcriptionally active dimers of STAT3. Moreover, acetyl-Src translocates into nuclei, where it forms the Src-STAT3 enhanceosome for gene regulation and cancer cell proliferation. Thus, c-Src acetylation in the N-terminal and C-terminal domains play distinct roles in Src activity and regulation. Significance: CBP-mediated acetylation of lysine clusters in both the N-terminal and C-terminal regions of c-Src provides additional levels of control over STAT3 transcriptional activity. Cancer Res; 78(11); 2825-38. ©2018 AACR . ©2018 American Association for Cancer Research.

Maintaining protein composition in cilia.

PubMed

Stephen, Louise A; Elmaghloob, Yasmin; Ismail, Shehab

2017-12-20

The primary cilium is a sensory organelle that is vital in regulating several signalling pathways. Unlike most organelles cilia are open to the rest of the cell, not enclosed by membranes. The distinct protein composition is crucial to the function of cilia and many signalling proteins and receptors are specifically concentrated within distinct compartments. To maintain this composition, a mechanism is required to deliver proteins to the cilium whilst another must counter the entropic tendency of proteins to distribute throughout the cell. The combination of the two mechanisms should result in the concentration of ciliary proteins to the cilium. In this review we will look at different cellular mechanisms that play a role in maintaining the distinct composition of cilia, including regulation of ciliary access and trafficking of ciliary proteins to, from and within the cilium.

Affective lability and difficulties with regulation are differentially associated with amygdala and prefrontal response in women with Borderline Personality Disorder

PubMed Central

Silvers, Jennifer A.; Hubbard, Alexa D.; Biggs, Emily; Shu, Jocelyn; Fertuck, Eric; Chaudhury, Sadia; Grunebaum, Michael F.; Weber, Jochen; Kober, Hedy; Chesin, Megan; Brodsky, Beth S.; Koenigsberg, Harold; Ochsner, Kevin N.; Stanley, Barbara

2016-01-01

The present neuroimaging study investigated two aspects of difficulties with emotion associated with Borderline Personality Disorder (BPD1): affective lability and difficulty regulating emotion. While these two characteristics have been previously linked to BPD symptomology, it remains unknown whether individual differences in affective lability and emotion regulation difficulties are subserved by distinct neural substrates within a BPD sample. To address this issue, sixty women diagnosed with BPD were scanned while completing a task that assessed baseline emotional reactivity as well as top-down emotion regulation. More affective instability, as measured by the Affective Lability Scale (ALS2), positively correlated with greater amygdala responses on trials assessing emotional reactivity. Greater difficulties with regulating emotion, as measured by the Difficulties with Emotion Regulation Scale (DERS3), was negatively correlated with left inferior frontal gyrus (IFG4) recruitment on trials assessing regulatory ability. These findings suggest that, within a sample of individuals with BPD, greater bottom-up amygdala activity is associated with heightened affective lability. By contrast, difficulties with emotion regulation are related to reduced IFG recruitment during emotion regulation. These results point to distinct neural mechanisms for different aspects of BPD symptomology. PMID:27379614

RNA regulators responding to ribosomal protein S15 are frequent in sequence space

PubMed Central

Slinger, Betty L.; Meyer, Michelle M.

2016-01-01

There are several natural examples of distinct RNA structures that interact with the same ligand to regulate the expression of homologous genes in different organisms. One essential question regarding this phenomenon is whether such RNA regulators are the result of convergent or divergent evolution. Are the RNAs derived from some common ancestor and diverged to the point where we cannot identify the similarity, or have multiple solutions to the same biological problem arisen independently? A key variable in assessing these alternatives is how frequently such regulators arise within sequence space. Ribosomal protein S15 is autogenously regulated via an RNA regulator in many bacterial species; four apparently distinct regulators have been functionally validated in different bacterial phyla. Here, we explore how frequently such regulators arise within a partially randomized sequence population. We find many RNAs that interact specifically with ribosomal protein S15 from Geobacillus kaustophilus with biologically relevant dissociation constants. Furthermore, of the six sequences we characterize, four show regulatory activity in an Escherichia coli reporter assay. Subsequent footprinting and mutagenesis analysis indicates that protein binding proximal to regulatory features such as the Shine–Dalgarno sequence is sufficient to enable regulation, suggesting that regulation in response to S15 is relatively easily acquired. PMID:27580716

Overlapping and Distinct Molecular Determinants Dictating the Antiviral Activities of TRIM56 against Flaviviruses and Coronavirus

PubMed Central

Liu, Baoming; Li, Nan L.; Wang, Jie; Shi, Pei-Yong; Wang, Tianyi; Miller, Mark A.

2014-01-01

ABSTRACT The tripartite motif-containing (TRIM) proteins have emerged as a new class of host antiviral restriction factors, with several demonstrating roles in regulating innate antiviral responses. Of >70 known TRIMs, TRIM56 inhibits replication of bovine viral diarrhea virus, a ruminant pestivirus of the family Flaviviridae, but has no appreciable effect on vesicular stomatitis virus (VSV), a rhabdovirus. Yet the antiviral spectrum of TRIM56 remains undefined. In particular, how TRIM56 impacts human-pathogenic viruses is unknown. Also unclear are the molecular determinants governing the antiviral activities of TRIM56. Herein, we show that TRIM56 poses a barrier to infections by yellow fever virus (YFV), dengue virus serotype 2 (DENV2), and human coronavirus virus (HCoV) OC43 but not encephalomyocarditis virus (EMCV). Moreover, by engineering cell lines conditionally expressing various TRIM56 mutants, we demonstrated that TRIM56's antiflavivirus effects required both the E3 ligase activity that lies in the N-terminal RING domain and the integrity of its C-terminal portion, while the restriction of HCoV-OC43 relied upon the TRIM56 E3 ligase activity alone. Furthermore, TRIM56 was revealed to impair YFV and DENV2 propagation by suppressing intracellular viral RNA accumulation but to compromise HCoV-OC43 infection at a later step in the viral life cycle, suggesting that distinct TRIM56 domains accommodate differing antiviral mechanisms. Altogether, TRIM56 is a versatile antiviral host factor that confers resistance to YFV, DENV2, and HCoV-OC43 through overlapping and distinct molecular determinants. IMPORTANCE We previously reported tripartite motif protein 56 (TRIM56) as a host restriction factor of bovine viral diarrhea virus, a ruminant pathogen. However, the impact of TRIM56 on human-pathogenic RNA viruses is unknown. Herein, we demonstrate that TRIM56 restricts two medically important flaviviruses, yellow fever virus (YFV) and dengue virus serotype 2 (DENV2), and a human coronavirus, HCoV-OC43, but not encephalomyocarditis virus, a picornavirus. Further, we show that TRIM56-mediated inhibition of HCoV-OC43 multiplication depends solely on its E3 ligase activity, whereas its restriction of YFV and DENV2 requires both the E3 ligase activity and integrity of the C-terminal portion. The differing molecular determinants appear to accommodate distinct antiviral mechanisms TRIM56 adopts to target different families of viruses; while TRIM56 curbs intracellular YFV/DENV2 RNA replication, it acts at a later step in HCoV-OC43 life cycle. These novel findings illuminate the molecular basis of the versatility and specificity of TRIM56's antiviral activities against positive-strand RNA viruses. PMID:25253338

Overlapping and distinct molecular determinants dictating the antiviral activities of TRIM56 against flaviviruses and coronavirus.

PubMed

Liu, Baoming; Li, Nan L; Wang, Jie; Shi, Pei-Yong; Wang, Tianyi; Miller, Mark A; Li, Kui

2014-12-01

The tripartite motif-containing (TRIM) proteins have emerged as a new class of host antiviral restriction factors, with several demonstrating roles in regulating innate antiviral responses. Of >70 known TRIMs, TRIM56 inhibits replication of bovine viral diarrhea virus, a ruminant pestivirus of the family Flaviviridae, but has no appreciable effect on vesicular stomatitis virus (VSV), a rhabdovirus. Yet the antiviral spectrum of TRIM56 remains undefined. In particular, how TRIM56 impacts human-pathogenic viruses is unknown. Also unclear are the molecular determinants governing the antiviral activities of TRIM56. Herein, we show that TRIM56 poses a barrier to infections by yellow fever virus (YFV), dengue virus serotype 2 (DENV2), and human coronavirus virus (HCoV) OC43 but not encephalomyocarditis virus (EMCV). Moreover, by engineering cell lines conditionally expressing various TRIM56 mutants, we demonstrated that TRIM56's antiflavivirus effects required both the E3 ligase activity that lies in the N-terminal RING domain and the integrity of its C-terminal portion, while the restriction of HCoV-OC43 relied upon the TRIM56 E3 ligase activity alone. Furthermore, TRIM56 was revealed to impair YFV and DENV2 propagation by suppressing intracellular viral RNA accumulation but to compromise HCoV-OC43 infection at a later step in the viral life cycle, suggesting that distinct TRIM56 domains accommodate differing antiviral mechanisms. Altogether, TRIM56 is a versatile antiviral host factor that confers resistance to YFV, DENV2, and HCoV-OC43 through overlapping and distinct molecular determinants. We previously reported tripartite motif protein 56 (TRIM56) as a host restriction factor of bovine viral diarrhea virus, a ruminant pathogen. However, the impact of TRIM56 on human-pathogenic RNA viruses is unknown. Herein, we demonstrate that TRIM56 restricts two medically important flaviviruses, yellow fever virus (YFV) and dengue virus serotype 2 (DENV2), and a human coronavirus, HCoV-OC43, but not encephalomyocarditis virus, a picornavirus. Further, we show that TRIM56-mediated inhibition of HCoV-OC43 multiplication depends solely on its E3 ligase activity, whereas its restriction of YFV and DENV2 requires both the E3 ligase activity and integrity of the C-terminal portion. The differing molecular determinants appear to accommodate distinct antiviral mechanisms TRIM56 adopts to target different families of viruses; while TRIM56 curbs intracellular YFV/DENV2 RNA replication, it acts at a later step in HCoV-OC43 life cycle. These novel findings illuminate the molecular basis of the versatility and specificity of TRIM56's antiviral activities against positive-strand RNA viruses. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Contributions of the three CYP1 monooxygenases to pro-inflammatory and inflammation-resolution lipid mediator pathways.

PubMed

Divanovic, Senad; Dalli, Jesmond; Jorge-Nebert, Lucia F; Flick, Leah M; Gálvez-Peralta, Marina; Boespflug, Nicholas D; Stankiewicz, Traci E; Fitzgerald, Jonathan M; Somarathna, Maheshika; Karp, Christopher L; Serhan, Charles N; Nebert, Daniel W

2013-09-15

All three cytochrome P450 1 (CYP1) monooxygenases are believed to participate in lipid mediator biosynthesis and/or their local inactivation; however, distinct metabolic steps are unknown. We used multiple-reaction monitoring and liquid chromatography-UV coupled with tandem mass spectrometry-based lipid-mediator metabololipidomics to identify and quantify three lipid-mediator metabolomes in basal peritoneal and zymosan-stimulated inflammatory exudates, comparing Cyp1a1/1a2/1b1(⁻/⁻) C57BL/6J-background triple-knockout mice with C57BL/6J wild-type mice. Significant differences between untreated triple-knockout and wild-type mice were not found for peritoneal cell number or type or for basal CYP1 activities involving 11 identified metabolic steps. Following zymosan-initiated inflammation, 18 lipid mediators were identified, including members of the eicosanoids and specialized proresolving mediators (i.e., resolvins and protectins). Compared with wild-type mice, Cyp1 triple-knockout mice exhibited increased neutrophil recruitment in zymosan-treated peritoneal exudates. Zymosan stimulation was associated with eight statistically significantly altered metabolic steps: increased arachidonic acid-derived leukotriene B₄ (LTB₄) and decreased 5S-hydroxyeicosatetraenoic acid; decreased docosahexaenoic acid-derived neuroprotectin D1/protectin D1, 17S-hydroxydocosahexaenoic acid, and 14S-hydroxydocosahexaenoic acid; and decreased eicosapentaenoic acid-derived 18R-hydroxyeicosapentaenoic acid (HEPE), 15S-HEPE, and 12S-HEPE. In neutrophils analyzed ex vivo, elevated LTB₄ levels were shown to parallel increased neutrophil numbers, and 20-hydroxy-LTB₄ formation was found to be deficient in Cyp1 triple-knockout mice. Together, these results demonstrate novel contributions of CYP1 enzymes to the local metabolite profile of lipid mediators that regulate neutrophilic inflammation.

Rapid and Efficient Isolation of High-Quality Small RNAs from Recalcitrant Plant Species Rich in Polyphenols and Polysaccharides

PubMed Central

Pu, Jinji; Guo, Jianrong; Fan, Zaifeng

2014-01-01

Small RNAs, including microRNAs (miRNAs) and small interfering RNAs (siRNAs), are important regulators of plant development and gene expression. The acquisition of high-quality small RNAs is the first step in the study of its expression and function analysis, yet the extraction method of small RNAs in recalcitrant plant tissues with various secondary metabolites is not well established, especially for tropical and subtropical plant species rich in polysaccharides and polyphenols. Here, we developed a simple and efficient method for high quality small RNAs extraction from recalcitrant plant species. Prior to RNA isolation, a precursory step with a CTAB-PVPP buffer system could efficiently remove compounds and secondary metabolites interfering with RNAs from homogenized lysates. Then, total RNAs were extracted by Trizol reagents followed by a differential precipitation of high-molecular-weight (HMW) RNAs using polyethylene glycol (PEG) 8000. Finally, small RNAs could be easily recovered from supernatant by ethanol precipitation without extra elimination steps. The isolated small RNAs from papaya showed high quality through a clear background on gel and a distinct northern blotting signal with miR159a probe, compared with other published protocols. Additionally, the small RNAs extracted from papaya were successfully used for validation of both predicted miRNAs and the putative conserved tasiARFs. Furthermore, the extraction method described here was also tested with several other subtropical and tropical plant tissues. The purity of the isolated small RNAs was sufficient for such applications as end-point stem-loop RT-PCR and northern blotting analysis, respectively. The simple and feasible extraction method reported here is expected to have excellent potential for isolation of small RNAs from recalcitrant plant tissues rich in polyphenols and polysaccharides. PMID:24787387

Functional complementation of anthocyanin sequestration in the vacuole by widely divergent glutathione S-transferases.

PubMed Central

Alfenito, M R; Souer, E; Goodman, C D; Buell, R; Mol, J; Koes, R; Walbot, V

1998-01-01

Glutathione S-transferases (GSTs) traditionally have been studied in plants and other organisms for their ability to detoxify chemically diverse herbicides and other toxic organic compounds. Anthocyanins are among the few endogenous substrates of plant GSTs that have been identified. The Bronze2 (Bz2) gene encodes a type III GST and performs the last genetically defined step of the maize anthocyanin pigment pathway. This step is the conjugation of glutathione to cyanidin 3-glucoside (C3G). Glutathionated C3G is transported to the vacuole via a tonoplast Mg-ATP-requiring glutathione pump (GS-X pump). Genetically, the comparable step in the petunia anthocyanin pathway is controlled by the Anthocyanin9 (An9) gene. An9 was cloned by transposon tagging and found to encode a type I plant GST. Bz2 and An9 have evolved independently from distinct types of GSTs, but each is regulated by the conserved transcriptional activators of the anthocyanin pathway. Here, a phylogenetic analysis is presented, with special consideration given to the origin of these genes and their relaxed substrate requirements. In particle bombardment tests, An9 and Bz2 functionally complement both mutants. Among several other GSTs tested, only soybean GmGST26A (previously called GmHsp26A and GH2/4) and maize GSTIII were found to confer vacuolar sequestration of anthocyanin. Previously, these genes had not been associated with the anthocyanin pathway. Requirements for An9 and Bz2 gene function were investigated by sequencing functional and nonfunctional germinal revertants of an9-T3529, bz2::Ds, and bz2::Mu1. PMID:9668133

Establishment of chronic hepatitis C virus infection: Translational evasion of oxidative defence

PubMed Central

Chan, Shiu-Wan

2014-01-01

Hepatitis C virus (HCV) causes a clinically important disease affecting 3% of the world population. HCV is a single-stranded, positive-sense RNA virus belonging to the genus Hepacivirus within the Flaviviridae family. The virus establishes a chronic infection in the face of an active host oxidative defence, thus adaptation to oxidative stress is key to virus survival. Being a small RNA virus with a limited genomic capacity, we speculate that HCV deploys a different strategy to evade host oxidative defence. Instead of counteracting oxidative stress, it utilizes oxidative stress to facilitate its own survival. Translation is the first step in the replication of a plus strand RNA virus so it would make sense if the virus can exploit the host oxidative defence in facilitating this very first step. This is particularly true when HCV utilizes an internal ribosome entry site element in translation, which is distinctive from that of cap-dependent translation of the vast majority of cellular genes, thus allowing selective translation of genes under conditions when global protein synthesis is compromised. Indeed, we were the first to show that HCV translation was stimulated by an important pro-oxidant-hydrogen peroxide in hepatocytes, suggesting that HCV is able to adapt to and utilize the host anti-viral response to facilitate its own translation thus allowing the virus to thrive under oxidative stress condition to establish chronicity. Understanding how HCV translation is regulated under oxidative stress condition will advance our knowledge on how HCV establishes chronicity. As chronicity is the initiator step in disease progression this will eventually lead to a better understanding of pathogenicity, which is particularly relevant to the development of anti-virals and improved treatments of HCV patients using anti-oxidants. PMID:24659872

a Migration Well Model for the Binding of Ligands to Heme Proteins.

NASA Astrophysics Data System (ADS)

Beece, Daniel Kenneth

The binding of carbon monoxide and dioxygen to heme proteins can be viewed as occurring in distinct stages: diffusion in the solvent, migration through the matrix, and occupation of the pocket before the final binding step. A model is presented which can explain the dominant kinetic behavior of several different heme protein-ligand systems. The model assumes that a ligand molecule in the solvent sequentially encounters discrete energy barriers on the way to the binding site. The rate to surmount each barrier is distributed, except for the pseudofirst order rate corresponding to the step into the protein from the solvent. The migration through the matrix is equivalent to a small number of distinct jumps. Quantitative analysis of the data permit estimates of the barrier heights, preexponentials and solvent coupling factors for each rate. A migration coefficient and a matrix occupation factor are defined.

Characterization of Two Late-Stage Enzymes Involved in Fosfomycin Biosynthesis in Pseudomonads.

PubMed

Olivares, Philip; Ulrich, Emily C; Chekan, Jonathan R; van der Donk, Wilfred A; Nair, Satish K

2017-02-17

The broad-spectrum phosphonate antibiotic fosfomycin is currently in use for clinical treatment of infections caused by both Gram-positive and Gram-negative uropathogens. The antibiotic is biosynthesized by various streptomycetes, as well as by pseudomonads. Notably, the biosynthetic strategies used by the two genera share only two steps: the first step in which primary metabolite phosphoenolpyruvate (PEP) is converted to phosphonopyruvate (PnPy) and the terminal step in which 2-hydroxypropylphosphonate (2-HPP) is converted to fosfomycin. Otherwise, distinct enzymatic paths are employed. Here, we biochemically confirm the last two steps in the fosfomycin biosynthetic pathway of Pseudomonas syringae PB-5123, showing that Psf3 performs the reduction of 2-oxopropylphosphonate (2-OPP) to (S)-2-HPP, followed by the Psf4-catalyzed epoxidation of (S)-2-HPP to fosfomycin. Psf4 can also accept (R)-2-HPP as a substrate but instead performs an oxidation to make 2-OPP. We show that the combined activities of Psf3 and Psf4 can be used to convert racemic 2-HPP to fosfomycin in an enantioconvergent process. X-ray structures of each enzyme with bound substrates provide insights into the stereospecificity of each conversion. These studies shed light on the reaction mechanisms of the two terminal enzymes in a distinct pathway employed by pseudomonads for the production of a potent antimicrobial agent.

Multiple intensity distributions from a single optical element

NASA Astrophysics Data System (ADS)

Berens, Michael; Bruneton, Adrien; Bäuerle, Axel; Traub, Martin; Wester, Rolf; Stollenwerk, Jochen; Loosen, Peter

2013-09-01

We report on an extension of the previously published two-step freeform optics tailoring algorithm using a Monge-Kantorovich mass transportation framework. The algorithm's ability to design multiple freeform surfaces allows for the inclusion of multiple distinct light paths and hence the implementation of multiple lighting functions in a single optical element. We demonstrate the procedure in the context of automotive lighting, in which a fog lamp and a daytime running lamp are integrated in a single optical element illuminated by two distinct groups of LEDs.

The recruitment of acetylated and unacetylated tropomyosin to distinct actin polymers permits the discrete regulation of specific myosins in fission yeast

PubMed Central

Coulton, Arthur T.; East, Daniel A.; Galinska-Rakoczy, Agnieszka; Lehman, William; Mulvihill, Daniel P.

2010-01-01

Tropomyosin (Tm) is a conserved dimeric coiled-coil protein, which forms polymers that curl around actin filaments in order to regulate actomyosin function. Acetylation of the Tm N-terminal methionine strengthens end-to-end bonds, which enhances actin binding as well as the ability of Tm to regulate myosin motor activity in both muscle and non-muscle cells. In this study we explore the function of each Tm form within fission yeast cells. Electron microscopy and live cell imaging revealed that acetylated and unacetylated Tm associate with distinct actin structures within the cell, and that each form has a profound effect upon the shape and integrity of the polymeric actin filament. We show that, whereas Tm acetylation is required to regulate the in vivo motility of class II myosins, acetylated Tm had no effect on the motility of class I and V myosins. These findings illustrate a novel Tm-acetylation-state-dependent mechanism for regulating specific actomyosin cytoskeletal interactions. PMID:20807799

No Longer Faces in the Crowd

ERIC Educational Resources Information Center

Myers, Anna

2012-01-01

Being distinctive has not always been critical for universities in the U.K. Until recently, significant economic and political forces--largely public funding and regulation--had pushed higher education institutions as a whole toward homogeneity. What is distinctiveness? For some, it's synonymous with being unique. What makes an organization…

Structure-mechanism-based engineering of chemical regulators targeting distinct pathological factors in Alzheimer's disease

NASA Astrophysics Data System (ADS)

Beck, Michael W.; Derrick, Jeffrey S.; Kerr, Richard A.; Oh, Shin Bi; Cho, Woo Jong; Lee, Shin Jung C.; Ji, Yonghwan; Han, Jiyeon; Tehrani, Zahra Aliakbar; Suh, Nayoung; Kim, Sujeong; Larsen, Scott D.; Kim, Kwang S.; Lee, Joo-Yong; Ruotolo, Brandon T.; Lim, Mi Hee

2016-10-01

The absence of effective therapeutics against Alzheimer's disease (AD) is a result of the limited understanding of its multifaceted aetiology. Because of the lack of chemical tools to identify pathological factors, investigations into AD pathogenesis have also been insubstantial. Here we report chemical regulators that demonstrate distinct specificity towards targets linked to AD pathology, including metals, amyloid-β (Aβ), metal-Aβ, reactive oxygen species, and free organic radicals. We obtained these chemical regulators through a rational structure-mechanism-based design strategy. We performed structural variations of small molecules for fine-tuning their electronic properties, such as ionization potentials and mechanistic pathways for reactivity towards different targets. We established in vitro and/or in vivo efficacies of the regulators for modulating their targets' reactivities, ameliorating toxicity, reducing amyloid pathology, and improving cognitive deficits. Our chemical tools show promise for deciphering AD pathogenesis and discovering effective drugs.

Neuronal activity determines distinct gliotransmitter release from a single astrocyte

PubMed Central

Covelo, Ana

2018-01-01

Accumulating evidence indicates that astrocytes are actively involved in brain function by regulating synaptic activity and plasticity. Different gliotransmitters, such as glutamate, ATP, GABA or D-serine, released form astrocytes have been shown to induce different forms of synaptic regulation. However, whether a single astrocyte may release different gliotransmitters is unknown. Here we show that mouse hippocampal astrocytes activated by endogenous (neuron-released endocannabinoids or GABA) or exogenous (single astrocyte Ca2+ uncaging) stimuli modulate putative single CA3-CA1 hippocampal synapses. The astrocyte-mediated synaptic modulation was biphasic and consisted of an initial glutamate-mediated potentiation followed by a purinergic-mediated depression of neurotransmitter release. The temporal dynamic properties of this biphasic synaptic regulation depended on the firing frequency and duration of the neuronal activity that stimulated astrocytes. Present results indicate that single astrocytes can decode neuronal activity and, in response, release distinct gliotransmitters to differentially regulate neurotransmission at putative single synapses. PMID:29380725

Structure-mechanism-based engineering of chemical regulators targeting distinct pathological factors in Alzheimer's disease.

PubMed

Beck, Michael W; Derrick, Jeffrey S; Kerr, Richard A; Oh, Shin Bi; Cho, Woo Jong; Lee, Shin Jung C; Ji, Yonghwan; Han, Jiyeon; Tehrani, Zahra Aliakbar; Suh, Nayoung; Kim, Sujeong; Larsen, Scott D; Kim, Kwang S; Lee, Joo-Yong; Ruotolo, Brandon T; Lim, Mi Hee

2016-10-13

The absence of effective therapeutics against Alzheimer's disease (AD) is a result of the limited understanding of its multifaceted aetiology. Because of the lack of chemical tools to identify pathological factors, investigations into AD pathogenesis have also been insubstantial. Here we report chemical regulators that demonstrate distinct specificity towards targets linked to AD pathology, including metals, amyloid-β (Aβ), metal-Aβ, reactive oxygen species, and free organic radicals. We obtained these chemical regulators through a rational structure-mechanism-based design strategy. We performed structural variations of small molecules for fine-tuning their electronic properties, such as ionization potentials and mechanistic pathways for reactivity towards different targets. We established in vitro and/or in vivo efficacies of the regulators for modulating their targets' reactivities, ameliorating toxicity, reducing amyloid pathology, and improving cognitive deficits. Our chemical tools show promise for deciphering AD pathogenesis and discovering effective drugs.

Endogenous opioids regulate moment-to-moment neuronal communication and excitability.

PubMed

Winters, Bryony L; Gregoriou, Gabrielle C; Kissiwaa, Sarah A; Wells, Oliver A; Medagoda, Danashi I; Hermes, Sam M; Burford, Neil T; Alt, Andrew; Aicher, Sue A; Bagley, Elena E

2017-03-22

Fear and emotional learning are modulated by endogenous opioids but the cellular basis for this is unknown. The intercalated cells (ITCs) gate amygdala output and thus regulate the fear response. Here we find endogenous opioids are released by synaptic stimulation to act via two distinct mechanisms within the main ITC cluster. Endogenously released opioids inhibit glutamate release through the δ-opioid receptor (DOR), an effect potentiated by a DOR-positive allosteric modulator. Postsynaptically, the opioids activate a potassium conductance through the μ-opioid receptor (MOR), suggesting for the first time that endogenously released opioids directly regulate neuronal excitability. Ultrastructural localization of endogenous ligands support these functional findings. This study demonstrates a new role for endogenously released opioids as neuromodulators engaged by synaptic activity to regulate moment-to-moment neuronal communication and excitability. These distinct actions through MOR and DOR may underlie the opposing effect of these receptor systems on anxiety and fear.

β-Catenin activity in the dermal papilla of the hair follicle regulates pigment-type switching

PubMed Central

Enshell-Seijffers, David; Lindon, Catherine; Wu, Eleanor; Taketo, Makoto M.; Morgan, Bruce A.

2010-01-01

The switch between black and yellow pigment is mediated by the interaction between Melanocortin receptor 1 (Mc1r) and its antagonist Agouti, but the genetic and developmental mechanisms that modify this interaction to obtain different coat color in distinct environments are poorly understood. Here, the role of Wnt/β-catenin signaling in the regulation of pigment-type switching was studied. Loss and gain of function of β-catenin in the dermal papilla (DP) of the hair follicle results in yellow and black animals, respectively. β-Catenin activity in the DP suppresses Agouti expression and activates Corin, a negative regulator of Agouti activity. In addition, β-catenin activity in the DP regulates melanocyte activity by a mechanism that is independent of both Agouti and Corin. The coordinate and inverse regulation of Agouti and Corin renders pelage pigmentation sensitive to changes in β-catenin activity in the DP that do not alter pelage structure. As a result, the signals that specify two biologically distinct quantitative traits are partially uncoupled despite their common regulation by the β-catenin pathway in the same cells. PMID:21098273

29 CFR 1956.62 - Completion of developmental steps and certification. [Reserved

Code of Federal Regulations, 2010 CFR

2010-07-01

... 29 Labor 9 2010-07-01 2010-07-01 false Completion of developmental steps and certification. [Reserved] 1956.62 Section 1956.62 Labor Regulations Relating to Labor (Continued) OCCUPATIONAL SAFETY AND... EMPLOYEE PLANS New Jersey § 1956.62 Completion of developmental steps and certification. [Reserved] ...

Synergistic Potentiation of Cystic Fibrosis Transmembrane Conductance Regulator Gating by Two Chemically Distinct Potentiators, Ivacaftor (VX-770) and 5-Nitro-2-(3-Phenylpropylamino) Benzoate.

PubMed

Lin, Wen-Ying; Sohma, Yoshiro; Hwang, Tzyh-Chang

2016-09-01

Cystic fibrosis (CF) is caused by loss-of-function mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene encoding a phosphorylation-activated but ATP-gated chloride channel. Previous studies suggested that VX-770 [ivacaftor, N-(2,4-di-tert-butyl-5-hydroxyphenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide], a CFTR potentiator now used in clinics, increases the open probability of CFTR by shifting the gating conformational changes to favor the open channel configuration. Recently the chloride channel blocker and CFTR potentiator 5-nitro-2-(3-phenylpropylamino) benzoate (NPPB) has been reported to enhance CFTR activity by a mechanism that exploits the ATP hydrolysis-driven, nonequilibrium gating mechanism unique to CFTR. Surprisingly however, NPPB increased the activity of nonhydrolytic G551D-CFTR, the third most common disease-associated mutation. Here, we further investigated the mechanism of NPPB's effects on CFTR gating by assessing its interaction with well-studied VX-770. Interestingly, once G551D-CFTR was maximally potentiated by VX-770, NPPB further increased its activity. However, quantitative analysis of this drug-drug interaction suggests that this pharmacologic synergism is not due to independent actions of NPPB and VX-770 on CFTR gating; instead, our data support a dependent mechanism involving two distinct binding sites. This latter idea is further supported by the observation that the locked-open time of a hydrolysis-deficient mutant K1250A was shortened by NPPB but prolonged by VX-770. In addition, the effectiveness of NPPB, but not of VX-770, was greatly diminished in a mutant whose second nucleotide-binding domain was completely removed. Interpreting these results under the framework of current understanding of CFTR gating not only reveals insights into the mechanism of action for different CFTR potentiators but also brings us one step forward to a more complete schematic for CFTR gating. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Synergistic Potentiation of Cystic Fibrosis Transmembrane Conductance Regulator Gating by Two Chemically Distinct Potentiators, Ivacaftor (VX-770) and 5-Nitro-2-(3-Phenylpropylamino) Benzoate

PubMed Central

Lin, Wen-Ying; Sohma, Yoshiro

2016-01-01

Cystic fibrosis (CF) is caused by loss-of-function mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene encoding a phosphorylation-activated but ATP-gated chloride channel. Previous studies suggested that VX-770 [ivacaftor, N-(2,4-di-tert-butyl-5-hydroxyphenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide], a CFTR potentiator now used in clinics, increases the open probability of CFTR by shifting the gating conformational changes to favor the open channel configuration. Recently the chloride channel blocker and CFTR potentiator 5-nitro-2-(3-phenylpropylamino) benzoate (NPPB) has been reported to enhance CFTR activity by a mechanism that exploits the ATP hydrolysis-driven, nonequilibrium gating mechanism unique to CFTR. Surprisingly however, NPPB increased the activity of nonhydrolytic G551D-CFTR, the third most common disease-associated mutation. Here, we further investigated the mechanism of NPPB’s effects on CFTR gating by assessing its interaction with well-studied VX-770. Interestingly, once G551D-CFTR was maximally potentiated by VX-770, NPPB further increased its activity. However, quantitative analysis of this drug–drug interaction suggests that this pharmacologic synergism is not due to independent actions of NPPB and VX-770 on CFTR gating; instead, our data support a dependent mechanism involving two distinct binding sites. This latter idea is further supported by the observation that the locked-open time of a hydrolysis-deficient mutant K1250A was shortened by NPPB but prolonged by VX-770. In addition, the effectiveness of NPPB, but not of VX-770, was greatly diminished in a mutant whose second nucleotide-binding domain was completely removed. Interpreting these results under the framework of current understanding of CFTR gating not only reveals insights into the mechanism of action for different CFTR potentiators but also brings us one step forward to a more complete schematic for CFTR gating. PMID:27413118

SRSF3 represses the expression of PDCD4 protein by coordinated regulation of alternative splicing, export and translation

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

Park, Seung Kuk; Jeong, Sunjoo, E-mail: sjsj@dankook.ac.kr

2016-02-05

Gene expression is regulated at multiple steps, such as transcription, splicing, export, degradation and translation. Considering diverse roles of SR proteins, we determined whether the tumor-related splicing factor SRSF3 regulates the expression of the tumor-suppressor protein, PDCD4, at multiple steps. As we have reported previously, knockdown of SRSF3 increased the PDCD4 protein level in SW480 colon cancer cells. More interestingly, here we showed that the alternative splicing and the nuclear export of minor isoforms of pdcd4 mRNA were repressed by SRSF3, but the translation step was unaffected. In contrast, only the translation step of the major isoform of pdcd4 mRNAmore » was repressed by SRSF3. Therefore, overexpression of SRSF3 might be relevant to the repression of all isoforms of PDCD4 protein levels in most types of cancer cell. We propose that SRSF3 could act as a coordinator of the expression of PDCD4 protein via two mechanisms on two alternatively spliced mRNA isoforms.« less

Synthetic studies of the zoanthamine alkaloids: the total syntheses of norzoanthamine and zoanthamine.

PubMed

Yoshimura, Fumihiko; Sasaki, Minoru; Hattori, Izumi; Komatsu, Kei; Sakai, Mio; Tanino, Keiji; Miyashita, Masaaki

2009-07-06

The zoanthamine alkaloids, a type of heptacyclic marine alkaloid isolated from colonial zoanthids of the genus Zoanthus sp., have distinctive biological and pharmacological properties in addition to their unique chemical structures with stereochemical complexity. Namely, norzoanthamine (1) can suppress the loss of bone weight and strength in ovariectomized mice and has been expected as a promising candidate for a new type of antiosteoporotic drug, while zoanthamine (2) has exhibited potent inhibitory activity toward phorbol myristate-induced inflammation in addition to powerful analgesic effects. Recently, norzoanthamine derivatives were demonstrated to inhibit strongly the growth of P-388 murine leukemia cell lines, in addition to their potent antiplatelet activities on human platelet aggregation. Their distinctive biological properties, combined with novel chemical structures, make this family of alkaloids extremely attractive targets for chemical synthesis. However, the chemical synthesis of the zoanthamine alkaloids has been impeded owing to their densely functionalized complex stereostructures. In this paper, we report the first and highly efficient total syntheses of norzoanthamine (1) and zoanthamine (2) in full detail, which involve stereoselective synthesis of the requisite triene (18) for an intramolecular Diels-Alder reaction via the sequential three-component coupling reactions, the key intramolecular Diels-Alder reaction, and subsequent crucial bis-aminoacetalization as the key steps. Ultimately, we achieved the total synthesis of norzoanthamine (1) in 41 steps with an overall yield of 3.5 % (an average of 92 % yield each step) and that of zoanthamine (2) in 43 steps with an overall yield of 2.2 % (an average of 91 % yield each step) starting from (R)-5-methylcyclohexenone (3), respectively.

A novel statistical approach for identification of the master regulator transcription factor.

PubMed

Sikdar, Sinjini; Datta, Susmita

2017-02-02

Transcription factors are known to play key roles in carcinogenesis and therefore, are gaining popularity as potential therapeutic targets in drug development. A 'master regulator' transcription factor often appears to control most of the regulatory activities of the other transcription factors and the associated genes. This 'master regulator' transcription factor is at the top of the hierarchy of the transcriptomic regulation. Therefore, it is important to identify and target the master regulator transcription factor for proper understanding of the associated disease process and identifying the best therapeutic option. We present a novel two-step computational approach for identification of master regulator transcription factor in a genome. At the first step of our method we test whether there exists any master regulator transcription factor in the system. We evaluate the concordance of two ranked lists of transcription factors using a statistical measure. In case the concordance measure is statistically significant, we conclude that there is a master regulator. At the second step, our method identifies the master regulator transcription factor, if there exists one. In the simulation scenario, our method performs reasonably well in validating the existence of a master regulator when the number of subjects in each treatment group is reasonably large. In application to two real datasets, our method ensures the existence of master regulators and identifies biologically meaningful master regulators. An R code for implementing our method in a sample test data can be found in http://www.somnathdatta.org/software . We have developed a screening method of identifying the 'master regulator' transcription factor just using only the gene expression data. Understanding the regulatory structure and finding the master regulator help narrowing the search space for identifying biomarkers for complex diseases such as cancer. In addition to identifying the master regulator our method provides an overview of the regulatory structure of the transcription factors which control the global gene expression profiles and consequently the cell functioning.

Two Distinct Types of E3 Ligases Work in Unison to Regulate Substrate Ubiquitylation.

PubMed

Scott, Daniel C; Rhee, David Y; Duda, David M; Kelsall, Ian R; Olszewski, Jennifer L; Paulo, Joao A; de Jong, Annemieke; Ovaa, Huib; Alpi, Arno F; Harper, J Wade; Schulman, Brenda A

2016-08-25

Hundreds of human cullin-RING E3 ligases (CRLs) modify thousands of proteins with ubiquitin (UB) to achieve vast regulation. Current dogma posits that CRLs first catalyze UB transfer from an E2 to their client substrates and subsequent polyubiquitylation from various linkage-specific E2s. We report an alternative E3-E3 tagging cascade: many cellular NEDD8-modified CRLs associate with a mechanistically distinct thioester-forming RBR-type E3, ARIH1, and rely on ARIH1 to directly add the first UB and, in some cases, multiple additional individual monoubiquitin modifications onto CRL client substrates. Our data define ARIH1 as a component of the human CRL system, demonstrate that ARIH1 can efficiently and specifically mediate monoubiquitylation of several CRL substrates, and establish principles for how two distinctive E3s can reciprocally control each other for simultaneous and joint regulation of substrate ubiquitylation. These studies have broad implications for CRL-dependent proteostasis and mechanisms of E3-mediated UB ligation. Copyright © 2016 Elsevier Inc. All rights reserved.

Distinct Contributions of Conserved Modules to Runt Transcription Factor Activity

PubMed Central

Walrad, Pegine B.; Hang, Saiyu; Joseph, Genevieve S.; Salas, Julia

2010-01-01

Runx proteins play vital roles in regulating transcription in numerous developmental pathways throughout the animal kingdom. Two Runx protein hallmarks are the DNA-binding Runt domain and a C-terminal VWRPY motif that mediates interaction with TLE/Gro corepressor proteins. A phylogenetic analysis of Runt, the founding Runx family member, identifies four distinct regions C-terminal to the Runt domain that are conserved in Drosophila and other insects. We used a series of previously described ectopic expression assays to investigate the functions of these different conserved regions in regulating gene expression during embryogenesis and in controlling axonal projections in the developing eye. The results indicate each conserved region is required for a different subset of activities and identify distinct regions that participate in the transcriptional activation and repression of the segmentation gene sloppy-paired-1 (slp1). Interestingly, the C-terminal VWRPY-containing region is not required for repression but instead plays a role in slp1 activation. Genetic experiments indicating that Groucho (Gro) does not participate in slp1 regulation further suggest that Runt's conserved C-terminus interacts with other factors to promote transcriptional activation. These results provide a foundation for further studies on the molecular interactions that contribute to the context-dependent properties of Runx proteins as developmental regulators. PMID:20462957

Autogenous cross-regulation of Quaking mRNA processing and translation balances Quaking functions in splicing and translation

PubMed Central

Liu, Naiyou; Fair, Jeffrey Haskell; Shiue, Lily; Katzman, Sol; Donohue, John Paul

2017-01-01

Quaking protein isoforms arise from a single Quaking gene and bind the same RNA motif to regulate splicing, translation, decay, and localization of a large set of RNAs. However, the mechanisms by which Quaking expression is controlled to ensure that appropriate amounts of each isoform are available for such disparate gene expression processes are unknown. Here we explore how levels of two isoforms, nuclear Quaking-5 (Qk5) and cytoplasmic Qk6, are regulated in mouse myoblasts. We found that Qk5 and Qk6 proteins have distinct functions in splicing and translation, respectively, enforced through differential subcellular localization. We show that Qk5 and Qk6 regulate distinct target mRNAs in the cell and act in distinct ways on their own and each other's transcripts to create a network of autoregulatory and cross-regulatory feedback controls. Morpholino-mediated inhibition of Qk translation confirms that Qk5 controls Qk RNA levels by promoting accumulation and alternative splicing of Qk RNA, whereas Qk6 promotes its own translation while repressing Qk5. This Qk isoform cross-regulatory network responds to additional cell type and developmental controls to generate a spectrum of Qk5/Qk6 ratios, where they likely contribute to the wide range of functions of Quaking in development and cancer. PMID:29021242

PSD-95 stabilizes NMDA receptors by inducing the degradation of STEP61.

PubMed

Won, Sehoon; Incontro, Salvatore; Nicoll, Roger A; Roche, Katherine W

2016-08-09

Phosphorylation regulates surface and synaptic expression of NMDA receptors (NMDARs). Both the tyrosine kinase Fyn and the tyrosine phosphatase striatal-enriched protein tyrosine phosphatase (STEP) are known to target the NMDA receptor subunit GluN2B on tyrosine 1472, which is a critical residue that mediates NMDAR endocytosis. STEP reduces the surface expression of NMDARs by promoting dephosphorylation of GluN2B Y1472, whereas the synaptic scaffolding protein postsynaptic density protein 95 (PSD-95) stabilizes the surface expression of NMDARs. However, nothing is known about a potential functional interaction between STEP and PSD-95. We now report that STEP61 binds to PSD-95 but not to other PSD-95 family members. We find that PSD-95 expression destabilizes STEP61 via ubiquitination and degradation by the proteasome. Using subcellular fractionation, we detect low amounts of STEP61 in the PSD fraction. However, STEP61 expression in the PSD is increased upon knockdown of PSD-95 or in vivo as detected in PSD-95-KO mice, demonstrating that PSD-95 excludes STEP61 from the PSD. Importantly, only extrasynaptic NMDAR expression and currents were increased upon STEP knockdown, as is consistent with low STEP61 localization in the PSD. Our findings support a dual role for PSD-95 in stabilizing synaptic NMDARs by binding directly to GluN2B but also by promoting synaptic exclusion and degradation of the negative regulator STEP61.

PSD-95 stabilizes NMDA receptors by inducing the degradation of STEP61

PubMed Central

Won, Sehoon; Incontro, Salvatore; Nicoll, Roger A.; Roche, Katherine W.

2016-01-01

Phosphorylation regulates surface and synaptic expression of NMDA receptors (NMDARs). Both the tyrosine kinase Fyn and the tyrosine phosphatase striatal-enriched protein tyrosine phosphatase (STEP) are known to target the NMDA receptor subunit GluN2B on tyrosine 1472, which is a critical residue that mediates NMDAR endocytosis. STEP reduces the surface expression of NMDARs by promoting dephosphorylation of GluN2B Y1472, whereas the synaptic scaffolding protein postsynaptic density protein 95 (PSD-95) stabilizes the surface expression of NMDARs. However, nothing is known about a potential functional interaction between STEP and PSD-95. We now report that STEP61 binds to PSD-95 but not to other PSD-95 family members. We find that PSD-95 expression destabilizes STEP61 via ubiquitination and degradation by the proteasome. Using subcellular fractionation, we detect low amounts of STEP61 in the PSD fraction. However, STEP61 expression in the PSD is increased upon knockdown of PSD-95 or in vivo as detected in PSD-95–KO mice, demonstrating that PSD-95 excludes STEP61 from the PSD. Importantly, only extrasynaptic NMDAR expression and currents were increased upon STEP knockdown, as is consistent with low STEP61 localization in the PSD. Our findings support a dual role for PSD-95 in stabilizing synaptic NMDARs by binding directly to GluN2B but also by promoting synaptic exclusion and degradation of the negative regulator STEP61. PMID:27457929

Use of a Phosphorylation Site Mutant To Identify Distinct Modes of Gene Repression by the Control of Virulence Regulator (CovR) in Streptococcus pyogenes.

PubMed

Horstmann, Nicola; Sahasrabhojane, Pranoti; Yao, Hui; Su, Xiaoping; Shelburne, Samuel A

2017-09-15

Control of the virulence regulator/sensor kinase (CovRS) two-component system (TCS) serves as a model for investigating the impact of signaling pathways on the pathogenesis of Gram-positive bacteria. However, the molecular mechanisms by which CovR, an OmpR/PhoB family response regulator, controls virulence gene expression are poorly defined, partly due to the labile nature of its aspartate phosphorylation site. To better understand the regulatory effect of phosphorylated CovR, we generated the phosphorylation site mutant strain 10870-CovR-D53E, which we predicted to have a constitutive CovR phosphorylation phenotype. Interestingly, this strain showed CovR activity only for a subset of the CovR regulon, which allowed for classification of CovR-influenced genes into D53E-regulated and D53E-nonregulated groups. Inspection of the promoter sequences of genes belonging to each group revealed distinct promoter architectures with respect to the location and number of putative CovR-binding sites. Electrophoretic mobility shift analysis demonstrated that recombinant CovR-D53E protein retains its ability to bind promoter DNA from both CovR-D53E-regulated and -nonregulated groups, implying that factors other than mere DNA binding are crucial for gene regulation. In fact, we found that CovR-D53E is incapable of dimerization, a process thought to be critical to OmpR/PhoB family regulator function. Thus, our global analysis of CovR-D53E indicates dimerization-dependent and dimerization-independent modes of CovR-mediated repression, thereby establishing distinct mechanisms by which this critical regulator coordinates virulence gene expression. IMPORTANCE Streptococcus pyogenes causes a wide variety of diseases, ranging from superficial skin and throat infections to life-threatening invasive infections. To establish these various disease manifestations, Streptococcus pyogenes requires tightly coordinated production of its virulence factor repertoire. Here, the response regulator CovR plays a crucial role. As an OmpR/PhoB family member, CovR is activated by phosphorylation on a conserved aspartate residue, leading to protein dimerization and subsequent binding to operator sites. Our transcriptome analysis using the monomeric phosphorylation mimic mutant CovR-D53E broadens this general notion by revealing dimerization-independent repression of a subset of CovR-regulated genes. Combined with promoter analyses, these data suggest distinct mechanisms of CovR transcriptional control, which allow for differential expression of virulence genes in response to environmental cues. Copyright © 2017 American Society for Microbiology.

MicroRNA networks in mouse lung organogenesis.

PubMed

Dong, Jie; Jiang, Guoqian; Asmann, Yan W; Tomaszek, Sandra; Jen, Jin; Kislinger, Thomas; Wigle, Dennis A

2010-05-26

MicroRNAs (miRNAs) are known to be important regulators of both organ development and tumorigenesis. MiRNA networks and their regulation of messenger RNA (mRNA) translation and protein expression in specific biological processes are poorly understood. We explored the dynamic regulation of miRNAs in mouse lung organogenesis. Comprehensive miRNA and mRNA profiling was performed encompassing all recognized stages of lung development beginning at embryonic day 12 and continuing to adulthood. We analyzed the expression patterns of dynamically regulated miRNAs and mRNAs using a number of statistical and computational approaches, and in an integrated manner with protein levels from an existing mass-spectrometry derived protein database for lung development. In total, 117 statistically significant miRNAs were dynamically regulated during mouse lung organogenesis and clustered into distinct temporal expression patterns. 11,220 mRNA probes were also shown to be dynamically regulated and clustered into distinct temporal expression patterns, with 3 major patterns accounting for 75% of all probes. 3,067 direct miRNA-mRNA correlation pairs were identified involving 37 miRNAs. Two defined correlation patterns were observed upon integration with protein data: 1) increased levels of specific miRNAs directly correlating with downregulation of predicted mRNA targets; and 2) increased levels of specific miRNAs directly correlating with downregulation of translated target proteins without detectable changes in mRNA levels. Of 1345 proteins analyzed, 55% appeared to be regulated in this manner with a direct correlation between miRNA and protein level, but without detectable change in mRNA levels. Systematic analysis of microRNA, mRNA, and protein levels over the time course of lung organogenesis demonstrates dynamic regulation and reveals 2 distinct patterns of miRNA-mRNA interaction. The translation of target proteins affected by miRNAs independent of changes in mRNA level appears to be a prominent mechanism of developmental regulation in lung organogenesis.

Immersive visualization of rail simulation data.

DOT National Transportation Integrated Search

2016-01-01

The prime objective of this project was to create scientific, immersive visualizations of a Rail-simulation. This project is a part of a larger initiative that consists of three distinct parts. The first step consists of performing a finite element a...

Interdisciplinary cognitive task analysis: a strategy to develop a comprehensive endoscopic retrograde cholangiopancreatography protocol for use in fellowship training.

PubMed

Canopy, Erin; Evans, Matt; Boehler, Margaret; Roberts, Nicole; Sanfey, Hilary; Mellinger, John

2015-10-01

Endoscopic retrograde cholangiopancreatography is a challenging procedure performed by surgeons and gastroenterologists. We employed cognitive task analysis to identify steps and decision points for this procedure. Standardized interviews were conducted with expert gastroenterologists (7) and surgeons (4) from 4 institutions. A procedural step and cognitive decision point protocol was created from audio-taped transcriptions and was refined by 5 additional surgeons. Conceptual elements, sequential actions, and decision points were iterated for 5 tasks: patient preparation, duodenal intubation, selective cannulation, imaging interpretation with related therapeutic intervention, and complication management. A total of 180 steps were identified. Gastroenterologists identified 34 steps not identified by surgeons, and surgeons identified 20 steps not identified by gastroenterologists. The findings suggest that for complex procedures performed by diverse practitioners, more experts may help delineate distinctive emphases differentiated by training background and type of practice. Copyright © 2015 Elsevier Inc. All rights reserved.

Investigation to biodiesel production by the two-step homogeneous base-catalyzed transesterification.

PubMed

Ye, Jianchu; Tu, Song; Sha, Yong

2010-10-01

For the two-step transesterification biodiesel production made from the sunflower oil, based on the kinetics model of the homogeneous base-catalyzed transesterification and the liquid-liquid phase equilibrium of the transesterification product, the total methanol/oil mole ratio, the total reaction time, and the split ratios of methanol and reaction time between the two reactors in the stage of the two-step reaction are determined quantitatively. In consideration of the transesterification intermediate product, both the traditional distillation separation process and the improved separation process of the two-step reaction product are investigated in detail by means of the rigorous process simulation. In comparison with the traditional distillation process, the improved separation process of the two-step reaction product has distinct advantage in the energy duty and equipment requirement due to replacement of the costly methanol-biodiesel distillation column. Copyright 2010 Elsevier Ltd. All rights reserved.

A Novel Mechanism of Regulating the ATPase VPS4 by Its Cofactor LIP5 and the Endosomal Sorting Complex Required for Transport (ESCRT)-III Protein CHMP5

DOE PAGES

Vild, Cody J.; Li, Yan; Guo, Emily Z.; ...

2015-01-30

Disassembly of the endosomal sorting complex required for transport (ESCRT) machinery from biological membranes is a critical final step in cellular processes that require the ESCRT function. This reaction is catalyzed by VPS4, an AAA-ATPase whose activity is tightly regulated by a host of proteins, including LIP5 and the ESCRT-III proteins. In this paper, we present structural and functional analyses of molecular interactions between human VPS4, LIP5, and the ESCRT-III proteins. The N-terminal domain of LIP5 (LIP5NTD) is required for LIP5-mediated stimulation of VPS4, and the ESCRT-III protein CHMP5 strongly inhibits the stimulation. Both of these observations are distinct frommore » what was previously described for homologous yeast proteins. The crystal structure of LIP5NTD in complex with the MIT (microtubule-interacting and transport)-interacting motifs of CHMP5 and a second ESCRT-III protein, CHMP1B, was determined at 1 Å resolution. It reveals an ESCRT-III binding induced moderate conformational change in LIP5NTD, which results from insertion of a conserved CHMP5 tyrosine residue (Tyr 182) at the core of LIP5NTD structure. Finally, mutation of Tyr 182 partially relieves the inhibition displayed by CHMP5. Together, these results suggest a novel mechanism of VPS4 regulation in metazoans, where CHMP5 functions as a negative allosteric switch to control LIP5-mediated stimulation of VPS4.« less

Diverse RNA-binding proteins interact with functionally related sets of RNAs, suggesting an extensive regulatory system.

PubMed

Hogan, Daniel J; Riordan, Daniel P; Gerber, André P; Herschlag, Daniel; Brown, Patrick O

2008-10-28

RNA-binding proteins (RBPs) have roles in the regulation of many post-transcriptional steps in gene expression, but relatively few RBPs have been systematically studied. We searched for the RNA targets of 40 proteins in the yeast Saccharomyces cerevisiae: a selective sample of the approximately 600 annotated and predicted RBPs, as well as several proteins not annotated as RBPs. At least 33 of these 40 proteins, including three of the four proteins that were not previously known or predicted to be RBPs, were reproducibly associated with specific sets of a few to several hundred RNAs. Remarkably, many of the RBPs we studied bound mRNAs whose protein products share identifiable functional or cytotopic features. We identified specific sequences or predicted structures significantly enriched in target mRNAs of 16 RBPs. These potential RNA-recognition elements were diverse in sequence, structure, and location: some were found predominantly in 3'-untranslated regions, others in 5'-untranslated regions, some in coding sequences, and many in two or more of these features. Although this study only examined a small fraction of the universe of yeast RBPs, 70% of the mRNA transcriptome had significant associations with at least one of these RBPs, and on average, each distinct yeast mRNA interacted with three of the RBPs, suggesting the potential for a rich, multidimensional network of regulation. These results strongly suggest that combinatorial binding of RBPs to specific recognition elements in mRNAs is a pervasive mechanism for multi-dimensional regulation of their post-transcriptional fate.

Structural Biology and Evolution of the TGF-β Family

PubMed Central

Hinck, Andrew P.; Mueller, Thomas D.; Springer, Timothy A.

2017-01-01

We review the evolution and structure of members of the transforming growth factor β (TGF-β) family, antagonistic or agonistic modulators, and receptors that regulate TGF-β signaling in extracellular environments. The growth factor (GF) domain common to all family members and many of their antagonists evolved from a common cystine knot growth factor (CKGF) domain. The CKGF superfamily comprises six distinct families in primitive metazoans, including the TGF-β and Dan families. Compared with Wnt/Frizzled and Notch/Delta families that also specify body axes, cell fate, tissues, and other families that contain CKGF domains that evolved in parallel, the TGF-β family was the most fruitful in evolution. Complexes between the prodomains and GFs of the TGF-β family suggest a new paradigm for regulating GF release by conversion from closed- to open-arm procomplex conformations. Ternary complexes of the final step in extracellular signaling show how TGF-β GF dimers bind type I and type II receptors on the cell surface, and enable understanding of much of the specificity and promiscuity in extracellular signaling. However, structures suggest that when GFs bind repulsive guidance molecule (RGM) family coreceptors, type I receptors do not bind until reaching an intracellular, membrane-enveloped compartment, blurring the line between extra- and intracellular signaling. Modulator protein structures show how structurally diverse antagonists including follistatins, noggin, and members of the chordin family bind GFs to regulate signaling; complexes with the Dan family remain elusive. Much work is needed to understand how these molecular components assemble to form signaling hubs in extracellular environments in vivo. PMID:27638177

Analysis of human soft palate morphogenesis supports regional regulation of palatal fusion

PubMed Central

Danescu, Adrian; Mattson, Melanie; Dool, Carly; Diewert, Virginia M; Richman, Joy M

2015-01-01

It is essential to complete palate closure at the correct time during fetal development, otherwise a serious malformation, cleft palate, will ensue. The steps in palate formation in humans take place between the 7th and 12th week and consist of outgrowth of palatal shelves from the paired maxillary prominences, reorientation of the shelves from vertical to horizontal, apposition of the medial surfaces, formation of a bilayered seam, degradation of the seam and bridging of mesenchyme. However, in the soft palate, the mechanism of closure is unclear. In previous studies it is possible to find support for both fusion and the alternative mechanism of merging. Here we densely sample the late embryonic-early fetal period between 54 and 74 days post-conception to determine the timing and mechanism of soft palate closure. We found the epithelial seam extends throughout the soft palates of 57-day specimens. Cytokeratin antibody staining detected the medial edge epithelium and distinguished clearly that cells in the midline retained their epithelial character. Compared with the hard palate, the epithelium is more rapidly degraded in the soft palate and only persists in the most posterior regions at 64 days. Our results are consistent with the soft palate following a developmentally more rapid program of fusion than the hard palate. Importantly, the two regions of the palate appear to be independently regulated and have their own internal clocks regulating the timing of seam removal. Considering data from human genetic and mouse studies, distinct anterior-posterior signaling mechanisms are likely to be at play in the human fetal palate. PMID:26299693

Diverse patterns of genomic targeting by transcriptional regulators in Drosophila melanogaster.

PubMed

Slattery, Matthew; Ma, Lijia; Spokony, Rebecca F; Arthur, Robert K; Kheradpour, Pouya; Kundaje, Anshul; Nègre, Nicolas; Crofts, Alex; Ptashkin, Ryan; Zieba, Jennifer; Ostapenko, Alexander; Suchy, Sarah; Victorsen, Alec; Jameel, Nader; Grundstad, A Jason; Gao, Wenxuan; Moran, Jennifer R; Rehm, E Jay; Grossman, Robert L; Kellis, Manolis; White, Kevin P

2014-07-01

Annotation of regulatory elements and identification of the transcription-related factors (TRFs) targeting these elements are key steps in understanding how cells interpret their genetic blueprint and their environment during development, and how that process goes awry in the case of disease. One goal of the modENCODE (model organism ENCyclopedia of DNA Elements) Project is to survey a diverse sampling of TRFs, both DNA-binding and non-DNA-binding factors, to provide a framework for the subsequent study of the mechanisms by which transcriptional regulators target the genome. Here we provide an updated map of the Drosophila melanogaster regulatory genome based on the location of 84 TRFs at various stages of development. This regulatory map reveals a variety of genomic targeting patterns, including factors with strong preferences toward proximal promoter binding, factors that target intergenic and intronic DNA, and factors with distinct chromatin state preferences. The data also highlight the stringency of the Polycomb regulatory network, and show association of the Trithorax-like (Trl) protein with hotspots of DNA binding throughout development. Furthermore, the data identify more than 5800 instances in which TRFs target DNA regions with demonstrated enhancer activity. Regions of high TRF co-occupancy are more likely to be associated with open enhancers used across cell types, while lower TRF occupancy regions are associated with complex enhancers that are also regulated at the epigenetic level. Together these data serve as a resource for the research community in the continued effort to dissect transcriptional regulatory mechanisms directing Drosophila development. © 2014 Slattery et al.; Published by Cold Spring Harbor Laboratory Press.

Genome-Wide Analysis Reveals the Unique Stem Cell Identity of Human Amniocytes

PubMed Central

Maguire, Colin T.; Demarest, Bradley L.; Hill, Jonathon T.; Palmer, James D.; Brothman, Arthur R.; Yost, H. Joseph; Condic, Maureen L.

2013-01-01

Human amniotic fluid contains cells that potentially have important stem cell characteristics, yet the programs controlling their developmental potency are unclear. Here, we provide evidence that amniocytes derived from multiple patients are marked by heterogeneity and variability in expression levels of pluripotency markers. Clonal analysis from multiple patients indicates that amniocytes have large pools of self-renewing cells that have an inherent property to give rise to a distinct amniocyte phenotype with a heterogeneity of pluripotent markers. Significant to their therapeutic potential, genome-wide profiles are distinct at different gestational ages and times in culture, but do not differ between genders. Based on hierarchical clustering and differential expression analyses of the entire transcriptome, amniocytes express canonical regulators associated with pluripotency and stem cell repression. Their profiles are distinct from human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and newborn foreskin fibroblasts. Amniocytes have a complex molecular signature, coexpressing trophoblastic, ectodermal, mesodermal, and endodermal cell-type-specific regulators. In contrast to the current view of the ground state of stem cells, ESCs and iPSCs also express high levels of a wide range of cell-type-specific regulators. The coexpression of multilineage differentiation markers combined with the strong expression of a subset of ES cell repressors in amniocytes suggests that these cells have a distinct phenotype that is unlike any other known cell-type or lineage. PMID:23326421

Phenotype- and Genotype-Specific Structural Alterations in Spasmodic Dysphonia

PubMed Central

Bianchi, Serena; Battistella, Giovanni; Huddleston, Hailey; Scharf, Rebecca; Fleysher, Lazar; Rumbach, Anna F.; Frucht, Steven J.; Blitzer, Andrew; Ozelius, Laurie J.; Simonyan, Kristina

2017-01-01

Background Spasmodic dysphonia is a focal dystonia characterized by involuntary spasms in the laryngeal muscles that occur selectively during speaking. Although hereditary trends have been reported in up to 16% of patients, the causative etiology of spasmodic dysphonia is unclear, and the influences of various phenotypes and genotypes on disorder pathophysiology are poorly understood. In this study, we examined structural alterations in cortical gray matter and white matter integrity in relationship to different phenotypes and putative genotypes of spasmodic dysphonia to elucidate the structural component of its complex pathophysiology. Methods Eighty-nine patients with spasmodic dysphonia underwent high-resolution magnetic resonance imaging and diffusion-weighted imaging to examine cortical thickness and white matter fractional anisotropy in adductor versus abductor forms (distinct phenotypes) and in sporadic versus familial cases (distinct genotypes). Results Phenotype-specific abnormalities were localized in the left sensorimotor cortex and angular gyrus and the white matter bundle of the right superior corona radiata. Genotype-specific alterations were found in the left superior temporal gyrus, supplementary motor area, and the arcuate portion of the left superior longitudinal fasciculus. Conclusions Our findings suggest that phenotypic differences in spasmodic dysphonia arise at the level of the primary and associative areas of motor control, whereas genotype-related pathophysiological mechanisms may be associated with dysfunction of regions regulating phonological and sensory processing. Identification of structural alterations specific to disorder phenotype and putative genotype provides an important step toward future delineation of imaging markers and potential targets for novel therapeutic interventions for spasmodic dysphonia. PMID:28186656

The Last Step of Syringyl Monolignol Biosynthesis in Angiosperms Is Regulated by a Novel Gene Encoding Sinapyl Alcohol Dehydrogenase

PubMed Central

Li, Laigeng; Cheng, Xiao Fei; Leshkevich, Jacqueline; Umezawa, Toshiaki; Harding, Scott A.; Chiang, Vincent L.

2001-01-01

Cinnamyl alcohol dehydrogenase (CAD; EC 1.1.1.195) has been thought to mediate the reduction of both coniferaldehyde and sinapaldehyde into guaiacyl and syringyl monolignols in angiosperms. Here, we report the isolation of a novel aspen gene (PtSAD) encoding sinapyl alcohol dehydrogenase (SAD), which is phylogenetically distinct from aspen CAD (PtCAD). Liquid chromatography–mass spectrometry-based enzyme functional analysis and substrate level–controlled enzyme kinetics consistently demonstrated that PtSAD is sinapaldehyde specific and that PtCAD is coniferaldehyde specific. The enzymatic efficiency of PtSAD for sinapaldehyde was ∼60 times greater than that of PtCAD. These data suggest that in addition to CAD, discrete SAD function is essential to the biosynthesis of syringyl monolignol in angiosperms. In aspen stem primary tissues, PtCAD was immunolocalized exclusively to xylem elements in which only guaiacyl lignin was deposited, whereas PtSAD was abundant in syringyl lignin–enriched phloem fiber cells. In the developing secondary stem xylem, PtCAD was most conspicuous in guaiacyl lignin–enriched vessels, but PtSAD was nearly absent from these elements and was conspicuous in fiber cells. In the context of additional protein immunolocalization and lignin histochemistry, these results suggest that the distinct CAD and SAD functions are linked spatiotemporally to the differential biosynthesis of guaiacyl and syringyl lignins in different cell types. SAD is required for the biosynthesis of syringyl lignin in angiosperms. PMID:11449052

The last step of syringyl monolignol biosynthesis in angiosperms is regulated by a novel gene encoding sinapyl alcohol dehydrogenase.

PubMed

Li, L; Cheng, X F; Leshkevich, J; Umezawa, T; Harding, S A; Chiang, V L

2001-07-01

Cinnamyl alcohol dehydrogenase (CAD; EC 1.1.1.195) has been thought to mediate the reduction of both coniferaldehyde and sinapaldehyde into guaiacyl and syringyl monolignols in angiosperms. Here, we report the isolation of a novel aspen gene (PtSAD) encoding sinapyl alcohol dehydrogenase (SAD), which is phylogenetically distinct from aspen CAD (PtCAD). Liquid chromatography-mass spectrometry-based enzyme functional analysis and substrate level-controlled enzyme kinetics consistently demonstrated that PtSAD is sinapaldehyde specific and that PtCAD is coniferaldehyde specific. The enzymatic efficiency of PtSAD for sinapaldehyde was approximately 60 times greater than that of PtCAD. These data suggest that in addition to CAD, discrete SAD function is essential to the biosynthesis of syringyl monolignol in angiosperms. In aspen stem primary tissues, PtCAD was immunolocalized exclusively to xylem elements in which only guaiacyl lignin was deposited, whereas PtSAD was abundant in syringyl lignin-enriched phloem fiber cells. In the developing secondary stem xylem, PtCAD was most conspicuous in guaiacyl lignin-enriched vessels, but PtSAD was nearly absent from these elements and was conspicuous in fiber cells. In the context of additional protein immunolocalization and lignin histochemistry, these results suggest that the distinct CAD and SAD functions are linked spatiotemporally to the differential biosynthesis of guaiacyl and syringyl lignins in different cell types. SAD is required for the biosynthesis of syringyl lignin in angiosperms.

5 CFR 534.203 - Maximum stipends.

Code of Federal Regulations, 2011 CFR

2011-01-01

... training program Maximums by grade and step 1 L-A Below high school graduation GS-1-1 (minus 3 steps). L-1... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS PAY UNDER OTHER SYSTEMS Student... year postgraduate predoctoral GS-7-1 (minus 3 steps). L-6 Third year medical school GS-7-1 (minus 3...

5 CFR 534.203 - Maximum stipends.

Code of Federal Regulations, 2012 CFR

2012-01-01

... training program Maximums by grade and step 1 L-A Below high school graduation GS-1-1 (minus 3 steps). L-1... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS PAY UNDER OTHER SYSTEMS Student... year postgraduate predoctoral GS-7-1 (minus 3 steps). L-6 Third year medical school GS-7-1 (minus 3...

5 CFR 534.203 - Maximum stipends.

Code of Federal Regulations, 2014 CFR

2014-01-01

... training program Maximums by grade and step 1 L-A Below high school graduation GS-1-1 (minus 3 steps). L-1... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS PAY UNDER OTHER SYSTEMS Student... year postgraduate predoctoral GS-7-1 (minus 3 steps). L-6 Third year medical school GS-7-1 (minus 3...

5 CFR 534.203 - Maximum stipends.

Code of Federal Regulations, 2013 CFR

2013-01-01

... training program Maximums by grade and step 1 L-A Below high school graduation GS-1-1 (minus 3 steps). L-1... Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT CIVIL SERVICE REGULATIONS PAY UNDER OTHER SYSTEMS Student... year postgraduate predoctoral GS-7-1 (minus 3 steps). L-6 Third year medical school GS-7-1 (minus 3...

45 CFR 2530.85 - What steps are necessary to revoke a transfer?

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 4 2010-10-01 2010-10-01 false What steps are necessary to revoke a transfer? 2530.85 Section 2530.85 Public Welfare Regulations Relating to Public Welfare (Continued) CORPORATION FOR NATIONAL AND COMMUNITY SERVICE TRANSFER OF EDUCATION AWARDS § 2530.85 What steps are necessary to...

45 CFR 2530.30 - What steps are necessary to transfer an education award?

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 4 2010-10-01 2010-10-01 false What steps are necessary to transfer an education award? 2530.30 Section 2530.30 Public Welfare Regulations Relating to Public Welfare (Continued) CORPORATION FOR NATIONAL AND COMMUNITY SERVICE TRANSFER OF EDUCATION AWARDS § 2530.30 What steps are necessary...

Molecular mechanism of ERK dephosphorylation by striatal-enriched protein tyrosine phosphatase (STEP)

PubMed Central

Li, Hui; Li, Kang-shuai; Su, Jing; Chen, Lai-Zhong; Xu, Yun-Fei; Wang, Hong-Mei; Gong, Zheng; Cui, Guo-Ying; Yu, Xiao; Wang, Kai; Yao, Wei; Xin, Tao; Li, Min-Yong; Xiao, Kun-Hong; An, Xiao-fei; Huo, Yuqing; Xu, Zhi-gang; Sun, Jin-Peng; Pang, Qi

2013-01-01

Striatal-enriched tyrosine phosphatase (STEP) is an important regulator of neuronal synaptic plasticity, and its abnormal level or activity contributes to cognitive disorders. One crucial downstream effector and direct substrate of STEP is extracellular signal-regulated protein kinase (ERK), which has important functions in spine stabilisation and action potential transmission. The inhibition of STEP activity toward phospho-ERK has the potential to treat neuronal diseases, but the detailed mechanism underlying the dephosphorylation of phospho-ERK by STEP is not known. Therefore, we examined STEP activity toward pNPP, phospho-tyrosine-containing peptides, and the full-length phospho-ERK protein using STEP mutants with different structural features. STEP was found to be a highly efficient ERK tyrosine phosphatase that required both its N-terminal regulatory region and key residues in its active site. Specifically, both KIM and KIS of STEP were required for ERK interaction. In addition to the N-terminal KIS region, S245, hydrophobic residues L249/L251, and basic residues R242/R243 located in the KIM region were important in controlling STEP activity toward phospho-ERK. Further kinetic experiments revealed subtle structural differences between STEP and HePTP that affected the interactions of their KIMs with ERK. Moreover, STEP recognised specific positions of a phospho-ERK peptide sequence through its active site, and the contact of STEP F311 with phospho-ERK V205 and T207 were crucial interactions. Taken together, our results not only provide the information for interactions between ERK and STEP, but will also help in the development of specific strategies to target STEP-ERK recognition, which could serve as a potential therapy for neurological disorders. PMID:24117863

Quantitative Characteristics of Gene Regulation by Small RNA

PubMed Central

Levine, Erel; Zhang, Zhongge; Kuhlman, Thomas; Hwa, Terence

2007-01-01

An increasing number of small RNAs (sRNAs) have been shown to regulate critical pathways in prokaryotes and eukaryotes. In bacteria, regulation by trans-encoded sRNAs is predominantly found in the coordination of intricate stress responses. The mechanisms by which sRNAs modulate expression of its targets are diverse. In common to most is the possibility that interference with the translation of mRNA targets may also alter the abundance of functional sRNAs. Aiming to understand the unique role played by sRNAs in gene regulation, we studied examples from two distinct classes of bacterial sRNAs in Escherichia coli using a quantitative approach combining experiment and theory. Our results demonstrate that sRNA provides a novel mode of gene regulation, with characteristics distinct from those of protein-mediated gene regulation. These include a threshold-linear response with a tunable threshold, a robust noise resistance characteristic, and a built-in capability for hierarchical cross-talk. Knowledge of these special features of sRNA-mediated regulation may be crucial toward understanding the subtle functions that sRNAs can play in coordinating various stress-relief pathways. Our results may also help guide the design of synthetic genetic circuits that have properties difficult to attain with protein regulators alone. PMID:17713988

48 CFR 3036.104-90 - Authority for one-step turn-key design-build contracting for the United States Coast Guard (USCG).

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 7 2010-10-01 2010-10-01 false Authority for one-step... General 3036.104-90 Authority for one-step turn-key design-build contracting for the United States Coast Guard (USCG). The Head of the Contracting Activity (HCA) of the U.S. Coast Guard may use one-step turn...

Improving government regulations: a guidebook for conservation and renewable energy

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

Neese, R. J.; Scheer, R. M.; Marasco, A. L.

1981-04-01

An integrated view of the Office of Conservation and Solar Energy (CS) policy making encompassing both administrative procedures and policy analysis is presented. Chapter One very briefly sketches each step in the development of a significant regulation, noting important requirements and participants. Chapter Two expands upon the Overview, providing the details of the process, the rationale and source of requirements, concurrence procedures, and advice on the timing and synchronization of steps. Chapter Three explains the types of analysis documents that may be required for a program. Regulatory Analyses, Environmental Impact Statements, Urban and Community Impact Analyses, and Regulatory Flexibility Analysesmore » are all discussed. Specific information to be included in the documents and the circumstances under which the documents need to be prepared are explained. Chapter Four is a step-by-step discussion of how to do good analysis. Use of models and data bases is discussed. Policy objectives, alternatives, and decision making are explained. In Chapter five guidance is provided on identifying the public that would most likely be interested in the regulation, involving its constituents in a dialogue with CS, evaluating and handling comments, and engineering the final response. Chapter Six provides direction on planning the evaluation, monitoring the regulation's success once it has been promulgated, and allowing for constructive support or criticism from outside DOE. (MCW)« less

Role of SKP1-CUL1-F-Box-Protein (SCF) E3 Ubiquitin Ligases in Skin Cancer

PubMed Central

Xie, Chuan-Ming; Wei, Wenyi; Sun, Yi

2013-01-01

Many biological processes such as cell proliferation, differentiation, and cell death depend precisely on the timely synthesis and degradation of key regulatory proteins. While protein synthesis can be regulated at multiple levels, protein degradation is mainly controlled by the ubiquitin—proteasome system (UPS), which consists of two distinct steps: (1) ubiquitylation of targeted protein by E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme and E3 ubiquitin ligase, and (2) subsequent degradation by the 26S proteasome. Among all E3 ubiquitin ligases, the SCF (SKP1-CUL1-F-box protein) E3 ligases are the largest family and are responsible for the turnover of many key regulatory proteins. Aberrant regulation of SCF E3 ligases is associated with various human diseases, such as cancers, including skin cancer. In this review, we provide a comprehensive overview of all currently published data to define a promoting role of SCF E3 ligases in the development of skin cancer. The future directions in this area of research are also discussed with an ultimate goal to develop small molecule inhibitors of SCF E3 ligases as a novel approach for the treatment of human skin cancer. Furthermore, altered components or substrates of SCF E3 ligases may also be developed as the biomarkers for early diagnosis or predicting prognosis. PMID:23522382

Pax2/8 proteins coordinate sequential induction of otic and epibranchial placodes through differential regulation of foxi1, sox3 and fgf24

PubMed Central

Padanad, Mahesh S.; Riley, Bruce B.

2011-01-01

Vertebrate cranial placodes contribute vitally to development of sensory structures of the head. Amongst posterior placodes, the otic placode forms the inner ear whereas nearby epibranchial placodes produce sensory ganglia within branchial clefts. Though diverse in fate, these placodes show striking similarities in their early regulation. In zebrafish, both are initiated by localized Fgf signaling plus the ubiquitous competence factor Foxi1, and both express pax8 and sox3 in response. It has been suggested that Fgf initially induces a common otic/epibranchial field, which later subdivides in response to other signals. However, we find that otic and epibranchial placodes form at different times and by distinct mechanisms. Initially, Fgf from surrounding tissues induces otic expression of pax8 and sox3, which cooperate synergistically to establish otic fate. Subsequently, pax8 works with related genes pax2a/pax2b to downregulate otic expression of foxi1, a necessary step for further otic development. Additionally, pax2/8 activate otic expression of fgf24, which induces epibranchial expression of sox3. Knockdown of fgf24 or sox3 causes severe epibranchial deficiencies but has little effect on otic development. These findings clarify the roles of pax8 and sox3 and support a model whereby the otic placode forms first and induces epibranchial placodes through an Fgf-relay. PMID:21215261

Ca2+ influx does not trigger glucose-induced traffic of the insulin granules and alteration of their distribution.

PubMed

Niki, Ichiro; Niwa, Tae; Yu, Wei; Budzko, Dorota; Miki, Takashi; Senda, Takao

2003-11-01

This study investigated mechanisms by which glucose increases readily releasable secretory granules via acting on preexocytotic steps, i.e., intracellular granule movement and granule access to the plasma membrane using a pancreatic beta-cell line, MIN6. Glucose-induced activation of the movement occurred at a substimulatory concentration with regard to insulin output. Glucose activation of the movement was inhibited by pretreatment with thapsigargin plus acetylcholine to suppress intracellular Ca2+ mobilization. Inhibitors of calmodulin and myosin light chain kinase also suppressed glucose activation of the movement. Simultaneous addition of glucose with Ca2+ channel blockers or the ATP-sensitive K+ channel opener diazoxide failed to suppress the traffic activation, and addition of these substances on top of glucose stimulation resulted in a further increase. Although stimulatory glucose had minimal changes in the intracellular granule distribution, inhibition of Ca2+ influx revealed increases by glucose of the granules in the cell periphery. In contrast, high K+ depolarization decreased the peripheral granules. Glucose-induced granule margination was abolished when the protein kinase C activity was downregulated. These findings indicate that preexocytotic control of insulin release is regulated by distinct mechanisms from Ca2+ influx, which triggers insulin exocytosis. The nature of the regulation by glucose may explain a part of potentiating effects of the hexose independent of the closure of the ATP-sensitive K+ channel.

Fission yeast Ccq1 is a modulator of telomerase activity

PubMed Central

Armstrong, Christine A; Moiseeva, Vera; Collopy, Laura C; Pearson, Siân R; Ullah, Tomalika R; Xi, Shidong T; Martin, Jennifer; Subramaniam, Shaan; Marelli, Sara; Amelina, Hanna

2018-01-01

Abstract Shelterin, the telomeric protein complex, plays a crucial role in telomere homeostasis. In fission yeast, telomerase is recruited to chromosome ends by the shelterin component Tpz1 and its binding partner Ccq1, where telomerase binds to the 3′ overhang to add telomeric repeats. Recruitment is initiated by the interaction of Ccq1 with the telomerase subunit Est1. However, how telomerase is released following elongation remains to be established. Here, we show that Ccq1 also has a role in the suppression of telomere elongation, when coupled with the Clr4 histone H3 methyl-transferase complex and the Clr3 histone deacetylase and nucleosome remodelling complex, SHREC. We have dissected the functions of Ccq1 by establishing a Ccq1-Est1 fusion system, which bypasses the telomerase recruitment step. We demonstrate that Ccq1 forms two distinct complexes for positive and negative telomerase regulation, with Est1 and Clr3 respectively. The negative form of Ccq1 promotes dissociation of Ccq1-telomerase from Tpz1, thereby restricting local telomerase activity. The Clr4 complex also has a negative regulation activity with Ccq1, independently of SHREC. Thus, we propose a model in which Ccq1-Est1 recruits telomerase to mediate telomere extension, whilst elongated telomeric DNA recruits Ccq1 with the chromatin-remodelling complexes, which in turn releases telomerase from the telomere. PMID:29216371

Competition between thyroid hormone receptor-associated protein (TRAP) 220 and transcriptional intermediary factor (TIF) 2 for binding to nuclear receptors. Implications for the recruitment of TRAP and p160 coactivator complexes.

PubMed

Treuter, E; Johansson, L; Thomsen, J S; Wärnmark, A; Leers, J; Pelto-Huikko, M; Sjöberg, M; Wright, A P; Spyrou, G; Gustafsson, J A

1999-03-05

Transcriptional activation by nuclear receptors (NRs) involves the concerted action of coactivators, chromatin components, and the basal transcription machinery. Crucial NR coactivators, which target primarily the conserved ligand-regulated activation (AF-2) domain, include p160 family members, such as TIF2, as well as p160-associated coactivators, such as CBP/p300. Because these coactivators possess intrinsic histone acetyltransferase activity, they are believed to function mainly by regulating chromatin-dependent transcriptional activation. Recent evidence suggests the existence of an additional NR coactivator complex, referred to as the thyroid hormone receptor-associated protein (TRAP) complex, which may function more directly as a bridging complex to the basal transcription machinery. TRAP220, the 220-kDa NR-binding subunit of the complex, has been identified in independent studies using both biochemical and genetic approaches. In light of the functional differences identified between p160 and TRAP coactivator complexes in NR activation, we have attempted to compare interaction and functional characteristics of TIF 2 and TRAP220. Our findings imply that competition between the NR-binding subunits of distinct coactivator complexes may act as a putative regulatory step in establishing either a sequential activation cascade or the formation of independent coactivator complexes.

Controlled free radical attack in the apoplast: A hypothesis for roles of O, N and S species in regulatory and polysaccharide cleavage events during rapid abscission by Azolla

PubMed Central

Cohen, Michael F.; Gurung, Sushma; Fukuto, Jon M.; Yamasaki, Hideo

2014-01-01

Shedding of organs by abscission is a key terminal step in plant development and stress responses. Cell wall (CW) loosening at the abscission zone can occur through a combination chain breakage of apoplastic polysaccharides and tension release of cellulose microfibrils. Two distinctly regulated abscission cleavage events are amenable to study in small water ferns of the genus Azolla; one is a rapid abscission induced by environmental stimuli such as heat or chemicals, and the other is an ethylene-induced process occurring more slowly through the action of hydrolytic enzymes. Although free radicals are suggested to be involved in the induction of rapid root abscission, its mechanism is not fully understood. The apoplast contains peroxidases, metal-binding proteins and phenolic compounds that potentially generate free radicals from H2O2 to cleave polysaccharides in the CW and middle lamella. Effects of various thiol-reactive agents implicate the action of apoplastic peroxidases having accessible cysteine thiols in rapid abscission. The Ca2+ dependency of rapid abscission may reflect the stabilization Ca2+ confers to peroxidase structure and binding to pectin. To spur further investigation, we present a hypothetical model for small signaling molecules H2O2 and NO and their derivatives in regulating, via modification of putative protein thiols, free radical attack of apoplastic polysaccharides. PMID:24467903

Association of CAD, a multifunctional protein involved in pyrimidine synthesis, with mLST8, a component of the mTOR complexes

PubMed Central

2013-01-01

Background mTOR is a genetically conserved serine/threonine protein kinase, which controls cell growth, proliferation, and survival. A multifunctional protein CAD, catalyzing the initial three steps in de novo pyrimidine synthesis, is regulated by the phosphorylation reaction with different protein kinases, but the relationship with mTOR protein kinase has not been known. Results CAD was recovered as a binding protein with mLST8, a component of the mTOR complexes, from HEK293 cells transfected with the FLAG-mLST8 vector. Association of these two proteins was confirmed by the co-immuoprecipitaiton followed by immunoblot analysis of transfected myc-CAD and FLAG-mLST8 as well as that of the endogenous proteins in the cells. Analysis using mutant constructs suggested that CAD has more than one region for the binding with mLST8, and that mLST8 recognizes CAD and mTOR in distinct ways. The CAD enzymatic activity decreased in the cells depleted of amino acids and serum, in which the mTOR activity is suppressed. Conclusion The results obtained indicate that mLST8 bridges between CAD and mTOR, and plays a role in the signaling mechanism where CAD is regulated in the mTOR pathway through the association with mLST8. PMID:23594158

Jmy regulates oligodendrocyte differentiation via modulation of actin cytoskeleton dynamics.

PubMed

Azevedo, Maria M; Domingues, Helena S; Cordelières, Fabrice P; Sampaio, Paula; Seixas, Ana I; Relvas, João B

2018-05-06

During central nervous system development, oligodendrocytes form structurally and functionally distinct actin-rich protrusions that contact and wrap around axons to assemble myelin sheaths. Establishment of axonal contact is a limiting step in myelination that relies on the oligodendrocyte's ability to locally coordinate cytoskeletal rearrangements with myelin production, under the control of a transcriptional differentiation program. The molecules that provide fine-tuning of actin dynamics during oligodendrocyte differentiation and axon ensheathment remain largely unidentified. We performed transcriptomics analysis of soma and protrusion fractions from rat brain oligodendrocyte progenitors and found a subcellular enrichment of mRNAs in newly-formed protrusions. Approximately 30% of protrusion-enriched transcripts encode proteins related to cytoskeleton dynamics, including the junction mediating and regulatory protein Jmy, a multifunctional regulator of actin polymerization. Here, we show that expression of Jmy is upregulated during myelination and is required for the assembly of actin filaments and protrusion formation during oligodendrocyte differentiation. Quantitative morphodynamics analysis of live oligodendrocytes showed that differentiation is driven by a stereotypical actin network-dependent "cellular shaping" program. Disruption of actin dynamics via knockdown of Jmy leads to a program fail resulting in oligodendrocytes that do not acquire an arborized morphology and are less efficient in contacting neurites and forming myelin wraps in co-cultures with neurons. Our findings provide new mechanistic insight into the relationship between cell shape dynamics and differentiation in development. © 2018 Wiley Periodicals, Inc.

Human physiological models of insomnia.

PubMed

Richardson, Gary S

2007-12-01

Despite the wide prevalence and important consequences of insomnia, remarkably little is known about its pathophysiology. Available models exist primarily in the psychological domain and derive from the demonstrated efficacy of behavioral treatment approaches to insomnia management. However, these models offer little specific prediction about the anatomic or physiological foundation of chronic primary insomnia. On the other hand, a growing body of data on the physiology of sleep supports a reasonably circumscribed overview of possible pathophysiological mechanisms, as well as the development of physiological models of insomnia to guide future research. As a pragmatic step, these models focus on primary insomnia, as opposed to comorbid insomnias, because the latter is by its nature a much more heterogeneous presentation, reflecting the effects of the distinct comorbid condition. Current understanding of the regulation of sleep and wakefulness in mammalian brain supports four broad candidate areas: 1) disruption of the sleep homeostat; 2) disruption of the circadian clock; 3) disruption of intrinsic systems responsible for the expression of sleep states; or 4) disruption (hyperactivity) of extrinsic systems capable of over-riding normal sleep-wake regulation. This review examines each of the four candidate pathophysiological mechanisms and the available data in support of each. While studies that directly test the viability of each model are not yet available, descriptive data on primary insomnia favor the involvement of dysfunctional extrinsic stress-response systems in the pathology of primary chronic insomnia.

Comparative functional analyses of ultrabithorax reveal multiple steps and paths to diversification of legs in the adaptive radiation of semi-aquatic insects.

PubMed

Khila, Abderrahman; Abouheif, Ehab; Rowe, Locke

2014-08-01

Invasion of new ecological habitats is often associated with lineage diversification, yet the genetic changes underlying invasions and radiations are poorly understood. Over 200 million years ago, the semi-aquatic insects invaded water surface from a common terrestrial ancestor and diversified to exploit a wide array of niches. Here, we uncover the changes in regulation and function of the gene Ultrabithorax associated with both the invasion of water surface and the subsequent diversification of the group. In the common ancestor of the semi-aquatic insects, a novel deployment of Ubx protein in the mid-legs increased their length, thereby enhancing their role in water surface walking. In derived lineages that specialize in rowing on the open water, additional changes in the timing of Ubx expression further elongated the mid-legs thereby facilitating their function as oars. In addition, Ubx protein function was selectively reversed to shorten specific rear-leg segments, thereby enabling their function as rudders. These changes in Ubx have generated distinct niche-specialized morphologies that account for the remarkable diversification of the semi-aquatic insects. Therefore, changes in the regulation and function of a key developmental gene may facilitate both the morphological change necessary to transition to novel habitats and fuel subsequent morphological diversification. © 2014 The Author(s). Evolution © 2014 The Society for the Study of Evolution.

Soluble N-Ethylmaleimide-Sensitive Factor Attachment Protein Receptor-Derived Peptides for Regulation of Mast Cell Degranulation.

PubMed

Yang, Yoosoo; Kong, Byoungjae; Jung, Younghoon; Park, Joon-Bum; Oh, Jung-Mi; Hwang, Jaesung; Cho, Jae Youl; Kweon, Dae-Hyuk

2018-01-01

Vesicle-associated V-soluble N -ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins and target membrane-associated T-SNAREs (syntaxin 4 and SNAP-23) assemble into a core trans -SNARE complex that mediates membrane fusion during mast cell degranulation. This complex plays pivotal roles at various stages of exocytosis from the initial priming step to fusion pore opening and expansion, finally resulting in the release of the vesicle contents. In this study, peptides with the sequences of various SNARE motifs were investigated for their potential inhibitory effects against SNARE complex formation and mast cell degranulation. The peptides with the sequences of the N-terminal regions of vesicle-associated membrane protein 2 (VAMP2) and VAMP8 were found to reduce mast cell degranulation by inhibiting SNARE complex formation. The fusion of protein transduction domains to the N-terminal of each peptide enabled the internalization of the fusion peptides into the cells equally as efficiently as cell permeabilization by streptolysin-O without any loss of their inhibitory activities. Distinct subsets of mast cell granules could be selectively regulated by the N-terminal-mimicking peptides derived from VAMP2 and VAMP8, and they effectively decreased the symptoms of atopic dermatitis in mouse models. These results suggest that the cell membrane fusion machinery may represent a therapeutic target for atopic dermatitis.

Protocol for the "four steps to control your fatigue (4-STEPS)" randomised controlled trial: a self-regulation based physical activity intervention for patients with unexplained chronic fatigue

PubMed Central

2012-01-01

Background Unexplained Chronic Fatigue is a medical condition characterized by the presence of persistent, severe and debilitating medically unexplained fatigue, leading to impaired functioning and lower quality of life. Research suggests that physical activity can contribute to the reduction of fatigue and other somatic symptoms and can thus significantly improve physical functioning and quality of life in these patients. Based on the self-regulation (SR) theory of behaviour change, we developed a brief physical activity program for patients suffering from unexplained chronic fatigue which focuses on the training of self-regulation skills, the "4-STEPS to control your fatigue" program. Methods/Design This is a multi-centre, randomised controlled trial (RCT) that will be carried out in local primary care centres and at the Portuguese Fibromyalgia and Chronic Fatigue Syndrome Patients Association. Patients aged between 18 and 65 and fulfilling operationalized criteria for Idiopathic Chronic Fatigue (ICF) and Chronic Fatigue Syndrome (CFS) will be recruited and randomly allocated to standard care (SC) or standard care plus a self-regulation based physical activity program (4-STEPS). Patients will be assessed at baseline, after the intervention (3 months) and at 12 months follow-up. The primary outcome is fatigue severity. Discussion The results of the RCT will provide information about the effectiveness of a brief self-regulation intervention for promoting physical activity in patients with unexplained chronic fatigue. If the program proves to be effective, it may be considered as an adjunctive treatment for these patients. Trial Registration ISRCTN: ISRCTN70763996 PMID:22429404

Moral Psychology Is Relationship Regulation: Moral Motives for Unity, Hierarchy, Equality, and Proportionality

ERIC Educational Resources Information Center

Rai, Tage Shakti; Fiske, Alan Page

2011-01-01

Genuine moral disagreement exists and is widespread. To understand such disagreement, we must examine the basic kinds of social relationships people construct across cultures and the distinct moral obligations and prohibitions these relationships entail. We extend relational models theory (Fiske, 1991) to identify 4 fundamental and distinct moral…

Expanding the Concepts in Protein Structure-Function Relationships and Enzyme Kinetics: Teaching Using Morpheeins

ERIC Educational Resources Information Center

Lawrence, Sarah H.; Jaffe, Eileen K.

2008-01-01

A morpheein is a homo-oligomeric protein that can exist as an ensemble of physiologically significant and functionally distinct alternate quaternary assemblies. Morpheeins exist in nature and use conformational equilibria between different tertiary structures to form distinct oligomers as a means of regulating their function. Notably, alternate…

Distinct transcriptome profiles differentiate NSAID-dependent from NSAID-independent food anaphylaxis

PubMed Central

Muñoz-Cano, Rosa; Pascal, Mariona; Bartra, Joan; Picado, Cesar; Valero, Antonio; Kim, Do-Kyun; Brooks, Stephen; Ombrello, Michael; Metcalfe, Dean D.; Rivera, Juan; Olivera, Ana

2015-01-01

Background Lipid transfer protein (LTP), an abundant protein in fruits, vegetables and nuts, is a common food allergen in Mediterranean areas causing diverse allergic reactions. Approximately 40% of food anaphylaxis induced by LTP require non-steroidal anti-inflammatory drugs (NSAIDs) as a triggering cofactor. Objective To better understand the determinants of NSAID-dependent (NSAID-LTP-A) and NSAID-independent LTP-anaphylaxis (LTP-A) Methods Selection of patients was based on a proven clinical history of NSAID-dependent or -independent anaphylaxis to LTP, positive skin prick test to LTP and serum LTP-IgE. Whole transcriptome (RNA-Seq) analysis of blood cells from 14 individuals with NSAID-LTP-A, 7 with LTP-A and 13 healthy controls was performed to identify distinct gene expression signatures. Results Expression of genes regulating gastrointestinal epithelium renewal was altered in both patient sets, particularly in LTP-A, who also presented gene expression profiles characteristic of an inflammatory syndrome. These included altered B cell pathways, increased neutrophil activation markers and elevated levels of reactive oxygen species. Increased expression of the IgG receptor (CD64) in LTP-A patients was mirrored by the presence of LTP-specific IgG1 and 3. Conversely, NSAID-LTP-A patients were characterized by reduced expression of IFN-γ-regulated genes and IFN-γ levels as well as up-regulated adenosine receptor 3 (ADORA3) expression and genes related to adenosine metabolism. Conclusions Gene ontology analysis suggests disturbances in gut epithelium homeostasis in both LTP-related anaphylaxis groups with potential integrity breaches in LTP-A that may explain their distinct inflammatory signature. Differential regulation in LTP-A and NSAID-LTP-A of the IFN-γ pathway, IgG receptors and ADORA3 may provide the pathogenic basis of their distinct responses. PMID:26194548

The evolution of a colluvial hollow to a fluvial channel with periodic steps following two transformational disturbances: A wildfire and a historic flood

NASA Astrophysics Data System (ADS)

Rengers, F. K.; McGuire, L. A.; Ebel, B. A.; Tucker, G. E.

2018-05-01

The transition of a colluvial hollow to a fluvial channel with discrete steps was observed after two landscape-scale disturbances. The first disturbance, a high-severity wildfire, changed the catchment hydrology to favor overland flow, which incised a colluvial hollow, creating a channel in the same location. This incised channel became armored with cobbles and boulders following repeated post-wildfire overland flow events. Three years after the fire, a record rainstorm produced regional flooding and generated sufficient fluvial erosion and sorting to produce a fluvial channel with periodically spaced steps. An analysis of the step spacing shows that after the flood, newly formed steps retained a similar spacing to the topographic roughness spacing in the original colluvial hollow (prior to channelization). This suggests that despite a distinct change in channel form roughness and bedform morphology, the endogenous roughness periodicity was conserved. Variations in sediment erodibility helped to create the emergent steps as the largest particles (>D84) remained immobile, becoming step features, and downstream soil was easily winnowed away.

The evolution of a colluvial hollow to a fluvial channel with periodic steps following two transformational disturbances: A wildfire and a historic flood

USGS Publications Warehouse

Rengers, Francis K.; McGuire, Luke; Ebel, Brian A.; Tucker, G. E.

2018-01-01

The transition of a colluvial hollow to a fluvial channel with discrete steps was observed after two landscape-scale disturbances. The first disturbance, a high-severity wildfire, changed the catchment hydrology to favor overland flow, which incised a colluvial hollow, creating a channel in the same location. This incised channel became armored with cobbles and boulders following repeated post-wildfire overland flow events. Three years after the fire, a record rainstorm produced regional flooding and generated sufficient fluvial erosion and sorting to produce a fluvial channel with periodically spaced steps. An analysis of the step spacing shows that after the flood, newly formed steps retained a similar spacing to the topographic roughness spacing in the original colluvial hollow (prior to channelization). This suggests that despite a distinct change in channel form roughness and bedform morphology, the endogenous roughness periodicity was conserved. Variations in sediment erodibility helped to create the emergent steps as the largest particles ( >D84) remained immobile, becoming step features, and downstream soil was easily winnowed away.

Live imaging and genetic analysis of mouse notochord formation reveals regional morphogenetic mechanisms.

PubMed

Yamanaka, Yojiro; Tamplin, Owen J; Beckers, Anja; Gossler, Achim; Rossant, Janet

2007-12-01

The node and notochord have been extensively studied as signaling centers in the vertebrate embryo. The morphogenesis of these tissues, particularly in mouse, is not well understood. Using time-lapse live imaging and cell lineage tracking, we show the notochord has distinct morphogenetic origins along the anterior-posterior axis. The anterior head process notochord arises independently of the node by condensation of dispersed cells. The trunk notochord is derived from the node and forms by convergent extension. The tail notochord forms by node-derived progenitors that actively migrate toward the posterior. We also reveal distinct genetic regulation within these different regions. We show that Foxa2 compensates for and genetically interacts with Noto in the trunk notochord, and that Noto has an evolutionarily conserved role in regulating axial versus paraxial cell fate. Therefore, we propose three distinct regions within the mouse notochord, each with unique morphogenetic origins.

South Africa’s Technology Sector

DTIC Science & Technology

2007-08-01

Somchem’s rocket motor propellant casting pits were destroyed and sealed with concrete .135 In taking this step, South Africa became the only country...principles of restraint, responsibility, and translucence .163 Translucence or semi-transparency is distinct from the more rigorous concept of total

The Epidemiology of Observed Temperament: Factor Structure and Demographic Group Differences

PubMed Central

Willoughby, Michael T.; Stifter, Cynthia A.; Gottfredson, Nisha C.

2015-01-01

This study investigated the factor structure of observational indicators of children’s temperament that were collected across the first three years of life in the Family Life Project (N = 1205) sample. A four-factor model (activity level, fear, anger, regulation), which corresponded broadly to Rothbart’s distinction between reactivity and regulation, provided an acceptable fit the observed data. Tests of measurement invariance demonstrated that a majority of the observational indicators exhibited comparable measurement properties for male vs. female, black vs. white, and poor vs. not-poor children, which improved the generalizability of these results. Unadjusted demographic group comparisons revealed small to moderate sized differences (Cohen ds = |.23 – .42|) in temperamental reactivity and moderate to large sized differences (Cohen ds = −.64 – −.97) in regulation. Collectively, demographic variables explained more of the variation in regulation (R2 = .25) than in reactivity (R2 = .02 – .06). Follow-up analyses demonstrated that race differences were substantially diminished in magnitude and better accounted for by poverty. These results help to validate the distinction between temperamental reactivity and regulation using observational indicators. PMID:25733489

Analysis of Strengths, Weaknesses, Opportunities, and Threats as a Tool for Translating Evidence into Individualized Medical Strategies (I-SWOT)

PubMed Central

von Kodolitsch, Yskert; Bernhardt, Alexander M.; Robinson, Peter N.; Kölbel, Tilo; Reichenspurner, Hermann; Debus, Sebastian; Detter, Christian

2015-01-01

Background It is the physicians’ task to translate evidence and guidelines into medical strategies for individual patients. Until today, however, there is no formal tool that is instrumental to perform this translation. Methods We introduce the analysis of strengths (S) and weaknesses (W) related to therapy with opportunities (O) and threats (T) related to individual patients as a tool to establish an individualized (I) medical strategy (I-SWOT). The I-SWOT matrix identifies four fundamental types of strategy. These comprise “SO” maximizing strengths and opportunities, “WT” minimizing weaknesses and threats, “WO” minimizing weaknesses and maximizing opportunities, and “ST” maximizing strengths and minimizing threats. Each distinct type of strategy may be considered for individualized medical strategies. Results We describe four steps of I-SWOT to establish an individualized medical strategy to treat aortic disease. In the first step, we define the goal of therapy and identify all evidence-based therapeutic options. In a second step, we assess strengths and weaknesses of each therapeutic option in a SW matrix form. In a third step, we assess opportunities and threats related to the individual patient, and in a final step, we use the I-SWOT matrix to establish an individualized medical strategy through matching “SW” with “OT”. As an example we present two 30-year-old patients with Marfan syndrome with identical medical history and aortic pathology. As a result of I-SWOT analysis of their individual opportunities and threats, we identified two distinct medical strategies in these patients. Conclusion I-SWOT is a formal but easy to use tool to translate medical evidence into individualized medical strategies. PMID:27069939

Analysis of Strengths, Weaknesses, Opportunities, and Threats as a Tool for Translating Evidence into Individualized Medical Strategies (I-SWOT).

PubMed

von Kodolitsch, Yskert; Bernhardt, Alexander M; Robinson, Peter N; Kölbel, Tilo; Reichenspurner, Hermann; Debus, Sebastian; Detter, Christian

2015-06-01

It is the physicians' task to translate evidence and guidelines into medical strategies for individual patients. Until today, however, there is no formal tool that is instrumental to perform this translation. We introduce the analysis of strengths (S) and weaknesses (W) related to therapy with opportunities (O) and threats (T) related to individual patients as a tool to establish an individualized (I) medical strategy (I-SWOT). The I-SWOT matrix identifies four fundamental types of strategy. These comprise "SO" maximizing strengths and opportunities, "WT" minimizing weaknesses and threats, "WO" minimizing weaknesses and maximizing opportunities, and "ST" maximizing strengths and minimizing threats. Each distinct type of strategy may be considered for individualized medical strategies. We describe four steps of I-SWOT to establish an individualized medical strategy to treat aortic disease. In the first step, we define the goal of therapy and identify all evidence-based therapeutic options. In a second step, we assess strengths and weaknesses of each therapeutic option in a SW matrix form. In a third step, we assess opportunities and threats related to the individual patient, and in a final step, we use the I-SWOT matrix to establish an individualized medical strategy through matching "SW" with "OT". As an example we present two 30-year-old patients with Marfan syndrome with identical medical history and aortic pathology. As a result of I-SWOT analysis of their individual opportunities and threats, we identified two distinct medical strategies in these patients. I-SWOT is a formal but easy to use tool to translate medical evidence into individualized medical strategies.

Ultrasound Microbubble Treatment Enhances Clathrin-Mediated Endocytosis and Fluid-Phase Uptake through Distinct Mechanisms.

PubMed

Fekri, Farnaz; Delos Santos, Ralph Christian; Karshafian, Raffi; Antonescu, Costin N

2016-01-01

Drug delivery to tumors is limited by several factors, including drug permeability of the target cell plasma membrane. Ultrasound in combination with microbubbles (USMB) is a promising strategy to overcome these limitations. USMB treatment elicits enhanced cellular uptake of materials such as drugs, in part as a result of sheer stress and formation of transient membrane pores. Pores formed upon USMB treatment are rapidly resealed, suggesting that other processes such as enhanced endocytosis may contribute to the enhanced material uptake by cells upon USMB treatment. How USMB regulates endocytic processes remains incompletely understood. Cells constitutively utilize several distinct mechanisms of endocytosis, including clathrin-mediated endocytosis (CME) for the internalization of receptor-bound macromolecules such as Transferrin Receptor (TfR), and distinct mechanism(s) that mediate the majority of fluid-phase endocytosis. Tracking the abundance of TfR on the cell surface and the internalization of its ligand transferrin revealed that USMB acutely enhances the rate of CME. Total internal reflection fluorescence microscopy experiments revealed that USMB treatment altered the assembly of clathrin-coated pits, the basic structural units of CME. In addition, the rate of fluid-phase endocytosis was enhanced, but with delayed onset upon USMB treatment relative to the enhancement of CME, suggesting that the two processes are distinctly regulated by USMB. Indeed, vacuolin-1 or desipramine treatment prevented the enhancement of CME but not of fluid phase endocytosis upon USMB, suggesting that lysosome exocytosis and acid sphingomyelinase, respectively, are required for the regulation of CME but not fluid phase endocytosis upon USMB treatment. These results indicate that USMB enhances both CME and fluid phase endocytosis through distinct signaling mechanisms, and suggest that strategies for potentiating the enhancement of endocytosis upon USMB treatment may improve targeted drug delivery.

Carboxyl-terminal Domain of Transient Receptor Potential Vanilloid 1 Contains Distinct Segments Differentially Involved in Capsaicin- and Heat-induced Desensitization*

PubMed Central

Joseph, John; Wang, Sen; Lee, Jongseok; Ro, Jin Y.; Chung, Man-Kyo

2013-01-01

Multiple Ca2+-dependent processes are involved in capsaicin-induced desensitization of transient receptor potential vanilloid 1 (TRPV1), but desensitization of TRPV1 by heat occurs even in the absence of extracellular Ca2+, although the mechanisms are unknown. In this study, we tested the hypothesis that capsaicin and heat desensitize TRPV1 through distinct mechanisms involving distinct structural segments of TRPV1. In HEK293 cells that heterologously express TRPV1, we found that heat-induced desensitization was not affected by the inclusion of intracellular ATP or alanine mutation of Lys155, both of which attenuate capsaicin-induced desensitization, suggesting that heat-induced desensitization occurs through mechanisms distinct from capsaicin-induced desensitization. To determine protein domains involved in heat-induced desensitization, we generated chimeric proteins between TRPV1 and TRPV3, a heat-gated channel lacking heat-induced desensitization. We found that TRPV1 with the carboxyl-terminal domain (CTD) of TRPV3 retained heat activation but was impaired in heat-induced desensitization. Further experiments using chimeric or deletion mutants within TRPV1 CTD indicated that the distal half of CTD regulates the activation and desensitization of TRPV1 in modality-specific manners. Within the distal CTD, we identified two segments that distinctly regulated capsaicin- and heat-induced desensitization. The results suggest that the activation and desensitization of TRPV1 by capsaicin and heat can be modulated differentially and disproportionally through different regions of TRPV1 CTD. Identifying the domains involved in thermal regulation of TRPV1 may facilitate the development of novel anti-hyperalgesic approaches aimed at attenuating activation and enhancing desensitization of TRPV1 by thermal stimuli. PMID:24174527

Distinct regulatory functions of SLP-76 and MIST in NK cell cytotoxicity and IFN-gamma production.

PubMed

Hidano, Shinya; Sasanuma, Hiroki; Ohshima, Keiko; Seino, Ken-ichiro; Kumar, Lalit; Hayashi, Katsuhiko; Hikida, Masaki; Kurosaki, Tomohiro; Taniguchi, Masaru; Geha, Raif S; Kitamura, Daisuke; Goitsuka, Ryo

2008-03-01

Activation of NK cells is triggered by multiple receptors. We demonstrate here that SLP-76 is required for CD16- and NKG2D-mediated NK cell cytotoxicity, while MIST negatively regulates these responses in an SLP-76-dependent manner. Exceptionally, MIST acts as a positive regulator of cytotoxicity against YAC-1 cells, although SLP-76 plays a more key role. SLP-76 acts as a dominant positive regulator for both NKG2D-mediated and YAC-1 cell-triggered IFN-gamma production. Although NKG2D-mediated IFN-gamma production depends on phospholipase C (PLC) gamma 2, YAC-1 cell-triggered IFN-gamma production is PLC gamma 2- and Syk/ZAP-70 independent and nuclear factor-kappa B mediated. SLP-76 is required for this process in the presence of MIST but is dispensable in the absence of MIST. Thus, YAC-1 cell-triggered NKG2D-independent IFN-gamma production appears to be regulated by SLP-76-dependent and -independent pathways, in which the latter is negatively regulated by MIST. Taken together, these results suggest that SLP-76 and MIST distinctly but interactively regulate NK cell cytotoxicity and IFN-gamma production.

The B-cell identity factor Pax5 regulates distinct transcriptional programmes in early and late B lymphopoiesis

PubMed Central

Revilla-i-Domingo, Roger; Bilic, Ivan; Vilagos, Bojan; Tagoh, Hiromi; Ebert, Anja; Tamir, Ido M; Smeenk, Leonie; Trupke, Johanna; Sommer, Andreas; Jaritz, Markus; Busslinger, Meinrad

2012-01-01

Pax5 controls the identity and development of B cells by repressing lineage-inappropriate genes and activating B-cell-specific genes. Here, we used genome-wide approaches to identify Pax5 target genes in pro-B and mature B cells. In these cell types, Pax5 bound to 40% of the cis-regulatory elements defined by mapping DNase I hypersensitive (DHS) sites, transcription start sites and histone modifications. Although Pax5 bound to 8000 target genes, it regulated only 4% of them in pro-B and mature B cells by inducing enhancers at activated genes and eliminating DHS sites at repressed genes. Pax5-regulated genes in pro-B cells account for 23% of all expression changes occurring between common lymphoid progenitors and committed pro-B cells, which identifies Pax5 as an important regulator of this developmental transition. Regulated Pax5 target genes minimally overlap in pro-B and mature B cells, which reflects massive expression changes between these cell types. Hence, Pax5 controls B-cell identity and function by regulating distinct target genes in early and late B lymphopoiesis. PMID:22669466

Distinct emotion regulation skills explain psychopathology and problems in social relationships following childhood emotional abuse and neglect.

PubMed

Berzenski, Sara R

2018-03-22

Efforts to differentiate between the developmental sequelae of childhood emotional abuse and childhood emotional neglect are critical to both research and practice efforts. As an oft-identified mechanism of the effects of child maltreatment on later adjustment, emotion dysregulation represents a key potential pathway. The present study explored a higher order factor model of specific emotion regulation skills, and the extent to which these skill sets would indicate distinct developmental pathways from unique emotional maltreatment experiences to multidomain adjustment. A sample of 500 ethnoracially diverse college students reported on their experiences. A two-factor model of emotion regulation skills based on subscales of the Difficulties in Emotion Regulation Scale was revealed. Significant indirect effects of childhood emotional abuse on psychopathology and problems in social relationships were found through response-focused difficulties in emotion regulation, whereas a significant indirect effect of childhood emotional neglect on problems in social relationships was found through antecedent-focused difficulties in emotion regulation. These results are consistent with theoretical models and empirical evidence suggesting differential effects of childhood emotional abuse and emotional neglect, and provide an important indication for developing targeted interventions focusing on specific higher order emotion dysregulation skill clusters.

Starch biosynthesis in cassava: a genome-based pathway reconstruction and its exploitation in data integration

PubMed Central

2013-01-01

Background Cassava is a well-known starchy root crop utilized for food, feed and biofuel production. However, the comprehension underlying the process of starch production in cassava is not yet available. Results In this work, we exploited the recently released genome information and utilized the post-genomic approaches to reconstruct the metabolic pathway of starch biosynthesis in cassava using multiple plant templates. The quality of pathway reconstruction was assured by the employed parsimonious reconstruction framework and the collective validation steps. Our reconstructed pathway is presented in the form of an informative map, which describes all important information of the pathway, and an interactive map, which facilitates the integration of omics data into the metabolic pathway. Additionally, to demonstrate the advantage of the reconstructed pathways beyond just the schematic presentation, the pathway could be used for incorporating the gene expression data obtained from various developmental stages of cassava roots. Our results exhibited the distinct activities of the starch biosynthesis pathway in different stages of root development at the transcriptional level whereby the activity of the pathway is higher toward the development of mature storage roots. Conclusions To expand its applications, the interactive map of the reconstructed starch biosynthesis pathway is available for download at the SBI group’s website (http://sbi.pdti.kmutt.ac.th/?page_id=33). This work is considered a big step in the quantitative modeling pipeline aiming to investigate the dynamic regulation of starch biosynthesis in cassava roots. PMID:23938102

The impact of phosphate scarcity on pharmaceutical protein production in S. cerevisiae: linking transcriptomic insights to phenotypic responses

PubMed Central

2011-01-01

Background The adaptation of unicellular organisms like Saccharomyces cerevisiae to alternating nutrient availability is of great fundamental and applied interest, as understanding how eukaryotic cells respond to variations in their nutrient supply has implications spanning from physiological insights to biotechnological applications. Results The impact of a step-wise restricted supply of phosphate on the physiological state of S. cerevisiae cells producing human Insulin was studied. The focus was to determine the changes within the global gene expression of cells being cultured to an industrially relevant high cell density of 33 g/l cell dry weight and under six distinct phosphate concentrations, ranging from 33 mM (unlimited) to 2.6 mM (limited). An increased flux through the secretory pathway, being induced by the PHO circuit during low Pi supplementation, proved to enhance the secretory production of the heterologous protein. The re-distribution of the carbon flux from biomass formation towards increased glycerol production under low phosphate led to increased transcript levels of the insulin gene, which was under the regulation of the TPI1 promoter. Conclusions Our study underlines the dynamic character of adaptive responses of cells towards a change in their nutrient access. The gradual decrease of the phosphate supply resulted in a step-wise modulated phenotypic response, thereby alternating the specific productivity and the secretory flux. Our work emphasizes the importance of reduced phosphate supply for improved secretory production of heterologous proteins. PMID:22151908

Starch biosynthesis in cassava: a genome-based pathway reconstruction and its exploitation in data integration.

PubMed

Saithong, Treenut; Rongsirikul, Oratai; Kalapanulak, Saowalak; Chiewchankaset, Porntip; Siriwat, Wanatsanan; Netrphan, Supatcharee; Suksangpanomrung, Malinee; Meechai, Asawin; Cheevadhanarak, Supapon

2013-08-10

Cassava is a well-known starchy root crop utilized for food, feed and biofuel production. However, the comprehension underlying the process of starch production in cassava is not yet available. In this work, we exploited the recently released genome information and utilized the post-genomic approaches to reconstruct the metabolic pathway of starch biosynthesis in cassava using multiple plant templates. The quality of pathway reconstruction was assured by the employed parsimonious reconstruction framework and the collective validation steps. Our reconstructed pathway is presented in the form of an informative map, which describes all important information of the pathway, and an interactive map, which facilitates the integration of omics data into the metabolic pathway. Additionally, to demonstrate the advantage of the reconstructed pathways beyond just the schematic presentation, the pathway could be used for incorporating the gene expression data obtained from various developmental stages of cassava roots. Our results exhibited the distinct activities of the starch biosynthesis pathway in different stages of root development at the transcriptional level whereby the activity of the pathway is higher toward the development of mature storage roots. To expand its applications, the interactive map of the reconstructed starch biosynthesis pathway is available for download at the SBI group's website (http://sbi.pdti.kmutt.ac.th/?page_id=33). This work is considered a big step in the quantitative modeling pipeline aiming to investigate the dynamic regulation of starch biosynthesis in cassava roots.

Hydrocarbons and energy from plants: Final report, 1984-1987

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

Calvin, M.; Otvos, J.; Taylor, S.E.

1988-08-01

Plant hydrocarbon (isoprenoid) production was investigated as an alternative source to fossil fuels. Because of their high triterpenoid (hydrocarbon) content of 4--8%, Euphorbia lathyris plants were used as a model system for this study. The structure of the E. lathyris triterpenoids was determined, and triterpenoid biosynthesis studied to better understand the metabolic regulation of isoprenoid production. Triterpenoid biosynthesis occurs in two distinct tissue types in E. lathyris plants: in the latex of the laticifer cells; and in the mesophyll cells of the leaf and stem. The latex has been fractionated by centrifugation, and it has been determined that the latermore » steps of isoprenoid biosynthesis, the conversion of mevalonic acid to the triterpenes, are compartmentized within a vacuole. Also identified was the conversion of hydroxymethyl glutaryl-CoA to mevalonic acid, catalyzed by the enzyme Hydroxymethyl glutaryl-CoA Reductase, as a key rate limiting step in isoprenoid biosynthesis. At least two isozymes of this enzyme, one in the latex and another in the leaf plastids, have been identified. Environmental stress has been applied to plants to study changes in carbon allocation. Salinity stress caused a large decrease in growth, smaller decreases in photosynthesis, resulting in a larger allocation of carbon to both hydrocarbon and sugar production. An increase in Hydroxymethyl glutaryl-CoA Reductase activity was also observed when isoprenoid production increased. Other species where also screened for the production of hydrogen rich products such as isoprenoids and glycerides, and their hydrocarbon composition was determined.« less

A Cholinergic-Regulated Circuit Coordinates the Maintenance and Bi-Stable States of a Sensory-Motor Behavior during Caenorhabditis elegans Male Copulation

PubMed Central

Liu, Yishi; LeBeouf, Brigitte; Guo, Xiaoyan; Correa, Paola A.; Gualberto, Daisy G.; Lints, Robyn; Garcia, L. Rene

2011-01-01

Penetration of a male copulatory organ into a suitable mate is a conserved and necessary behavioral step for most terrestrial matings; however, the detailed molecular and cellular mechanisms for this distinct social interaction have not been elucidated in any animal. During mating, the Caenorhabditis elegans male cloaca is maintained over the hermaphrodite's vulva as he attempts to insert his copulatory spicules. Rhythmic spicule thrusts cease when insertion is sensed. Circuit components consisting of sensory/motor neurons and sex muscles for these steps have been previously identified, but it was unclear how their outputs are integrated to generate a coordinated behavior pattern. Here, we show that cholinergic signaling between the cloacal sensory/motor neurons and the posterior sex muscles sustains genital contact between the sexes. Simultaneously, via gap junctions, signaling from these muscles is transmitted to the spicule muscles, thus coupling repeated spicule thrusts with vulval contact. To transit from rhythmic to sustained muscle contraction during penetration, the SPC sensory-motor neurons integrate the signal of spicule's position in the vulva with inputs from the hook and cloacal sensilla. The UNC-103 K+ channel maintains a high excitability threshold in the circuit, so that sustained spicule muscle contraction is not stimulated by fewer inputs. We demonstrate that coordination of sensory inputs and motor outputs used to initiate, maintain, self-monitor, and complete an innate behavior is accomplished via the coupling of a few circuit components. PMID:21423722

Self-Regulated Strategy Development. What Works Clearinghouse Intervention Report

ERIC Educational Resources Information Center

What Works Clearinghouse, 2017

2017-01-01

"Self-Regulated Strategy Development" ("SRSD") is an intervention designed to improve students' academic skills through a six-step process that teaches students specific academic strategies and self-regulation skills. The practice is especially appropriate for students with learning disabilities, the focal population of the…

PTHrP and Indian hedgehog control differentiation of growth plate chondrocytes at multiple steps.

PubMed

Kobayashi, Tatsuya; Chung, Ung-Il; Schipani, Ernestina; Starbuck, Michael; Karsenty, Gerard; Katagiri, Takenobu; Goad, Dale L; Lanske, Beate; Kronenberg, Henry M

2002-06-01

In developing murine growth plates, chondrocytes near the articular surface (periarticular chondrocytes) proliferate, differentiate into flat column-forming proliferating cells (columnar chondrocytes), stop dividing and finally differentiate into hypertrophic cells. Indian hedgehog (Ihh), which is predominantly expressed in prehypertrophic cells, stimulates expression of parathyroid hormone (PTH)-related peptide (PTHrP) which negatively regulates terminal chondrocyte differentiation through the PTH/PTHrP receptor (PPR). However, the roles of PTHrP and Ihh in regulating earlier steps in chondrocyte differentiation are unclear. We present novel mouse models with PPR abnormalities that help clarify these roles. In mice with chondrocyte-specific PPR ablation and mice with reduced PPR expression, chondrocyte differentiation was accelerated not only at the terminal step but also at an earlier step: periarticular to columnar differentiation. In these models, upregulation of Ihh action in the periarticular region was also observed. In the third model in which the PPR was disrupted in about 30% of columnar chondrocytes, Ihh action in the periarticular chondrocytes was upregulated because of ectopically differentiated hypertrophic chondrocytes that had lost PPR. Acceleration of periarticular to columnar differentiation was also noted in this mouse, while most of periarticular chondrocytes retained PPR signaling. These data suggest that Ihh positively controls differentiation of periarticular chondrocytes independently of PTHrP. Thus, chondrocyte differentiation is controlled at multiple steps by PTHrP and Ihh through the mutual regulation of their activities.

Different CHD chromatin remodelers are required for expression of distinct gene sets and specific stages during development of Dictyostelium discoideum

PubMed Central

Platt, James L.; Rogers, Benjamin J.; Rogers, Kelley C.; Harwood, Adrian J.; Kimmel, Alan R.

2013-01-01

Control of chromatin structure is crucial for multicellular development and regulation of cell differentiation. The CHD (chromodomain-helicase-DNA binding) protein family is one of the major ATP-dependent, chromatin remodeling factors that regulate nucleosome positioning and access of transcription factors and RNA polymerase to the eukaryotic genome. There are three mammalian CHD subfamilies and their impaired functions are associated with several human diseases. Here, we identify three CHD orthologs (ChdA, ChdB and ChdC) in Dictyostelium discoideum. These CHDs are expressed throughout development, but with unique patterns. Null mutants lacking each CHD have distinct phenotypes that reflect their expression patterns and suggest functional specificity. Accordingly, using genome-wide (RNA-seq) transcriptome profiling for each null strain, we show that the different CHDs regulate distinct gene sets during both growth and development. ChdC is an apparent ortholog of the mammalian Class III CHD group that is associated with the human CHARGE syndrome, and GO analyses of aberrant gene expression in chdC nulls suggest defects in both cell-autonomous and non-autonomous signaling, which have been confirmed through analyses of chdC nulls developed in pure populations or with low levels of wild-type cells. This study provides novel insight into the broad function of CHDs in the regulation development and disease, through chromatin-mediated changes in directed gene expression. PMID:24301467

Autogenous cross-regulation of Quaking mRNA processing and translation balances Quaking functions in splicing and translation.

PubMed

Fagg, W Samuel; Liu, Naiyou; Fair, Jeffrey Haskell; Shiue, Lily; Katzman, Sol; Donohue, John Paul; Ares, Manuel

2017-09-15

Quaking protein isoforms arise from a single Quaking gene and bind the same RNA motif to regulate splicing, translation, decay, and localization of a large set of RNAs. However, the mechanisms by which Quaking expression is controlled to ensure that appropriate amounts of each isoform are available for such disparate gene expression processes are unknown. Here we explore how levels of two isoforms, nuclear Quaking-5 (Qk5) and cytoplasmic Qk6, are regulated in mouse myoblasts. We found that Qk5 and Qk6 proteins have distinct functions in splicing and translation, respectively, enforced through differential subcellular localization. We show that Qk5 and Qk6 regulate distinct target mRNAs in the cell and act in distinct ways on their own and each other's transcripts to create a network of autoregulatory and cross-regulatory feedback controls. Morpholino-mediated inhibition of Qk translation confirms that Qk5 controls Qk RNA levels by promoting accumulation and alternative splicing of Qk RNA, whereas Qk6 promotes its own translation while repressing Qk5. This Qk isoform cross-regulatory network responds to additional cell type and developmental controls to generate a spectrum of Qk5/Qk6 ratios, where they likely contribute to the wide range of functions of Quaking in development and cancer. © 2017 Fagg et al.; Published by Cold Spring Harbor Laboratory Press.

Clonal selection versus clonal cooperation: the integrated perception of immune objects

PubMed Central

Nataf, Serge

2016-01-01

Analogies between the immune and nervous systems were first envisioned by the immunologist Niels Jerne who introduced the concepts of antigen "recognition" and immune "memory". However, since then, it appears that only the cognitive immunology paradigm proposed by Irun Cohen, attempted to further theorize the immune system functions through the prism of neurosciences. The present paper is aimed at revisiting this analogy-based reasoning. In particular, a parallel is drawn between the brain pathways of visual perception and the processes allowing the global perception of an "immune object". Thus, in the visual system, distinct features of a visual object (shape, color, motion) are perceived separately by distinct neuronal populations during a primary perception task. The output signals generated during this first step instruct then an integrated perception task performed by other neuronal networks. Such a higher order perception step is by essence a cooperative task that is mandatory for the global perception of visual objects. Based on a re-interpretation of recent experimental data, it is suggested that similar general principles drive the integrated perception of immune objects in secondary lymphoid organs (SLOs). In this scheme, the four main categories of signals characterizing an immune object (antigenic, contextual, temporal and localization signals) are first perceived separately by distinct networks of immunocompetent cells.  Then, in a multitude of SLO niches, the output signals generated during this primary perception step are integrated by TH-cells at the single cell level. This process eventually generates a multitude of T-cell and B-cell clones that perform, at the scale of SLOs, an integrated perception of immune objects. Overall, this new framework proposes that integrated immune perception and, consequently, integrated immune responses, rely essentially on clonal cooperation rather than clonal selection. PMID:27830060

To "do the right thing" or to "just do it": locomotion and assessment as distinct self-regulatory imperatives.

PubMed

Kruglanski, A W; Thompson, E P; Higgins, E T; Atash, M N; Pierro, A; Shah, J Y; Spiegel, S

2000-11-01

An integrated series of studies investigated 2 functional dimensions of self-regulation referred to as assessment and locomotion (E. T. Higgins and A. W. Kruglanski, 1995). Assessment constitutes the comparative aspect of self-regulation that critically evaluates alternative goals or means to decide which are best to pursue and appraises performance. Locomotion constitutes the aspect of self-regulation concerned with movement from state to state, including commitment of psychological resources to initiate and maintain such movement. Two separate scales were developed to measure individual differences in these tendencies. Psychometric work attested to the scales' unidimensionality, internal consistency, and temporal stability. The authors found that (a) locomotion and assessment are relatively independent of each other, (b) both are needed for self-regulatory success, and (c) each relates to distinct task orientations and motivational emphases.

Mother-Infant Responsiveness: Timing, Mutual Regulation, and Interactional Context.

ERIC Educational Resources Information Center

Van Egeren, Laurie A,; Barratt, Marguerite S.; Roach, Mary A.

2001-01-01

Investigated from a dynamic systems perspective mutual regulation during naturalistic interaction of mothers with their 4-month-olds. Found that mothers and infants communicated primarily through vocal signals and responses. Levels of contingent responsiveness between partners were significantly associated and occurred within distinct behavioral…

Molecular Regulation of Endothelial Cells by NF-1

DTIC Science & Technology

2013-01-01

cancer progression. The mammalian target of rapamycin (mTOR) is a serine threonine kinase, that exists in two distinct signaling complexes: mTORC1 and...abnormalities such as diabetes , with known vascular complications. Thus mTOR may be a significant regulator of endothelial cell functions

An integrated view of suppressor T cell subsets in immunoregulation

PubMed Central

Jiang, Hong; Chess, Leonard

2004-01-01

The immune system evolved to protect organisms from a virtually infinite variety of disease-causing agents but to avoid harmful responses to self. Because immune protective mechanisms include the elaboration of potent inflammatory molecules, antibodies, and killer cell activation — which together can not only destroy invading microorganisms, pathogenic autoreactive cells, and tumors, but also mortally injure normal cells — the immune system is inherently a “double-edged sword” and must be tightly regulated. Immune response regulation includes homeostatic mechanisms intrinsic to the activation and differentiation of antigen-triggered immunocompetent cells and extrinsic mechanisms mediated by suppressor cells. This review series will focus on recent advances indicating that distinct subsets of regulatory CD4+ and CD8+ T cells as well as NK T cells control the outgrowth of potentially pathogenic antigen-reactive T cells and will highlight the evidence that these suppressor T cells may play potentially important clinical roles in preventing and treating immune-mediated disease. Here we provide a historical overview of suppressor cells and the experimental basis for the existence of functionally and phenotypically distinct suppressor subsets. Finally, we will speculate on how the distinct suppressor cell subsets may function in concert to regulate immune responses. PMID:15520848

Distinct requirements for TrkB and TrkC signaling in target innervation by sensory neurons

NASA Technical Reports Server (NTRS)

Postigo, Antonio; Calella, Anna Maria; Fritzsch, Bernd; Knipper, Marlies; Katz, David; Eilers, Andreas; Schimmang, Thomas; Lewin, Gary R.; Klein, Rudiger; Minichiello, Liliana

2002-01-01

Signaling by brain-derived neurotrophic factor (BDNF) via the TrkB receptor, or by neurotrophin-3 (NT3) through the TrkC receptor support distinct populations of sensory neurons. The intracellular signaling pathways activated by Trk (tyrosine kinase) receptors, which in vivo promote neuronal survival and target innervation, are not well understood. Using mice with TrkB or TrkC receptors lacking the docking site for Shc adaptors (trkB(shc/shc) and trkC(shc/shc) mice), we show that TrkB and TrkC promote survival of sensory neurons mainly through Shc site-independent pathways, suggesting that these receptors use similar pathways to prevent apoptosis. In contrast, the regulation of target innervation appears different: in trkB(shc/shc) mice neurons lose target innervation, whereas in trkC(shc/shc) mice the surviving TrkC-dependent neurons maintain target innervation and function. Biochemical analysis indicates that phosphorylation at the Shc site positively regulates autophosphorylation of TrkB, but not of TrkC. Our findings show that although TrkB and TrkC signals mediating survival are largely similar, TrkB and TrkC signals required for maintenance of target innervation in vivo are regulated by distinct mechanisms.

Distinct Protein Expression Profiles of Solid-Pseudopapillary Neoplasms of the Pancreas.

PubMed

Park, Minhee; Lim, Jong-Sun; Lee, Hyoung-Joo; Na, Keun; Lee, Min Jung; Kang, Chang Moo; Paik, Young-Ki; Kim, Hoguen

2015-08-07

Solid-pseudopapillary neoplasm (SPN) is an uncommon pancreatic tumor with mutation in CTNNB1 and distinct clinical and pathological features. We compared the proteomic profiles of SPN to mRNA expression. Pooled SPNs and pooled non-neoplastic pancreatic tissues were examined with high-resolution mass spectrometry. We identified 329 (150 up-regulated and 179 down-regulated) differentially expressed proteins in SPN. We identified 191 proteins (58.1% of the 329 dysregulated proteins) with the same expression tendencies in SPN based on mRNA data. Many overexpressed proteins were related to signaling pathways known to be activated in SPNs. We found that several proteins involved in Wnt signaling, including DKK4 and β-catenin, and proteins that bind β-catenin, such as FUS and NONO, were up-regulated in SPNs. Molecules involved in glycolysis, including PKM2, ENO2, and HK1, were overexpressed in accordance to their mRNA levels. In summary, SPN showed (1) distinct protein expression changes that correlated with mRNA expression, (2) overexpression of Wnt signaling proteins and proteins that bind directly to β-catenin, and (3) overexpression of proteins involved in metabolism. These findings may help develop early diagnostic biomarkers and molecular targets.

Cohesin Function in Cohesion, Condensation, and DNA Repair Is Regulated by Wpl1p via a Common Mechanism in Saccharomyces cerevisiae.

PubMed

Bloom, Michelle S; Koshland, Douglas; Guacci, Vincent

2018-01-01

Cohesin tethers DNA to mediate sister chromatid cohesion, chromosome condensation, and DNA repair. How the cell regulates cohesin to perform these distinct functions remains to be elucidated. One cohesin regulator, Wpl1p, was characterized in Saccharomyces cerevisiae as a promoter of efficient cohesion and an inhibitor of condensation. Wpl1p is also required for resistance to DNA-damaging agents. Here, we provide evidence that Wpl1p promotes the timely repair of DNA damage induced during S-phase. Previous studies have indicated that Wpl1p destabilizes cohesin's binding to DNA by modulating the interface between the cohesin subunits Mcd1p and Smc3p Our results suggest that Wpl1p likely modulates this interface to regulate all of cohesin's biological functions. Furthermore, we show that Wpl1p regulates cohesion and condensation through the formation of a functional complex with another cohesin-associated factor, Pds5p In contrast, Wpl1p regulates DNA repair independently of its interaction with Pds5p Together, these results suggest that Wpl1p regulates distinct biological functions of cohesin by Pds5p-dependent and -independent modulation of the Smc3p/Mcd1p interface. Copyright © 2018 by the Genetics Society of America.

Cohesin Function in Cohesion, Condensation, and DNA Repair Is Regulated by Wpl1p via a Common Mechanism in Saccharomyces cerevisiae

PubMed Central

Bloom, Michelle S.; Koshland, Douglas; Guacci, Vincent

2018-01-01

Cohesin tethers DNA to mediate sister chromatid cohesion, chromosome condensation, and DNA repair. How the cell regulates cohesin to perform these distinct functions remains to be elucidated. One cohesin regulator, Wpl1p, was characterized in Saccharomyces cerevisiae as a promoter of efficient cohesion and an inhibitor of condensation. Wpl1p is also required for resistance to DNA-damaging agents. Here, we provide evidence that Wpl1p promotes the timely repair of DNA damage induced during S-phase. Previous studies have indicated that Wpl1p destabilizes cohesin’s binding to DNA by modulating the interface between the cohesin subunits Mcd1p and Smc3p. Our results suggest that Wpl1p likely modulates this interface to regulate all of cohesin’s biological functions. Furthermore, we show that Wpl1p regulates cohesion and condensation through the formation of a functional complex with another cohesin-associated factor, Pds5p. In contrast, Wpl1p regulates DNA repair independently of its interaction with Pds5p. Together, these results suggest that Wpl1p regulates distinct biological functions of cohesin by Pds5p-dependent and -independent modulation of the Smc3p/Mcd1p interface. PMID:29158426

75 FR 52398 - Migratory Bird Hunting; Proposed Frameworks for Late-Season Migratory Bird Hunting Regulations

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-25

... late-season hunting regulations for certain migratory game birds. We annually prescribe frameworks, or... migratory game birds under Sec. Sec. 20.101 through 20.107, 20.109, and 20.110 of subpart K. Major steps in... game birds and developed recommendations for the 2010-11 regulations for these species plus regulations...

Modeling-independent elucidation of inactivation pathways in recombinant and native A-type Kv channels

PubMed Central

Fineberg, Jeffrey D.; Ritter, David M.

2012-01-01

A-type voltage-gated K+ (Kv) channels self-regulate their activity by inactivating directly from the open state (open-state inactivation [OSI]) or by inactivating before they open (closed-state inactivation [CSI]). To determine the inactivation pathways, it is often necessary to apply several pulse protocols, pore blockers, single-channel recording, and kinetic modeling. However, intrinsic hurdles may preclude the standardized application of these methods. Here, we implemented a simple method inspired by earlier studies of Na+ channels to analyze macroscopic inactivation and conclusively deduce the pathways of inactivation of recombinant and native A-type Kv channels. We investigated two distinct A-type Kv channels expressed heterologously (Kv3.4 and Kv4.2 with accessory subunits) and their native counterparts in dorsal root ganglion and cerebellar granule neurons. This approach applies two conventional pulse protocols to examine inactivation induced by (a) a simple step (single-pulse inactivation) and (b) a conditioning step (double-pulse inactivation). Consistent with OSI, the rate of Kv3.4 inactivation (i.e., the negative first derivative of double-pulse inactivation) precisely superimposes on the profile of the Kv3.4 current evoked by a single pulse because the channels must open to inactivate. In contrast, the rate of Kv4.2 inactivation is asynchronous, already changing at earlier times relative to the profile of the Kv4.2 current evoked by a single pulse. Thus, Kv4.2 inactivation occurs uncoupled from channel opening, indicating CSI. Furthermore, the inactivation time constant versus voltage relation of Kv3.4 decreases monotonically with depolarization and levels off, whereas that of Kv4.2 exhibits a J-shape profile. We also manipulated the inactivation phenotype by changing the subunit composition and show how CSI and CSI combined with OSI might affect spiking properties in a full computational model of the hippocampal CA1 neuron. This work unambiguously elucidates contrasting inactivation pathways in neuronal A-type Kv channels and demonstrates how distinct pathways might impact neurophysiological activity. PMID:23109714

43 CFR 17.322 - Assurance of compliance and recipient assessment of age distinctions.

Code of Federal Regulations, 2010 CFR

2010-10-01

... Nondiscrimination on the Basis of Age Duties of Doi Recipients § 17.322 Assurance of compliance and recipient assessment of age distinctions. (a) Each recipient of Federal financial assistance from DOI shall sign a written assurance as specified by DOI that it will comply with the Act and these regulations. (b...

43 CFR 17.322 - Assurance of compliance and recipient assessment of age distinctions.

Code of Federal Regulations, 2011 CFR

2011-10-01

... Nondiscrimination on the Basis of Age Duties of Doi Recipients § 17.322 Assurance of compliance and recipient assessment of age distinctions. (a) Each recipient of Federal financial assistance from DOI shall sign a written assurance as specified by DOI that it will comply with the Act and these regulations. (b...

A Tipping Point

ERIC Educational Resources Information Center

Heid, Susan D.

2007-01-01

It's near-impossible to think about course management systems (CMS) without thinking about innovation, collaboration, and the sharing of ideas across institutions and even from vendor to vendor. Yet, "the next step" in CMS now means distinctly different things to various colleges and universities as, going forward, they consider their…

On Euler's Theorem for Homogeneous Functions and Proofs Thereof.

ERIC Educational Resources Information Center

Tykodi, R. J.

1982-01-01

Euler's theorem for homogenous functions is useful when developing thermodynamic distinction between extensive and intensive variables of state and when deriving the Gibbs-Duhem relation. Discusses Euler's theorem and thermodynamic applications. Includes six-step instructional strategy for introducing the material to students. (Author/JN)

Studying Psychosocial Barriers to Drug Treatment Among Chinese Methamphetamine Users Using A 3-Step Latent Class Analysis.

PubMed

Wang, Jichuan; Kelly, Brian C; Liu, Tieqiao; Hao, Wei

2016-03-01

Given the growth in methamphetamine use in China during the 21st century, we assessed perceived psychosocial barriers to drug treatment among this population. Using a sample of 303 methamphetamine users recruited via Respondent Driven Sampling, we use Latent Class Analysis (LCA) to identify possible distinct latent groups among Chinese methamphetamine users on the basis of their perceptions of psychosocial barriers to drug treatment. After covariates were included to predict latent class membership, the 3-step modeling approach was applied. Our findings indicate that the Chinese methamphetamine using population was heterogeneous on perceptions of drug treatment barriers; four distinct latent classes (subpopulations) were identified--Unsupported Deniers, Deniers, Privacy Anxious, and Low Barriers--and individual characteristics shaped the probability of class membership. Efforts to link Chinese methamphetamine users to treatment may require a multi-faceted approach that attends to differing perceptions about impediments to drug treatment. Copyright © 2015. Published by Elsevier Inc.

Beyond fitness tracking: The use of consumer-grade wearable data from normal volunteers in cardiovascular and lipidomics research

PubMed Central

Teo, Jing Xian; Yang, Chengxi; Pua, Chee Jian; Blöcker, Christopher; Lim, Jing Quan; Ching, Jianhong; Yap, Jonathan Jiunn Liang; Tan, Swee Yaw; Sahlén, Anders; Chin, Calvin Woon-Loong; Teh, Bin Tean; Rozen, Steven G.; Cook, Stuart Alexander; Yeo, Khung Keong; Tan, Patrick

2018-01-01

The use of consumer-grade wearables for purposes beyond fitness tracking has not been comprehensively explored. We generated and analyzed multidimensional data from 233 normal volunteers, integrating wearable data, lifestyle questionnaires, cardiac imaging, sphingolipid profiling, and multiple clinical-grade cardiovascular and metabolic disease markers. We show that subjects can be stratified into distinct clusters based on daily activity patterns and that these clusters are marked by distinct demographic and behavioral patterns. While resting heart rates (RHRs) performed better than step counts in being associated with cardiovascular and metabolic disease markers, step counts identified relationships between physical activity and cardiac remodeling, suggesting that wearable data may play a role in reducing overdiagnosis of cardiac hypertrophy or dilatation in active individuals. Wearable-derived activity levels can be used to identify known and novel activity-modulated sphingolipids that are in turn associated with insulin sensitivity. Our findings demonstrate the potential for wearables in biomedical research and personalized health. PMID:29485983

Localization and roles of Ski8p protein in Sordaria meiosis and delineation of three mechanistically distinct steps of meiotic homolog juxtaposition.

PubMed

Tessé, Sophie; Storlazzi, Aurora; Kleckner, Nancy; Gargano, Silvana; Zickler, Denise

2003-10-28

Ski8p is implicated in degradation of non-poly(A) and double-stranded RNA, and in meiotic DNA recombination. We have identified the Sordaria macrospora SKI8 gene. Ski8p is cytoplasmically localized in all vegetative and sexual cycle cells, and is nuclear localized, specifically in early-mid-meiotic prophase, in temporal correlation with Spo11p, the meiotic double-strand break (DSB) transesterase. Localizations of Ski8p and Spo11p are mutually interdependent. ski8 mutants exhibit defects in vegetative growth, entry into the sexual program, and sporulation. Diverse meiotic defects, also seen in spo11 mutants, are diagnostic of DSB absence, and they are restored by exogenous DSBs. These results suggest that Ski8p promotes meiotic DSB formation by acting directly within meiotic prophase chromosomes. Mutant phenotypes also divide meiotic homolog juxtaposition into three successive, mechanistically distinct steps; recognition, presynaptic alignment, and synapsis, which are distinguished by their differential dependence on DSBs.

Localization and roles of Ski8p protein in Sordaria meiosis and delineation of three mechanistically distinct steps of meiotic homolog juxtaposition

PubMed Central

Tessé, Sophie; Storlazzi, Aurora; Kleckner, Nancy; Gargano, Silvana; Zickler, Denise

2003-01-01

Ski8p is implicated in degradation of non-poly(A) and double-stranded RNA, and in meiotic DNA recombination. We have identified the Sordaria macrospora SKI8 gene. Ski8p is cytoplasmically localized in all vegetative and sexual cycle cells, and is nuclear localized, specifically in early-mid-meiotic prophase, in temporal correlation with Spo11p, the meiotic double-strand break (DSB) transesterase. Localizations of Ski8p and Spo11p are mutually interdependent. ski8 mutants exhibit defects in vegetative growth, entry into the sexual program, and sporulation. Diverse meiotic defects, also seen in spo11 mutants, are diagnostic of DSB absence, and they are restored by exogenous DSBs. These results suggest that Ski8p promotes meiotic DSB formation by acting directly within meiotic prophase chromosomes. Mutant phenotypes also divide meiotic homolog juxtaposition into three successive, mechanistically distinct steps; recognition, presynaptic alignment, and synapsis, which are distinguished by their differential dependence on DSBs. PMID:14563920

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

PubMed Central

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

2016-01-01

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

Dynamic Cytology and Transcriptional Regulation of Rice Lamina Joint Development1[OPEN

PubMed Central

2017-01-01

Rice (Oryza sativa) leaf angle is determined by lamina joint and is an important agricultural trait determining leaf erectness and, hence, the photosynthesis efficiency and grain yield. Genetic studies reveal a complex regulatory network of lamina joint development; however, the morphological changes, cytological transitions, and underlying transcriptional programming remain to be elucidated. A systemic morphological and cytological study reveals a dynamic developmental process and suggests a common but distinct regulation of the lamina joint. Successive and sequential cell division and expansion, cell wall thickening, and programmed cell death at the adaxial or abaxial sides form the cytological basis of the lamina joint, and the increased leaf angle results from the asymmetric cell proliferation and elongation. Analysis of the gene expression profiles at four distinct developmental stages ranging from initiation to senescence showed that genes related to cell division and growth, hormone synthesis and signaling, transcription (transcription factors), and protein phosphorylation (protein kinases) exhibit distinct spatiotemporal patterns during lamina joint development. Phytohormones play crucial roles by promoting cell differentiation and growth at early stages or regulating the maturation and senescence at later stages, which is consistent with the quantitative analysis of hormones at different stages. Further comparison with the gene expression profile of leaf inclination1, a mutant with decreased auxin and increased leaf angle, indicates the coordinated effects of hormones in regulating lamina joint. These results reveal a dynamic cytology of rice lamina joint that is fine-regulated by multiple factors, providing informative clues for illustrating the regulatory mechanisms of leaf angle and plant architecture. PMID:28500269

Dynamic Cytology and Transcriptional Regulation of Rice Lamina Joint Development.

PubMed

Zhou, Li-Juan; Xiao, Lang-Tao; Xue, Hong-Wei

2017-07-01

Rice ( Oryza sativa ) leaf angle is determined by lamina joint and is an important agricultural trait determining leaf erectness and, hence, the photosynthesis efficiency and grain yield. Genetic studies reveal a complex regulatory network of lamina joint development; however, the morphological changes, cytological transitions, and underlying transcriptional programming remain to be elucidated. A systemic morphological and cytological study reveals a dynamic developmental process and suggests a common but distinct regulation of the lamina joint. Successive and sequential cell division and expansion, cell wall thickening, and programmed cell death at the adaxial or abaxial sides form the cytological basis of the lamina joint, and the increased leaf angle results from the asymmetric cell proliferation and elongation. Analysis of the gene expression profiles at four distinct developmental stages ranging from initiation to senescence showed that genes related to cell division and growth, hormone synthesis and signaling, transcription (transcription factors), and protein phosphorylation (protein kinases) exhibit distinct spatiotemporal patterns during lamina joint development. Phytohormones play crucial roles by promoting cell differentiation and growth at early stages or regulating the maturation and senescence at later stages, which is consistent with the quantitative analysis of hormones at different stages. Further comparison with the gene expression profile of leaf inclination1 , a mutant with decreased auxin and increased leaf angle, indicates the coordinated effects of hormones in regulating lamina joint. These results reveal a dynamic cytology of rice lamina joint that is fine-regulated by multiple factors, providing informative clues for illustrating the regulatory mechanisms of leaf angle and plant architecture. © 2017 American Society of Plant Biologists. All Rights Reserved.

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

PubMed

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

2016-06-15

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

MicroRNA dynamics in the stages of tumorigenesis correlate with hallmark capabilities of cancer.

PubMed

Olson, Peter; Lu, Jun; Zhang, Hao; Shai, Anny; Chun, Matthew G; Wang, Yucheng; Libutti, Steven K; Nakakura, Eric K; Golub, Todd R; Hanahan, Douglas

2009-09-15

While altered expression of microRNAs (miRs) in tumors has been well documented, it remains unclear how the miR transcriptome intersects neoplastic progression. By profiling the miR transcriptome we identified miR expression signatures associated with steps in tumorigenesis and the acquisition of hallmark capabilities in a prototypical mouse model of cancer. Metastases and a rare subset of primary tumors shared a distinct miR signature, implicating a discrete lineage for metastatic tumors. The miR-200 family is strongly down-regulated in metastases and met-like primary tumors, thereby relieving repression of the mesenchymal transcription factor Zeb1, which in turn suppresses E-cadherin. Treatment with a clinically approved angiogenesis inhibitor normalized angiogenic signature miRs in primary tumors, while altering expression of metastatic signature miRs similarly to liver metastases, suggesting their involvement in adaptive resistance to anti-angiogenic therapy via enhanced metastasis. Many of the miR changes associated with specific stages and hallmark capabilities in the mouse model are similarly altered in human tumors, including cognate pancreatic neuroendocrine tumors, implying a generality.

Functional divisions for visual processing in the central brain of flying Drosophila

PubMed Central

Weir, Peter T.; Dickinson, Michael H.

2015-01-01

Although anatomy is often the first step in assigning functions to neural structures, it is not always clear whether architecturally distinct regions of the brain correspond to operational units. Whereas neuroarchitecture remains relatively static, functional connectivity may change almost instantaneously according to behavioral context. We imaged panneuronal responses to visual stimuli in a highly conserved central brain region in the fruit fly, Drosophila, during flight. In one substructure, the fan-shaped body, automated analysis revealed three layers that were unresponsive in quiescent flies but became responsive to visual stimuli when the animal was flying. The responses of these regions to a broad suite of visual stimuli suggest that they are involved in the regulation of flight heading. To identify the cell types that underlie these responses, we imaged activity in sets of genetically defined neurons with arborizations in the targeted layers. The responses of this collection during flight also segregated into three sets, confirming the existence of three layers, and they collectively accounted for the panneuronal activity. Our results provide an atlas of flight-gated visual responses in a central brain circuit. PMID:26324910

Both endonucleolytic and exonucleolytic cleavage mediate ITS1 removal during human ribosomal RNA processing

PubMed Central

Sloan, Katherine E.; Mattijssen, Sandy; Lebaron, Simon; Tollervey, David; Pruijn, Ger J.M.

2013-01-01

Human ribosome production is up-regulated during tumorogenesis and is defective in many genetic diseases (ribosomopathies). We have undertaken a detailed analysis of human precursor ribosomal RNA (pre-rRNA) processing because surprisingly little is known about this important pathway. Processing in internal transcribed spacer 1 (ITS1) is a key step that separates the rRNA components of the large and small ribosomal subunits. We report that this was initiated by endonuclease cleavage, which required large subunit biogenesis factors. This was followed by 3′ to 5′ exonucleolytic processing by RRP6 and the exosome, an enzyme complex not previously linked to ITS1 removal. In contrast, RNA interference–mediated knockdown of the endoribonuclease MRP did not result in a clear defect in ITS1 processing. Despite the apparently high evolutionary conservation of the pre-rRNA processing pathway and ribosome synthesis factors, each of these features of human ITS1 processing is distinct from those in budding yeast. These results also provide significant insight into the links between ribosomopathies and ribosome production in human cells. PMID:23439679

Cofactor Editing by the G-protein Metallochaperone Domain Regulates the Radical B12 Enzyme IcmF.

PubMed

Li, Zhu; Kitanishi, Kenichi; Twahir, Umar T; Cracan, Valentin; Chapman, Derrell; Warncke, Kurt; Banerjee, Ruma

2017-03-10

IcmF is a 5'-deoxyadenosylcobalamin (AdoCbl)-dependent enzyme that catalyzes the carbon skeleton rearrangement of isobutyryl-CoA to butyryl-CoA. It is a bifunctional protein resulting from the fusion of a G-protein chaperone with GTPase activity and the cofactor- and substrate-binding mutase domains with isomerase activity. IcmF is prone to inactivation during catalytic turnover, thus setting up its dependence on a cofactor repair system. Herein, we demonstrate that the GTPase activity of IcmF powers the ejection of the inactive cob(II)alamin cofactor and requires the presence of an acceptor protein, adenosyltransferase, for receiving it. Adenosyltransferase in turn converts cob(II)alamin to AdoCbl in the presence of ATP and a reductant. The repaired cofactor is then reloaded onto IcmF in a GTPase-gated step. The mechanistic details of cofactor loading and offloading from the AdoCbl-dependent IcmF are distinct from those of the better characterized and homologous methylmalonyl-CoA mutase/G-protein chaperone system. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Specific Btk inhibition suppresses B cell- and myeloid cell-mediated arthritis.

PubMed

Di Paolo, Julie A; Huang, Tao; Balazs, Mercedesz; Barbosa, James; Barck, Kai H; Bravo, Brandon J; Carano, Richard A D; Darrow, James; Davies, Douglas R; DeForge, Laura E; Diehl, Lauri; Ferrando, Ronald; Gallion, Steven L; Giannetti, Anthony M; Gribling, Peter; Hurez, Vincent; Hymowitz, Sarah G; Jones, Randall; Kropf, Jeffrey E; Lee, Wyne P; Maciejewski, Patricia M; Mitchell, Scott A; Rong, Hong; Staker, Bart L; Whitney, J Andrew; Yeh, Sherry; Young, Wendy B; Yu, Christine; Zhang, Juan; Reif, Karin; Currie, Kevin S

2011-01-01

Bruton's tyrosine kinase (Btk) is a therapeutic target for rheumatoid arthritis, but the cellular and molecular mechanisms by which Btk mediates inflammation are poorly understood. Here we describe the discovery of CGI1746, a small-molecule Btk inhibitor chemotype with a new binding mode that stabilizes an inactive nonphosphorylated enzyme conformation. CGI1746 has exquisite selectivity for Btk and inhibits both auto- and transphosphorylation steps necessary for enzyme activation. Using CGI1746, we demonstrate that Btk regulates inflammatory arthritis by two distinct mechanisms. CGI1746 blocks B cell receptor-dependent B cell proliferation and in prophylactic regimens reduces autoantibody levels in collagen-induced arthritis. In macrophages, Btk inhibition abolishes FcγRIII-induced TNFα, IL-1β and IL-6 production. Accordingly, in myeloid- and FcγR-dependent autoantibody-induced arthritis, CGI1746 decreases cytokine levels within joints and ameliorates disease. These results provide new understanding of the function of Btk in both B cell- or myeloid cell-driven disease processes and provide a compelling rationale for targeting Btk in rheumatoid arthritis.

Induction of the neural crest state: Control of stem cell attributes by gene regulatory, post-transcriptional and epigenetic interactions

PubMed Central

Prasad, Maneeshi S.; Sauka-Spengler, Tatjana; LaBonne, Carole

2012-01-01

Neural crest cells are a population of multipotent stem cell-like progenitors that arise at the neural plate border in vertebrates, migrate extensively, and give rise to diverse derivatives such as melanocytes, craniofacial cartilage and bone, smooth muscle, peripheral and enteric neurons and glia. The neural crest gene regulatory network (NC-GRN) includes a number of key factors that are used reiteratively to control multiple steps in the development of neural crest cells, including the acquisition of stem cell attributes. It is therefore essential to understand the mechanisms that control the distinct functions of such reiteratively used factors in different cellular contexts. The context-dependent control of neural crest specification is achieved through combinatorial interaction with other factors, post-transcriptional and post-translational modifications, and the epigenetic status and chromatin state of target genes. Here we review the current understanding of the NC-GRN, including the role of the neural crest specifiers, their links to the control of “stemness,” and their dynamic context-dependent regulation during the formation of neural crest progenitors. PMID:22583479

Mechanisms of nuclear pore complex assembly - two different ways of building one molecular machine.

PubMed

Otsuka, Shotaro; Ellenberg, Jan

2018-02-01

The nuclear pore complex (NPC) mediates all macromolecular transport across the nuclear envelope. In higher eukaryotes that have an open mitosis, NPCs assemble at two points in the cell cycle: during nuclear assembly in late mitosis and during nuclear growth in interphase. How the NPC, the largest nonpolymeric protein complex in eukaryotic cells, self-assembles inside cells remained unclear. Recent studies have started to uncover the assembly process, and evidence has been accumulating that postmitotic and interphase NPC assembly use fundamentally different mechanisms; the duration, structural intermediates, and regulation by molecular players are different and different types of membrane deformation are involved. In this Review, we summarize the current understanding of these two modes of NPC assembly and discuss the structural and regulatory steps that might drive the assembly processes. We furthermore integrate understanding of NPC assembly with the mechanisms for rapid nuclear growth in embryos and, finally, speculate on the evolutionary origin of the NPC implied by the presence of two distinct assembly mechanisms. © 2017 The Authors. FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.

Specific Btk inhibition suppresses B cell- and myeloid cell-mediated arthritis

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

Di Paolo, Julie A; Huang, Tao; Balazs, Mercedesz

Bruton's tyrosine kinase (Btk) is a therapeutic target for rheumatoid arthritis, but the cellular and molecular mechanisms by which Btk mediates inflammation are poorly understood. Here we describe the discovery of CGI1746, a small-molecule Btk inhibitor chemotype with a new binding mode that stabilizes an inactive nonphosphorylated enzyme conformation. CGI1746 has exquisite selectivity for Btk and inhibits both auto- and transphosphorylation steps necessary for enzyme activation. Using CGI1746, we demonstrate that Btk regulates inflammatory arthritis by two distinct mechanisms. CGI1746 blocks B cell receptor–dependent B cell proliferation and in prophylactic regimens reduces autoantibody levels in collagen-induced arthritis. In macrophages, Btkmore » inhibition abolishes FcγRIII-induced TNFα, IL-1β and IL-6 production. Accordingly, in myeloid- and FcγR-dependent autoantibody-induced arthritis, CGI1746 decreases cytokine levels within joints and ameliorates disease. These results provide new understanding of the function of Btk in both B cell– or myeloid cell–driven disease processes and provide a compelling rationale for targeting Btk in rheumatoid arthritis.« less

Differential expression of syndecan isoforms during mouse incisor amelogenesis.

PubMed

Muto, Taro; Miyoshi, Keiko; Munesue, Seiichi; Nakada, Hiroshi; Okayama, Minoru; Matsuo, Takashi; Noma, Takafumi

2007-08-01

Syndecans are transmembranous heparan sulfate proteoglycans (HSPGs) with covalently attached glycosaminoglycan side-chains located on the cell surface. The mammalian syndecan family is composed of four types of syndecans (syndecan-1 to -4). Syndecans interact with the intracellular cytoskeleton through the cytoplasmic domains of their core proteins and membrane proteins, extracellular enzymes, growth factors, and matrix components, through their heparan-sulfate chains, to regulate developmental processes.Here, as a first step to assess the possible roles of syndecan proteins in amelogenesis, we examined the expression patterns of all syndecan isoforms in continuously growing mouse incisors, in which we can overview major differentiation stages of amelogenesis at a glance. Understanding the expression domain of each syndecan isoform during specific developmental stages seems useful for investigating their physiological roles in amelogenesis. Immunohistochemical analysis of syndecan core proteins in the lower incisors from postnatal day 1 mice revealed spatially and temporally specific expression patterns, with syndecan-1 expressed in undifferentiated epithelial and mesenchymal cells, and syndecan-2, -3, and -4 in more differentiated cells. These findings suggest that each syndecan isoform functions distinctly during the amelogenesis of the incisors of mice.

Gene expression profiling at early organogenesis reveals both common and diverse mechanisms in foregut patterning

PubMed Central

Fagman, Henrik; Amendola, Elena; Parrillo, Luca; Zoppoli, Pietro; Marotta, Pina; Scarfò, Marzia; De Luca, Pasquale; de Carvalho, Denise Pires; Ceccarelli, Michele; De Felice, Mario; Di Lauro, Roberto

2011-01-01

The thyroid and lungs originate as neighboring bud shaped outgrowths from the midline of the embryonic foregut. When and how organ specific programs regulate development into structures of distinct shapes, positions and functions is incompletely understood. To characterize, at least in part, the genetic basis of these events, we have employed laser capture microdissection and microarray analysis to define gene expression in the mouse thyroid and lung primordia at E10.5. By comparing the transcriptome of each bud to that of the whole embryo as well as to each other, we broadly describe the genes that are preferentially expressed in each developing organ as well as those with an enriched expression common to both. The results thus obtained provide a valuable resource for further analysis of genes previously unrecognized to participate in thyroid and lung morphogenesis and to discover organ specific as well as common developmental mechanisms. As an initial step in this direction we describe a regulatory pathway involving the anti-apoptotic gene Bcl2 that controls cell survival in early thyroid development. PMID:21924257

Restoring Ureagenesis in Hepatocytes by CRISPR/Cas9-mediated Genomic Addition to Arginase-deficient Induced Pluripotent Stem Cells.

PubMed

Lee, Patrick C; Truong, Brian; Vega-Crespo, Agustin; Gilmore, W Blake; Hermann, Kip; Angarita, Stephanie Ak; Tang, Jonathan K; Chang, Katherine M; Wininger, Austin E; Lam, Alex K; Schoenberg, Benjamen E; Cederbaum, Stephen D; Pyle, April D; Byrne, James A; Lipshutz, Gerald S

2016-11-29

Urea cycle disorders are incurable enzymopathies that affect nitrogen metabolism and typically lead to hyperammonemia. Arginase deficiency results from a mutation in Arg1, the enzyme regulating the final step of ureagenesis and typically results in developmental disabilities, seizures, spastic diplegia, and sometimes death. Current medical treatments for urea cycle disorders are only marginally effective, and for proximal disorders, liver transplantation is effective but limited by graft availability. Advances in human induced pluripotent stem cell research has allowed for the genetic modification of stem cells for potential cellular replacement therapies. In this study, we demonstrate a universally-applicable CRISPR/Cas9-based strategy utilizing exon 1 of the hypoxanthine-guanine phosphoribosyltransferase locus to genetically modify and restore arginase activity, and thus ureagenesis, in genetically distinct patient-specific human induced pluripotent stem cells and hepatocyte-like derivatives. Successful strategies restoring gene function in patient-specific human induced pluripotent stem cells may advance applications of genetically modified cell therapy to treat urea cycle and other inborn errors of metabolism.

Human Genomic Signatures of Brain Oscillations During Memory Encoding.

PubMed

Berto, Stefano; Wang, Guang-Zhong; Germi, James; Lega, Bradley C; Konopka, Genevieve

2018-05-01

Memory encoding is an essential step for all learning. However, the genetic and molecular mechanisms underlying human memory encoding remain poorly understood, and how this molecular framework permits the emergence of specific patterns of brain oscillations observed during mnemonic processing is unknown. Here, we directly compare intracranial electroencephalography recordings from the neocortex in individuals performing an episodic memory task with human gene expression from the same areas. We identify genes correlated with oscillatory memory effects across 6 frequency bands. These genes are enriched for autism-related genes and have preferential expression in neurons, in particular genes encoding synaptic proteins and ion channels, supporting the idea that the genes regulating voltage gradients are involved in the modulation of oscillatory patterns during successful memory encoding across brain areas. Memory-related genes are distinct from those correlated with other forms of cognitive processing and resting state fMRI. These data are the first to identify correlations between gene expression and active human brain states as well as provide a molecular window into memory encoding oscillations in the human brain.

Functional divisions for visual processing in the central brain of flying Drosophila.

PubMed

Weir, Peter T; Dickinson, Michael H

2015-10-06

Although anatomy is often the first step in assigning functions to neural structures, it is not always clear whether architecturally distinct regions of the brain correspond to operational units. Whereas neuroarchitecture remains relatively static, functional connectivity may change almost instantaneously according to behavioral context. We imaged panneuronal responses to visual stimuli in a highly conserved central brain region in the fruit fly, Drosophila, during flight. In one substructure, the fan-shaped body, automated analysis revealed three layers that were unresponsive in quiescent flies but became responsive to visual stimuli when the animal was flying. The responses of these regions to a broad suite of visual stimuli suggest that they are involved in the regulation of flight heading. To identify the cell types that underlie these responses, we imaged activity in sets of genetically defined neurons with arborizations in the targeted layers. The responses of this collection during flight also segregated into three sets, confirming the existence of three layers, and they collectively accounted for the panneuronal activity. Our results provide an atlas of flight-gated visual responses in a central brain circuit.

Specific Btk inhibition suppresses B cell- and myeloid cell-mediated arthritis

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

Di Paolo, Julie A.; Huang, Tao; Balazs, Mercedesz

Bruton's tyrosine kinase (Btk) is a therapeutic target for rheumatoid arthritis, but the cellular and molecular mechanisms by which Btk mediates inflammation are poorly understood. Here we describe the discovery of CGI1746, a small-molecule Btk inhibitor chemotype with a new binding mode that stabilizes an inactive nonphosphorylated enzyme conformation. CGI1746 has exquisite selectivity for Btk and inhibits both auto- and transphosphorylation steps necessary for enzyme activation. Using CGI1746, we demonstrate that Btk regulates inflammatory arthritis by two distinct mechanisms. CGI1746 blocks B cell receptor-dependent B cell proliferation and in prophylactic regimens reduces autoantibody levels in collagen-induced arthritis. In macrophages, Btkmore » inhibition abolishes Fc{gamma}RIII-induced TNF{alpha}, IL-1{beta} and IL-6 production. Accordingly, in myeloid- and Fc{gamma}R-dependent autoantibody-induced arthritis, CGI1746 decreases cytokine levels within joints and ameliorates disease. These results provide new understanding of the function of Btk in both B cell- or myeloid cell-driven disease processes and provide a compelling rationale for targeting Btk in rheumatoid arthritis.« less

Synaptic UNC13A protein variant causes increased neurotransmission and dyskinetic movement disorder.

PubMed

Lipstein, Noa; Verhoeven-Duif, Nanda M; Michelassi, Francesco E; Calloway, Nathaniel; van Hasselt, Peter M; Pienkowska, Katarzyna; van Haaften, Gijs; van Haelst, Mieke M; van Empelen, Ron; Cuppen, Inge; van Teeseling, Heleen C; Evelein, Annemieke M V; Vorstman, Jacob A; Thoms, Sven; Jahn, Olaf; Duran, Karen J; Monroe, Glen R; Ryan, Timothy A; Taschenberger, Holger; Dittman, Jeremy S; Rhee, Jeong-Seop; Visser, Gepke; Jans, Judith J; Brose, Nils

2017-03-01

Munc13 proteins are essential regulators of neurotransmitter release at nerve cell synapses. They mediate the priming step that renders synaptic vesicles fusion-competent, and their genetic elimination causes a complete block of synaptic transmission. Here we have described a patient displaying a disorder characterized by a dyskinetic movement disorder, developmental delay, and autism. Using whole-exome sequencing, we have shown that this condition is associated with a rare, de novo Pro814Leu variant in the major human Munc13 paralog UNC13A (also known as Munc13-1). Electrophysiological studies in murine neuronal cultures and functional analyses in Caenorhabditis elegans revealed that the UNC13A variant causes a distinct dominant gain of function that is characterized by increased fusion propensity of synaptic vesicles, which leads to increased initial synaptic vesicle release probability and abnormal short-term synaptic plasticity. Our study underscores the critical importance of fine-tuned presynaptic control in normal brain function. Further, it adds the neuronal Munc13 proteins and the synaptic vesicle priming process that they control to the known etiological mechanisms of psychiatric and neurological synaptopathies.

Synaptic UNC13A protein variant causes increased neurotransmission and dyskinetic movement disorder

PubMed Central

Lipstein, Noa; Verhoeven-Duif, Nanda M.; Calloway, Nathaniel; van Hasselt, Peter M.; Pienkowska, Katarzyna; van Haelst, Mieke M.; van Empelen, Ron; Cuppen, Inge; van Teeseling, Heleen C.; Evelein, Annemieke M.V.; Vorstman, Jacob A.; Jahn, Olaf; Duran, Karen J.; Monroe, Glen R.; Ryan, Timothy A.; Taschenberger, Holger; Rhee, Jeong-Seop; Visser, Gepke; Jans, Judith J.

2017-01-01

Munc13 proteins are essential regulators of neurotransmitter release at nerve cell synapses. They mediate the priming step that renders synaptic vesicles fusion-competent, and their genetic elimination causes a complete block of synaptic transmission. Here we have described a patient displaying a disorder characterized by a dyskinetic movement disorder, developmental delay, and autism. Using whole-exome sequencing, we have shown that this condition is associated with a rare, de novo Pro814Leu variant in the major human Munc13 paralog UNC13A (also known as Munc13-1). Electrophysiological studies in murine neuronal cultures and functional analyses in Caenorhabditis elegans revealed that the UNC13A variant causes a distinct dominant gain of function that is characterized by increased fusion propensity of synaptic vesicles, which leads to increased initial synaptic vesicle release probability and abnormal short-term synaptic plasticity. Our study underscores the critical importance of fine-tuned presynaptic control in normal brain function. Further, it adds the neuronal Munc13 proteins and the synaptic vesicle priming process that they control to the known etiological mechanisms of psychiatric and neurological synaptopathies. PMID:28192369

A Rab5 GTPase module is important for autophagosome closure

PubMed Central

Lipatova, Zhanna; Sun, Dan; Zhu, Xiaolong; Li, Rui; Wu, Zulin; You, Weiming; Cong, Xiaoxia; Zhou, Yiting; Gyurkovska, Valeriya; Liu, Yutao; Li, Qunli; Li, Wenjing; Cheng, Jie; Segev, Nava

2017-01-01

In the conserved autophagy pathway, the double-membrane autophagosome (AP) engulfs cellular components to be delivered for degradation in the lysosome. While only sealed AP can productively fuse with the lysosome, the molecular mechanism of AP closure is currently unknown. Rab GTPases, which regulate all intracellular trafficking pathways in eukaryotes, also regulate autophagy. Rabs function in GTPase modules together with their activators and downstream effectors. In yeast, an autophagy-specific Ypt1 GTPase module, together with a set of autophagy-related proteins (Atgs) and a phosphatidylinositol-3-phosphate (PI3P) kinase, regulates AP formation. Fusion of APs and endosomes with the vacuole (the yeast lysosome) requires the Ypt7 GTPase module. We have previously shown that the Rab5-related Vps21, within its endocytic GTPase module, regulates autophagy. However, it was not clear which autophagy step it regulates. Here, we show that this module, which includes the Vps9 activator, the Rab5-related Vps21, the CORVET tethering complex, and the Pep12 SNARE, functions after AP expansion and before AP closure. Whereas APs are not formed in mutant cells depleted for Atgs, sealed APs accumulate in cells depleted for the Ypt7 GTPase module members. Importantly, depletion of individual members of the Vps21 module results in a novel phenotype: accumulation of unsealed APs. In addition, we show that Vps21-regulated AP closure precedes another AP maturation step, the previously reported PI3P phosphatase-dependent Atg dissociation. Our results delineate three successive steps in the autophagy pathway regulated by Rabs, Ypt1, Vps21 and Ypt7, and provide the first insight into the upstream regulation of AP closure. PMID:28934205

Homo sapiens exhibit a distinct pattern of CNV genes regulation: an important role of miRNAs and SNPs in expression plasticity.

PubMed

Dweep, Harsh; Kubikova, Nada; Gretz, Norbert; Voskarides, Konstantinos; Felekkis, Kyriacos

2015-07-16

Gene expression regulation is a complex and highly organized process involving a variety of genomic factors. It is widely accepted that differences in gene expression can contribute to the phenotypic variability between species, and that their interpretation can aid in the understanding of the physiologic variability. CNVs and miRNAs are two major players in the regulation of expression plasticity and may be responsible for the unique phenotypic characteristics observed in different lineages. We have previously demonstrated that a close interaction between these two genomic elements may have contributed to the regulation of gene expression during evolution. This work presents the molecular interactions between CNV and non CNV genes with miRNAs and other genomic elements in eight different species. A comprehensive analysis of these interactions indicates a unique nature of human CNV genes regulation as compared to other species. By using genes with short 3' UTR that abolish the "canonical" miRNA-dependent regulation, as a model, we demonstrate a distinct and tight regulation of human genes that might explain some of the unique features of human physiology. In addition, comparison of gene expression regulation between species indicated that there is a significant difference between humans and mice possibly questioning the effectiveness of the latest as experimental models of human diseases.

Homo sapiens exhibit a distinct pattern of CNV genes regulation: an important role of miRNAs and SNPs in expression plasticity

PubMed Central

Dweep, Harsh; Kubikova, Nada; Gretz, Norbert; Voskarides, Konstantinos; Felekkis, Kyriacos

2015-01-01

Gene expression regulation is a complex and highly organized process involving a variety of genomic factors. It is widely accepted that differences in gene expression can contribute to the phenotypic variability between species, and that their interpretation can aid in the understanding of the physiologic variability. CNVs and miRNAs are two major players in the regulation of expression plasticity and may be responsible for the unique phenotypic characteristics observed in different lineages. We have previously demonstrated that a close interaction between these two genomic elements may have contributed to the regulation of gene expression during evolution. This work presents the molecular interactions between CNV and non CNV genes with miRNAs and other genomic elements in eight different species. A comprehensive analysis of these interactions indicates a unique nature of human CNV genes regulation as compared to other species. By using genes with short 3′ UTR that abolish the “canonical” miRNA-dependent regulation, as a model, we demonstrate a distinct and tight regulation of human genes that might explain some of the unique features of human physiology. In addition, comparison of gene expression regulation between species indicated that there is a significant difference between humans and mice possibly questioning the effectiveness of the latest as experimental models of human diseases. PMID:26178010

Cell cycle arrest in plants: what distinguishes quiescence, dormancy and differentiated G1?

PubMed

Velappan, Yazhini; Signorelli, Santiago; Considine, Michael J

2017-10-17

Quiescence is a fundamental feature of plant life, which enables plasticity, renewal and fidelity of the somatic cell line. Cellular quiescence is defined by arrest in a particular phase of the cell cycle, typically G1 or G2; however, the regulation of quiescence and proliferation can also be considered across wider scales in space and time. As such, quiescence is a defining feature of plant development and phenology, from meristematic stem cell progenitors to terminally differentiated cells, as well as dormant or suppressed seeds and buds. While the physiology of each of these states differs considerably, each is referred to as 'cell cycle arrest' or 'G1 arrest'. Here the physiology and molecular regulation of (1) meristematic quiescence, (2) dormancy and (3) terminal differentiation (cell cycle exit) are considered in order to determine whether and how the molecular decisions guiding these nuclear states are distinct. A brief overview of the canonical cell cycle regulators is provided, and the genetic and genomic, as well as physiological, evidence is considered regarding two primary questions: (1) Are the canonical cell cycle regulators superior or subordinate in the regulation of quiescence? (2) Are these three modes of quiescence governed by distinct molecular controls? Meristematic quiescence, dormancy and terminal differentiation are each predominantly characterized by G1 arrest but regulated distinctly, at a level largely superior to the canonical cell cycle. Meristematic quiescence is intrinsically linked to non-cell-autonomous regulation of meristem cell identity, and particularly through the influence of ubiquitin-dependent proteolysis, in partnership with reactive oxygen species, abscisic acid and auxin. The regulation of terminal differentiation shares analogous features with meristematic quiescence, albeit with specific activators and a greater role for cytokinin signalling. Dormancy meanwhile appears to be regulated at the level of chromatin accessibility, by Polycomb group-type histone modifications of particular dormancy genes. © The Author 2017. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Combating omission errors through task analysis and good reminders.

PubMed

Reason, J

2002-03-01

Leaving out necessary task steps is the single most common human error type. Certain task steps possess characteristics that are more likely to provoke omissions than others, and can be identified in advance. The paper reports two studies. The first, involving a simple photocopier, established that failing to remove the last page of the original is the commonest omission. This step possesses four distinct error-provoking features that combine their effects in an additive fashion. The second study examined the degree to which everyday memory aids satisfy five features of a good reminder: conspicuity, contiguity, content, context, and countability. A close correspondence was found between the percentage use of strategies and the degree to which they satisfied these five criteria. A three stage omission management programme was outlined: task analysis (identifying discrete task steps) of some safety critical activity; assessing the omission likelihood of each step; and the choice and application of a suitable reminder. Such a programme is applicable to a variety of healthcare procedures.

Functional analysis of the interactions between reovirus particles and various proteases in vitro

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

Sargent, M.D.; Long, D.G.; Borsa, J.

1977-01-01

The digestion of purified reovirus particles by various proteases including chymotrypsin, trypsin, pronase, papain, bromelain, proteinase K, and fibrinolysin has been examined as it relates to virion transcriptase activation and alteration of infectivity. In every case uncoating to the level of active transcriptase proceeds via two mechanistically distinct steps. All the proteases tested serve to mediate only the first of the two steps, converting intact virions to intermediate subviral particles (ISVP) in which the transcriptase is retained in a latent state. The second step of the uncoating process is mediated by a K/sup +/ ion-triggered, endogenous mechanism and results inmore » conversion of ISVP to cores, concomitant with transcriptase activation and loss of infectivity. All of the tested enzymes, except trypsin, reversibly block the second step of uncoating. These results indicate the generality, with respect to protease employed, of the two-step process for reovirus uncoating and transcriptase activation demonstrated previously with chymotrypsin.« less

Measurements of gluconeogenesis and glycogenolysis: A methodological review

USDA-ARS?s Scientific Manuscript database

Gluconeogenesis is a complex metabolic process that involves multiple enzymatic steps regulated by myriad factors, including substrate concentrations, the redox state, activation and inhibition of specific enzyme steps, and hormonal modulation. At present, the most widely accepted technique to deter...

Does my step look big in this? A visual illusion leads to safer stepping behaviour.

PubMed

Elliott, David B; Vale, Anna; Whitaker, David; Buckley, John G

2009-01-01

Tripping is a common factor in falls and a typical safety strategy to avoid tripping on steps or stairs is to increase foot clearance over the step edge. In the present study we asked whether the perceived height of a step could be increased using a visual illusion and whether this would lead to the adoption of a safer stepping strategy, in terms of greater foot clearance over the step edge. The study also addressed the controversial question of whether motor actions are dissociated from visual perception. 21 young, healthy subjects perceived the step to be higher in a configuration of the horizontal-vertical illusion compared to a reverse configuration (p = 0.01). During a simple stepping task, maximum toe elevation changed by an amount corresponding to the size of the visual illusion (p<0.001). Linear regression analyses showed highly significant associations between perceived step height and maximum toe elevation for all conditions. The perceived height of a step can be manipulated using a simple visual illusion, leading to the adoption of a safer stepping strategy in terms of greater foot clearance over a step edge. In addition, the strong link found between perception of a visual illusion and visuomotor action provides additional support to the view that the original, controversial proposal by Goodale and Milner (1992) of two separate and distinct visual streams for perception and visuomotor action should be re-evaluated.

Butyrate induced IGF2 activation correlated with distinct chromatin landscapes due to histone modification

USDA-ARS?s Scientific Manuscript database

Histone modification has emerged as a very important mechanism regulating the transcriptional status of the genome. Insulin-like growth factor 2 (IGF2) is a peptide hormone controlling various cellular processes such as proliferation and apoptosis. IGF2 and H19 are reciprocally regulated imprinted ...

Examining Hypermedia Learning: The Role of Cognitive Load and Self-Regulated Learning

ERIC Educational Resources Information Center

Moos, Daniel

2013-01-01

Distinct theoretical perspectives, Cognitive Load Theory and Self-Regulated Learning (SRL) theory, have been used to examine individual differences the challenges faced with hypermedia learning. However, research has tended to use these theories independently, resulting in less robust explanations of hypermedia learning. This study examined the…

Homework Emotion Regulation Scale: Psychometric Properties for Middle School Students

ERIC Educational Resources Information Center

Xu, Jianzhong; Fan, Xitao; Du, Jianxia

2016-01-01

The goal of the present investigation is to evaluate the psychometric properties of the Homework Emotion Regulation Scale (HERS) using 796 middle school students in China. Confirmatory factor analyses (CFAs) supported the existence of two distinct yet related subscales for the HERS: Emotion Management and Cognitive Reappraisal. Concerning the…

Emotion Regulation in Mathematics Homework: An Empirical Study

ERIC Educational Resources Information Center

Xu, Jianzhong

2018-01-01

The author examined 2 distinctive aspects of emotion regulation in mathematics homework, including emotion management and cognitive reappraisal. Participants were 1,799 high school students from 46 classes in China. Two multilevel models were run, 1 with emotion management and another with cognitive reappraisal as the dependent variable. Both…

Multiple roles for the actin cytoskeleton during regulated exocytosis

PubMed Central

Porat-Shliom, Natalie; Milberg, Oleg; Masedunskas, Andrius; Weigert, Roberto

2014-01-01

Regulated exocytosis is the main mechanism utilized by specialized secretory cells to deliver molecules to the cell surface by virtue of membranous containers (i.e. secretory vesicles). The process involves a series of highly coordinated and sequential steps, which include the biogenesis of the vesicles, their delivery to the cell periphery, their fusion with the plasma membrane and the release of their content into the extracellular space. Each of these steps is regulated by the actin cytoskeleton. In this review, we summarize the current knowledge regarding the involvement of actin and its associated molecules during each of the exocytic steps in vertebrates, and suggest that the overall role of the actin cytoskeleton during regulated exocytosis is linked to the architecture and the physiology of the secretory cells under examination. Specifically, in neurons, neuroendocrine, endocrine, and hematopoietic cells, which contain small secretory vesicles that undergo rapid exocytosis (on the order of milliseconds), the actin cytoskeleton plays a role in pre-fusion events, where it acts primarily as a functional barrier and facilitates docking. In exocrine and other secretory cells, which contain large secretory vesicles that undergo slow exocytosis (seconds to minutes), the actin cytoskeleton plays a role in post-fusion events, where it regulates the dynamics of the fusion pore, facilitates the integration of the vesicles into the plasma membrane, provides structural support, and promotes the expulsion of large cargo molecules. PMID:22986507

Regulating RNA polymerase pausing and transcription elongation in embryonic stem cells

PubMed Central

Min, Irene M.; Waterfall, Joshua J.; Core, Leighton J.; Munroe, Robert J.; Schimenti, John; Lis, John T.

2011-01-01

Transitions between pluripotent stem cells and differentiated cells are executed by key transcription regulators. Comparative measurements of RNA polymerase distribution over the genome's primary transcription units in different cell states can identify the genes and steps in the transcription cycle that are regulated during such transitions. To identify the complete transcriptional profiles of RNA polymerases with high sensitivity and resolution, as well as the critical regulated steps upon which regulatory factors act, we used genome-wide nuclear run-on (GRO-seq) to map the density and orientation of transcriptionally engaged RNA polymerases in mouse embryonic stem cells (ESCs) and mouse embryonic fibroblasts (MEFs). In both cell types, progression of a promoter-proximal, paused RNA polymerase II (Pol II) into productive elongation is a rate-limiting step in transcription of ∼40% of mRNA-encoding genes. Importantly, quantitative comparisons between cell types reveal that transcription is controlled frequently at paused Pol II's entry into elongation. Furthermore, “bivalent” ESC genes (exhibiting both active and repressive histone modifications) bound by Polycomb group complexes PRC1 (Polycomb-repressive complex 1) and PRC2 show dramatically reduced levels of paused Pol II at promoters relative to an average gene. In contrast, bivalent promoters bound by only PRC2 allow Pol II pausing, but it is confined to extremely 5′ proximal regions. Altogether, these findings identify rate-limiting targets for transcription regulation during cell differentiation. PMID:21460038

Regulation and competition without privatization: Norway`s experience

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

Moen, J.; Hamrin, J.

The competitive market for the hydro-based Norwegian electricity system is working well, with end-user prices only slightly above the wholesale market. Pool prices are reflecting only weather-related variations, and no market power abuses are evident. The challenge now is to restructure ownership of the wires and retail suppliers to lower wheeling costs and avoid cross-subsidization. Since the Norwegian Energy Act came into effect in 1991, the electricity industry in Norway has operated as one of the most deregulated electricity industries in the world. The Energy Act introduced third party access to the retail market and competition in electricity production. Themore » generation, sale and purchase of electricity is now highly competitive, with customers free to buy electricity from any generator, trader or the electricity Pool. Transmission pricing was separated from power purchasing arrangements, so that the buying and selling of electricity as a product is distinct from the transmission of electricity as a service. Transmission and distribution networks continue to maintain natural monopolies, with network owners providing wheeling service across their networks to customers who are connected to them. These monopoly sectors of the industry are subject to regulation by the government-appointed regulatory body, Norwegian Water Resources and Energy Administration (NVE). Regulation is on a cost-of-service basis, with the revenue allowance determined by NVE. The main force behind the Norwegian reform was the desire for efficiency gains to be achieved through a total restructure of the commercial character of the energy service industry (ESI). Unlike the U.K., in Norway the monopoly franchise for both generation and retail supply was removed in one step without any transition period, and the old pool was reformed to provide the needed structure for this new competitive energy market.« less

Mechanisms and Regulation of Alternative Pre-mRNA Splicing

PubMed Central

Lee, Yeon

2015-01-01

Precursor messenger RNA (pre-mRNA) splicing is a critical step in the posttranscriptional regulation of gene expression, providing significant expansion of the functional proteome of eukaryotic organisms with limited gene numbers. Split eukaryotic genes contain intervening sequences or introns disrupting protein-coding exons, and intron removal occurs by repeated assembly of a large and highly dynamic ribonucleoprotein complex termed the spliceosome, which is composed of five small nuclear ribonucleoprotein particles, U1, U2, U4/U6, and U5. Biochemical studies over the past 10 years have allowed the isolation as well as compositional, functional, and structural analysis of splicing complexes at distinct stages along the spliceosome cycle. The average human gene contains eight exons and seven introns, producing an average of three or more alternatively spliced mRNA isoforms. Recent high-throughput sequencing studies indicate that 100% of human genes produce at least two alternative mRNA isoforms. Mechanisms of alternative splicing include RNA–protein interactions of splicing factors with regulatory sites termed silencers or enhancers, RNA–RNA base-pairing interactions, or chromatin-based effects that can change or determine splicing patterns. Disease-causing mutations can often occur in splice sites near intron borders or in exonic or intronic RNA regulatory silencer or enhancer elements, as well as in genes that encode splicing factors. Together, these studies provide mechanistic insights into how spliceosome assembly, dynamics, and catalysis occur; how alternative splicing is regulated and evolves; and how splicing can be disrupted by cis- and trans-acting mutations leading to disease states. These findings make the spliceosome an attractive new target for small-molecule, antisense, and genome-editing therapeutic interventions. PMID:25784052

EBI1/CCR7 is a new member of dendritic cell chemokine receptor that is up-regulated upon maturation.

PubMed

Yanagihara, S; Komura, E; Nagafune, J; Watarai, H; Yamaguchi, Y

1998-09-15

Dendritic cells (DC) that are stimulated with inflammatory mediators can maturate and migrate from nonlymphoid tissues to lymphoid organs to initiate T cell-mediated immune responses. This migratory step is closely related to the maturation of the DC. In an attempt to identify chemokine receptors that might influence migration and are selectively expressed in mature DC, we have discovered that the chemokine receptor, EBI1/CCR7, is strikingly up-regulated upon maturation in three distinct culture systems: 1) mouse bone marrow-derived DC, 2) mouse epidermal Langerhans cells, and 3) human monocyte-derived DC. The EBI1/CCR7 expressed in mature DC is functional because ELC/MIP-3beta, recently identified as a ligand of EBI1/CCR7, induces a rise in intracellular free calcium concentrations and directional migration of human monocyte-derived mature DC (HLA-DRhigh, CD1a(low), CD14-, CD25+, CD83+, and CD86high) in a dose-dependent manner, but not of immature DC (HLA-DRlow, CD1a(high), CD14-, CD25-, CD83-, and CD86-). In contrast, macrophage inflammatory protein-1alpha (MIP-1alpha), monocyte chemotactic protein-3 (MCP-3), and RANTES are active on immature DC but not on mature DC. Thus, it seems likely that MIP-1alpha, MCP-3, and RANTES can mediate the migration of immature DC located in peripheral sites, whereas ELC/MIP-3beta can direct the migration of Ag-carrying DC from peripheral inflammatory sites, where DC are stimulated to up-regulate the expression of EBI1/CCR7, to lymphoid organs. It is postulated that different chemokines and chemokine receptors are involved in DC migration in vivo, depending on the maturation state of DC.

Ca2+-dependent dephosphorylation of kinesin heavy chain on beta-granules in pancreatic beta-cells. Implications for regulated beta-granule transport and insulin exocytosis

NASA Technical Reports Server (NTRS)

Donelan, Matthew J.; Morfini, Gerardo; Julyan, Richard; Sommers, Scott; Hays, Lori; Kajio, Hiroshi; Briaud, Isabelle; Easom, Richard A.; Molkentin, Jeffery D.; Brady, Scott T.;

The structure of bradyzoite-specific enolase from Toxoplasma gondii reveals insights into its dual cytoplasmic and nuclear functions

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

Ruan, Jiapeng; Mouveaux, Thomas; Light, Samuel H.

2015-03-01

The second crystal structure of a parasite protein preferentially enriched in the brain cyst of T. gondii has been solved at 2.75 Å resolution. Bradyzoite enolase 1 is reported to have differential functions as a glycolytic enzyme and a transcriptional regulator in bradyzoites. In addition to catalyzing a central step in glycolysis, enolase assumes a remarkably diverse set of secondary functions in different organisms, including transcription regulation as documented for the oncogene c-Myc promoter-binding protein 1. The apicomplexan parasite Toxoplasma gondii differentially expresses two nuclear-localized, plant-like enolases: enolase 1 (TgENO1) in the latent bradyzoite cyst stage and enolase 2 (TgENO2)more » in the rapidly replicative tachyzoite stage. A 2.75 Å resolution crystal structure of bradyzoite enolase 1, the second structure to be reported of a bradyzoite-specific protein in Toxoplasma, captures an open conformational state and reveals that distinctive plant-like insertions are located on surface loops. The enolase 1 structure reveals that a unique residue, Glu164, in catalytic loop 2 may account for the lower activity of this cyst-stage isozyme. Recombinant TgENO1 specifically binds to a TTTTCT DNA motif present in the cyst matrix antigen 1 (TgMAG1) gene promoter as demonstrated by gel retardation. Furthermore, direct physical interactions of both nuclear TgENO1 and TgENO2 with the TgMAG1 gene promoter are demonstrated in vivo using chromatin immunoprecipitation (ChIP) assays. Structural and biochemical studies reveal that T. gondii enolase functions are multifaceted, including the coordination of gene regulation in parasitic stage development. Enolase 1 provides a potential lead in the design of drugs against Toxoplasma brain cysts.« less

Genome-Wide Temporal Expression Profiling in Caenorhabditis elegans Identifies a Core Gene Set Related to Long-Term Memory.

PubMed

Freytag, Virginie; Probst, Sabine; Hadziselimovic, Nils; Boglari, Csaba; Hauser, Yannick; Peter, Fabian; Gabor Fenyves, Bank; Milnik, Annette; Demougin, Philippe; Vukojevic, Vanja; de Quervain, Dominique J-F; Papassotiropoulos, Andreas; Stetak, Attila

2017-07-12

The identification of genes related to encoding, storage, and retrieval of memories is a major interest in neuroscience. In the current study, we analyzed the temporal gene expression changes in a neuronal mRNA pool during an olfactory long-term associative memory (LTAM) in Caenorhabditis elegans hermaphrodites. Here, we identified a core set of 712 (538 upregulated and 174 downregulated) genes that follows three distinct temporal peaks demonstrating multiple gene regulation waves in LTAM. Compared with the previously published positive LTAM gene set (Lakhina et al., 2015), 50% of the identified upregulated genes here overlap with the previous dataset, possibly representing stimulus-independent memory-related genes. On the other hand, the remaining genes were not previously identified in positive associative memory and may specifically regulate aversive LTAM. Our results suggest a multistep gene activation process during the formation and retrieval of long-term memory and define general memory-implicated genes as well as conditioning-type-dependent gene sets. SIGNIFICANCE STATEMENT The identification of genes regulating different steps of memory is of major interest in neuroscience. Identification of common memory genes across different learning paradigms and the temporal activation of the genes are poorly studied. Here, we investigated the temporal aspects of Caenorhabditis elegans gene expression changes using aversive olfactory associative long-term memory (LTAM) and identified three major gene activation waves. Like in previous studies, aversive LTAM is also CREB dependent, and CREB activity is necessary immediately after training. Finally, we define a list of memory paradigm-independent core gene sets as well as conditioning-dependent genes. Copyright © 2017 the authors 0270-6474/17/376661-12$15.00/0.

Coregulation of Terpenoid Pathway Genes and Prediction of Isoprene Production in Bacillus subtilis Using Transcriptomics.

PubMed

Hess, Becky M; Xue, Junfeng; Markillie, Lye Meng; Taylor, Ronald C; Wiley, H Steven; Ahring, Birgitte K; Linggi, Bryan

2013-01-01

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes, as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding system level regulation and control of the pathway. To address these limitations, we examined Bacillus subtilis grown under multiple conditions and determined the relationship between altered isoprene production and gene expression patterns. We found that with respect to the amount of isoprene produced, terpenoid genes fall into two distinct subsets with opposing correlations. The group whose expression levels positively correlated with isoprene production included dxs, which is responsible for the commitment step in the pathway, ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome-wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. These analyses showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model that accurately predicts production of this secondary metabolite across many simulated environmental conditions.

Coregulation of Terpenoid Pathway Genes and Prediction of Isoprene Production in Bacillus subtilis Using Transcriptomics

PubMed Central

Hess, Becky M.; Xue, Junfeng; Markillie, Lye Meng; Taylor, Ronald C.; Wiley, H. Steven; Ahring, Birgitte K.; Linggi, Bryan

2013-01-01

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes, as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding system level regulation and control of the pathway. To address these limitations, we examined Bacillus subtilis grown under multiple conditions and determined the relationship between altered isoprene production and gene expression patterns. We found that with respect to the amount of isoprene produced, terpenoid genes fall into two distinct subsets with opposing correlations. The group whose expression levels positively correlated with isoprene production included dxs, which is responsible for the commitment step in the pathway, ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome-wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. These analyses showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model that accurately predicts production of this secondary metabolite across many simulated environmental conditions. PMID:23840410

The Phosphatidylinositol 3,5-Bisphosphate (PI(3,5)P2)-dependent Tup1 Conversion (PIPTC) Regulates Metabolic Reprogramming from Glycolysis to Gluconeogenesis*

PubMed Central

Han, Bong-Kwan; Emr, Scott D.

2013-01-01

Glucose/carbon metabolism is a fundamental cellular process in living cells. In response to varying environments, eukaryotic cells reprogram their glucose/carbon metabolism between aerobic or anaerobic glycolysis, oxidative phosphorylation, and/or gluconeogenesis. The distinct type of glucose/carbon metabolism that a cell carries out has significant effects on the cell's proliferation and differentiation. However, it is poorly understood how the reprogramming of glucose/carbon metabolism is regulated. Here, we report a novel endosomal PI(3,5)P2 lipid-dependent regulatory mechanism that is required for metabolic reprogramming from glycolysis to gluconeogenesis in Saccharomyces cerevisiae. Certain gluconeogenesis genes, such as FBP1 (encoding fructose-1,6-bisphosphatase 1) and ICL1 (encoding isocitrate lyase 1) are under control of the Mig1 repressor and Cyc8-Tup1 corepressor complex. We previously identified the PI(3,5)P2-dependent Tup1 conversion (PIPTC), a mechanism to convert Cyc8-Tup1 corepressor to Cti6-Cyc8-Tup1 coactivator. We demonstrate that the PIPTC plays a critical role for transcriptional activation of FBP1 and ICL1. Furthermore, without the PIPTC, the Cat8 and Sip4 transcriptional activators cannot be efficiently recruited to the promoters of FBP1 and ICL1, suggesting a key role for the PIPTC in remodulating the chromatin architecture at the promoters. Our findings expand our understanding of the regulatory mechanisms for metabolic reprogramming in eukaryotes to include key regulation steps outside the nucleus. Given that Tup1 and the metabolic enzymes that control PI(3,5)P2 are highly conserved among eukaryotes, our findings may provide important insights toward understanding glucose/carbon metabolic reprogramming in other eukaryotes, including humans. PMID:23733183

The phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2)-dependent Tup1 conversion (PIPTC) regulates metabolic reprogramming from glycolysis to gluconeogenesis.

PubMed

Han, Bong-Kwan; Emr, Scott D

2013-07-12

Glucose/carbon metabolism is a fundamental cellular process in living cells. In response to varying environments, eukaryotic cells reprogram their glucose/carbon metabolism between aerobic or anaerobic glycolysis, oxidative phosphorylation, and/or gluconeogenesis. The distinct type of glucose/carbon metabolism that a cell carries out has significant effects on the cell's proliferation and differentiation. However, it is poorly understood how the reprogramming of glucose/carbon metabolism is regulated. Here, we report a novel endosomal PI(3,5)P2 lipid-dependent regulatory mechanism that is required for metabolic reprogramming from glycolysis to gluconeogenesis in Saccharomyces cerevisiae. Certain gluconeogenesis genes, such as FBP1 (encoding fructose-1,6-bisphosphatase 1) and ICL1 (encoding isocitrate lyase 1) are under control of the Mig1 repressor and Cyc8-Tup1 corepressor complex. We previously identified the PI(3,5)P2-dependent Tup1 conversion (PIPTC), a mechanism to convert Cyc8-Tup1 corepressor to Cti6-Cyc8-Tup1 coactivator. We demonstrate that the PIPTC plays a critical role for transcriptional activation of FBP1 and ICL1. Furthermore, without the PIPTC, the Cat8 and Sip4 transcriptional activators cannot be efficiently recruited to the promoters of FBP1 and ICL1, suggesting a key role for the PIPTC in remodulating the chromatin architecture at the promoters. Our findings expand our understanding of the regulatory mechanisms for metabolic reprogramming in eukaryotes to include key regulation steps outside the nucleus. Given that Tup1 and the metabolic enzymes that control PI(3,5)P2 are highly conserved among eukaryotes, our findings may provide important insights toward understanding glucose/carbon metabolic reprogramming in other eukaryotes, including humans.

Coregulation of terpenoid pathway genes and prediction of isoprene production in Bacillus subtilis using transcriptomics

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

Hess, Becky M.; Xue, Junfeng; Markillie, Lye Meng

2013-06-19

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding the system level regulation and control of the pathway. To address this limitation, we examined Bacillus subtilis grown under multiple conditions and then determined the relationship between altered isoprene production and the pattern of gene expression. Wemore » found that terpenoid genes appeared to fall into two distinct subsets with opposing correlations with respect to the amount of isoprene produced. The group whose expression levels positively correlated with isoprene production included dxs, the gene responsible for the commitment step in the pathway, as well as ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. This analysis showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model which accurately predicts production of this secondary metabolite across many simulated environmental conditions.« less

Mating and male pheromone kill Caenorhabditis males through distinct mechanisms.

PubMed

Shi, Cheng; Runnels, Alexi M; Murphy, Coleen T

2017-03-14

Differences in longevity between sexes is a mysterious yet general phenomenon across great evolutionary distances. To test the roles of responses to environmental cues and sexual behaviors in longevity regulation, we examined Caenorhabditis male lifespan under solitary, grouped, and mated conditions. We find that neurons and the germline are required for male pheromone-dependent male death. Hermaphrodites with a masculinized nervous system secrete male pheromone and are susceptible to male pheromone killing. Male pheromone-mediated killing is unique to androdioecious Caenorhabditis , and may reduce the number of males in hermaphroditic populations; neither males nor females of gonochoristic species are susceptible to male pheromone killing. By contrast, mating-induced death, which is characterized by germline-dependent shrinking, glycogen loss, and ectopic vitellogenin expression, utilizes distinct molecular pathways and is shared between the sexes and across species. The study of sex- and species-specific regulation of aging reveals deeply conserved mechanisms of longevity and population structure regulation.

Distinct bone marrow blood vessels differentially regulate haematopoiesis.

PubMed

Itkin, Tomer; Gur-Cohen, Shiri; Spencer, Joel A; Schajnovitz, Amir; Ramasamy, Saravana K; Kusumbe, Anjali P; Ledergor, Guy; Jung, Yookyung; Milo, Idan; Poulos, Michael G; Kalinkovich, Alexander; Ludin, Aya; Kollet, Orit; Shakhar, Guy; Butler, Jason M; Rafii, Shahin; Adams, Ralf H; Scadden, David T; Lin, Charles P; Lapidot, Tsvee

2016-04-21

Bone marrow endothelial cells (BMECs) form a network of blood vessels that regulate both leukocyte trafficking and haematopoietic stem and progenitor cell (HSPC) maintenance. However, it is not clear how BMECs balance these dual roles, and whether these events occur at the same vascular site. We found that mammalian bone marrow stem cell maintenance and leukocyte trafficking are regulated by distinct blood vessel types with different permeability properties. Less permeable arterial blood vessels maintain haematopoietic stem cells in a low reactive oxygen species (ROS) state, whereas the more permeable sinusoids promote HSPC activation and are the exclusive site for immature and mature leukocyte trafficking to and from the bone marrow. A functional consequence of high permeability of blood vessels is that exposure to blood plasma increases bone marrow HSPC ROS levels, augmenting their migration and differentiation, while compromising their long-term repopulation and survival. These findings may have relevance for clinical haematopoietic stem cell transplantation and mobilization protocols.

Tissue organization by cadherin adhesion molecules: dynamic molecular and cellular mechanisms of morphogenetic regulation

PubMed Central

Niessen, Carien M.; Leckband, Deborah; Yap, Alpha S.

2013-01-01

This review addresses the cellular and molecular mechanisms of cadherin-based tissue morphogenesis. Tissue physiology is profoundly influenced by the distinctive organizations of cells in organs and tissues. In metazoa, adhesion receptors of the classical cadherin family play important roles in establishing and maintaining such tissue organization. Indeed, it is apparent that cadherins participate in a range of morphogenetic events that range from support of tissue integrity to dynamic cellular rearrangements. A comprehensive understanding of cadherin-based morphogenesis must then define the molecular and cellular mechanisms that support these distinct cadherin biologies. Here we focus on four key mechanistic elements: the molecular basis for adhesion through cadherin ectodomains; the regulation of cadherin expression at the cell surface; cooperation between cadherins and the actin cytoskeleton; and regulation by cell signaling. We discuss current progress and outline issues for further research in these fields. PMID:21527735

Generation of multiple cell types in Bacillus subtilis.

PubMed

Lopez, Daniel; Vlamakis, Hera; Kolter, Roberto

2009-01-01

Bacillus subtilis is a Gram-positive bacterium that is well known for its ability to differentiate into metabolically inactive spores that are highly resistant to environmental stresses. In fact, populations of genetically identical B. subtilis comprise numerous distinct cell types. In addition to spores, cells can become genetically competent, motile, produce extracellular matrix or degradative enzymes, or secrete toxins that allow them to cannibalize their neighbors. Many of the cell fates listed above appear to be mutually exclusive. In this review, we discuss how individual cells within a population control their gene expression to ensure that proper regulation of differentiation occurs. These different cell fates are regulated by an intricate network that relies primarily on the activity of three major transcriptional regulators: Spo0A, DegU, and ComK. While individual cells must choose distinct cell fates, the population as a whole exhibits a spectrum of phenotypes whose diversity may increase fitness.

29 CFR Appendix A to Part 35 - Age Distinctions in Statutes Affecting Financial Assistance Administered by DOL

Code of Federal Regulations, 2010 CFR

2010-07-01

... implementing regulations as “a worker at least 16 years of age, * * *, who is employed to learn a skilled trade... 29 Labor 1 2010-07-01 2010-07-01 true Age Distinctions in Statutes Affecting Financial Assistance... BASIS OF AGE IN PROGRAMS OR ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE FROM THE DEPARTMENT OF...

45 CFR 2528.20 - What steps are necessary to use an education award to repay a qualified student loan?

Code of Federal Regulations, 2010 CFR

2010-10-01

... 45 Public Welfare 4 2010-10-01 2010-10-01 false What steps are necessary to use an education award to repay a qualified student loan? 2528.20 Section 2528.20 Public Welfare Regulations Relating to....20 What steps are necessary to use an education award to repay a qualified student loan? (a) Required...

Simulation of load traffic and steeped speed control of conveyor

NASA Astrophysics Data System (ADS)

Reutov, A. A.

2017-10-01

The article examines the possibilities of the step control simulation of conveyor speed within Mathcad, Simulink, Stateflow software. To check the efficiency of the control algorithms and to more accurately determine the characteristics of the control system, it is necessary to simulate the process of speed control with real values of traffic for a work shift or for a day. For evaluating the belt workload and absence of spillage it is necessary to use empirical values of load flow in a shorter period of time. The analytical formulas for optimal speed step values were received using empirical values of load. The simulation checks acceptability of an algorithm, determines optimal parameters of regulation corresponding to load flow characteristics. The average speed and the number of speed switching during simulation are admitted as criteria of regulation efficiency. The simulation example within Mathcad software is implemented. The average conveyor speed decreases essentially by two-step and three-step control. A further increase in the number of regulatory steps decreases average speed insignificantly but considerably increases the intensity of the speed switching. Incremental algorithm of speed regulation uses different number of stages for growing and reducing load traffic. This algorithm allows smooth control of the conveyor speed changes with monotonic variation of the load flow. The load flow oscillation leads to an unjustified increase or decrease of speed. Work results can be applied at the design of belt conveyors with adjustable drives.

Mesenchymal Stem Cells and Myeloid Derived Suppressor Cells: Common Traits in Immune Regulation

PubMed Central

Nikolaev, Alexander

2016-01-01

To protect host against immune-mediated damage, immune responses are tightly regulated. The regulation of immune responses is mediated by various populations of mature immune cells, such as T regulatory cells and B regulatory cells, but also by immature cells of different origins. In this review, we discuss regulatory properties and mechanisms whereby two distinct populations of immature cells, mesenchymal stem cells, and myeloid derived suppressor cells mediate immune regulation, focusing on their similarities, discrepancies, and potential clinical applications. PMID:27529074

ChIP-Seq Analysis for Identifying Genome-Wide Histone Modifications Associated with Stress-Responsive Genes in Plants.

PubMed

Li, Guosheng; Jagadeeswaran, Guru; Mort, Andrew; Sunkar, Ramanjulu

2017-01-01

Histone modifications represent the crux of epigenetic gene regulation essential for most biological processes including abiotic stress responses in plants. Thus, identification of histone modifications at the genome-scale can provide clues for how some genes are 'turned-on' while some others are "turned-off" in response to stress. This chapter details a step-by-step protocol for identifying genome-wide histone modifications associated with stress-responsive gene regulation using chromatin immunoprecipitation (ChIP) followed by sequencing of the DNA (ChIP-seq).

Abstract Interpreters for Free

NASA Astrophysics Data System (ADS)

Might, Matthew

In small-step abstract interpretations, the concrete and abstract semantics bear an uncanny resemblance. In this work, we present an analysis-design methodology that both explains and exploits that resemblance. Specifically, we present a two-step method to convert a small-step concrete semantics into a family of sound, computable abstract interpretations. The first step re-factors the concrete state-space to eliminate recursive structure; this refactoring of the state-space simultaneously determines a store-passing-style transformation on the underlying concrete semantics. The second step uses inference rules to generate an abstract state-space and a Galois connection simultaneously. The Galois connection allows the calculation of the "optimal" abstract interpretation. The two-step process is unambiguous, but nondeterministic: at each step, analysis designers face choices. Some of these choices ultimately influence properties such as flow-, field- and context-sensitivity. Thus, under the method, we can give the emergence of these properties a graph-theoretic characterization. To illustrate the method, we systematically abstract the continuation-passing style lambda calculus to arrive at two distinct families of analyses. The first is the well-known k-CFA family of analyses. The second consists of novel "environment-centric" abstract interpretations, none of which appear in the literature on static analysis of higher-order programs.

Adsorption properties of argon on Ti doped SBA-15.

PubMed

Kim, Euikwoun; Lee, Sang-Hwa; Kim, Jaeyong

2014-11-01

Thermodynamic properties of argon on Ti doped Santa barbara amorphous No. 15 (SBA-15) were investigated in the temperature range of 77-89 K to understand the interaction of gas molecules with porous materials. When the total amount of adsorbed molecules is plotted as a function of the equilibrium vapor pressure of the adsorbed Ar, the results exhibit two distinct isotherm steps. The first step appears at the beginning of the isotherm while the second step locates at 0.7 of the normalized pressure. The existence of the second isotherm step which spanned in the normalized pressure from 0.7 to 0.9 is confirmed when the isotherm data were plotted in terms of the 2-dimensional compressibility values. The total amount of adsorbed molecules forming the second isotherm step is 2.5 times greater than the one for the first step. These adsorption behaviors are typical patterns noted from porous materials and far different from the ones observed from non-pore materials. Our observations demonstrate that most of adsorbed molecules reside in the pores and the height of the second isotherm step is strongly associated with filling pores with gas molecules.

Identification of distinct molecular phenotypes in acute megakaryoblastic leukemia by gene expression profiling

PubMed Central

Bourquin, Jean-Pierre; Subramanian, Aravind; Langebrake, Claudia; Reinhardt, Dirk; Bernard, Olivier; Ballerini, Paola; Baruchel, André; Cavé, Hélène; Dastugue, Nicole; Hasle, Henrik; Kaspers, Gertjan L.; Lessard, Michel; Michaux, Lucienne; Vyas, Paresh; van Wering, Elisabeth; Zwaan, Christian M.; Golub, Todd R.; Orkin, Stuart H.

2006-01-01

Individuals with Down syndrome (DS) are predisposed to develop acute megakaryoblastic leukemia (AMKL), characterized by expression of truncated GATA1 transcription factor protein (GATA1s) due to somatic mutation. The treatment outcome for DS-AMKL is more favorable than for AMKL in non-DS patients. To gain insight into gene expression differences in AMKL, we compared 24 DS and 39 non-DS AMKL samples. We found that non-DS-AMKL samples cluster in two groups, characterized by differences in expression of HOX/TALE family members. Both of these groups are distinct from DS-AMKL, independent of chromosome 21 gene expression. To explore alterations of the GATA1 transcriptome, we used cross-species comparison with genes regulated by GATA1 expression in murine erythroid precursors. Genes repressed after GATA1 induction in the murine system, most notably GATA-2, MYC, and KIT, show increased expression in DS-AMKL, suggesting that GATA1s fail to repress this class of genes. Only a subset of genes that are up-regulated upon GATA1 induction in the murine system show increased expression in DS-AMKL, including GATA1 and BACH1, a probable negative regulator of megakaryocytic differentiation located on chromosome 21. Surprisingly, expression of the chromosome 21 gene RUNX1, a known regulator of megakaryopoiesis, was not elevated in DS-AMKL. Our results identify relevant signatures for distinct AMKL entities and provide insight into gene expression changes associated with these related leukemias. PMID:16492768

Subcellular localization of the Snf1 kinase is regulated by specific β subunits and a novel glucose signaling mechanism

PubMed Central

Vincent, Olivier; Townley, Robert; Kuchin, Sergei; Carlson, Marian

2001-01-01

The Snf1/AMP-activated protein kinase family has broad roles in transcriptional, metabolic, and developmental regulation in response to stress. In Saccharomyces cerevisiae, Snf1 is required for the response to glucose limitation. Snf1 kinase complexes contain the α (catalytic) subunit Snf1, one of the three related β subunits Gal83, Sip1, or Sip2, and the γ subunit Snf4. We present evidence that the β subunits regulate the subcellular localization of the Snf1 kinase. Green fluorescent protein fusions to Gal83, Sip1, and Sip2 show different patterns of localization to the nucleus, vacuole, and/or cytoplasm. We show that Gal83 directs Snf1 to the nucleus in a glucose-regulated manner. We further identify a novel signaling pathway that controls this nuclear localization in response to glucose phosphorylation. This pathway is distinct from the glucose signaling pathway that inhibits Snf1 kinase activity and responds not only to glucose but also to galactose and sucrose. Such independent regulation of the localization and the activity of the Snf1 kinase, combined with the distinct localization of kinases containing different β subunits, affords versatility in regulating physiological responses. PMID:11331606

Addiction, Family Treatment, and Healing Resources: An Interview with David Berenson.

ERIC Educational Resources Information Center

Morgan, Oliver J.

1998-01-01

Interviews Berenson on his distinctive approach to therapy with families and couples affected by addiction and provides references. Considers background and theoretical influences, and changes over time. Discusses the use of "phasing," collaboration with Twelve Step programs, and integration of a spiritual perspective into family and…

Association of distinct mutational signatures with correlates of increased immune activity in pancreatic ductal adenocarcinoma

DOE PAGES

Connor, Ashton A.; Denroche, Robert E.; Jang, Gun Ho; ...

2016-10-20

Outcomes for patients with pancreatic ductal adenocarcinoma (PDAC) remain poor. In addition, advances in next-generation sequencing provide a route to therapeutic approaches, and integrating DNA and RNA analysis with clinicopathologic data may be a crucial step toward personalized treatment strategies for this disease.

Hypermedia as a Distinct Medium: Challenges for Designers.

ERIC Educational Resources Information Center

Clark, Barbara I.; Knupfer, Nancy Nelson

As multimedia development software becomes easier to use and more powerful, instructional designers can establish ways of incorporating the Internet into their lessons. This paper introduces some questions that should be considered prior to stepping into that next level of instructional design. Specifically the paper addresses some of the…

Towards Distinctive and Developmental Curricula at UoTs: The STEPS process at CUT

ERIC Educational Resources Information Center

Mthembu, T. Z.; Orkin, M.; Gering, M.

2012-01-01

Universities of technology (UoTs) achieve developmental impact through differentiated curricula, allowing graduates to undertake mid-level occupations in the workplace. This mandate differs from that at traditional universities in six respects: diploma-level entrants, labour market focus, workplace-oriented learning, applied research and…

Too proud to regulate: The differential effect of pride versus joy on children's ability to delay gratification.

PubMed

Shimoni, Einav; Asbe, Marwa; Eyal, Tal; Berger, Andrea

2016-01-01

We examined the effect of the distinct positive emotions pride and joy on children's self-regulation, focusing on their ability to delay gratification (i.e., resist a temptation in favor of a long-term goal). We hypothesized that because pride corresponds to the attainment of long-term goals and joy corresponds to the attainment of immediate desires, the experience of pride may signal sufficient progress toward a long-term goal, resulting in less delay of gratification than the experience of joy. To test this hypothesis, we induced an experience of pride or joy in 8-year-old children. At this age, the ability to self-regulate--and to experience pride and joy distinctively--is relatively mature. We then measured performance in a delay discounting task. We found that, compared with the joy condition and a control condition, children who experienced pride performed worse on the delay discounting task (p=.045), indicating poorer self-regulation. This result suggests that emotions may function as cues for sufficient goal pursuit, thereby influencing self-regulation from a very young age. Copyright © 2015 Elsevier Inc. All rights reserved.

Prospects for inhibiting the post-transcriptional regulation of gene expression in hepatitis B virus

PubMed Central

Chen, Augustine; Panjaworayan T-Thienprasert, Nattanan; Brown, Chris M

2014-01-01

There is a continuing need for novel antivirals to treat hepatitis B virus (HBV) infection, as it remains a major health problem worldwide. Ideally new classes of antivirals would target multiple steps in the viral lifecycle. In this review, we consider the steps in which HBV RNAs are processed, exported from the nucleus and translated. These are often overlooked steps in the HBV life-cycle. HBV, like retroviruses, incorporates a number of unusual steps in these processes, which use a combination of viral and host cellular machinery. Some of these unusual steps deserve a closer scrutiny. They may provide alternative targets to existing antiviral therapies, which are associated with increasing drug resistance. The RNA post-transcriptional regulatory element identified 20 years ago promotes nucleocytoplasmic export of all unspliced HBV RNAs. There is evidence that inhibition of this step is part of the antiviral action of interferon. Similarly, the structured RNA epsilon element situated at the 5’ end of the polycistronic HBV pregenomic RNA also performs key roles during HBV replication. The pregenomic RNA, which is the template for translation of both the viral core and polymerase proteins, is also encapsidated and used in replication. This complex process, regulated at the epsilon element, also presents an attractive antiviral target. These RNA elements that mediate and regulate gene expression are highly conserved and could be targeted using novel strategies employing RNAi, miRNAs or aptamers. Such approaches targeting these functionally constrained genomic regions should avoid escape mutations. Therefore understanding these regulatory elements, along with providing potential targets, may also facilitate the development of other new classes of antiviral drugs. PMID:25009369

Phenotype- and genotype-specific structural alterations in spasmodic dysphonia.

PubMed

Bianchi, Serena; Battistella, Giovanni; Huddleston, Hailey; Scharf, Rebecca; Fleysher, Lazar; Rumbach, Anna F; Frucht, Steven J; Blitzer, Andrew; Ozelius, Laurie J; Simonyan, Kristina

2017-04-01

Spasmodic dysphonia is a focal dystonia characterized by involuntary spasms in the laryngeal muscles that occur selectively during speaking. Although hereditary trends have been reported in up to 16% of patients, the causative etiology of spasmodic dysphonia is unclear, and the influences of various phenotypes and genotypes on disorder pathophysiology are poorly understood. In this study, we examined structural alterations in cortical gray matter and white matter integrity in relationship to different phenotypes and putative genotypes of spasmodic dysphonia to elucidate the structural component of its complex pathophysiology. Eighty-nine patients with spasmodic dysphonia underwent high-resolution magnetic resonance imaging and diffusion-weighted imaging to examine cortical thickness and white matter fractional anisotropy in adductor versus abductor forms (distinct phenotypes) and in sporadic versus familial cases (distinct genotypes). Phenotype-specific abnormalities were localized in the left sensorimotor cortex and angular gyrus and the white matter bundle of the right superior corona radiata. Genotype-specific alterations were found in the left superior temporal gyrus, supplementary motor area, and the arcuate portion of the left superior longitudinal fasciculus. Our findings suggest that phenotypic differences in spasmodic dysphonia arise at the level of the primary and associative areas of motor control, whereas genotype-related pathophysiological mechanisms may be associated with dysfunction of regions regulating phonological and sensory processing. Identification of structural alterations specific to disorder phenotype and putative genotype provides an important step toward future delineation of imaging markers and potential targets for novel therapeutic interventions for spasmodic dysphonia. © 2017 International Parkinson and Movement Disorder Society. © 2017 International Parkinson and Movement Disorder Society.

Linear-Quadratic-Gaussian Regulator Developed for a Magnetic Bearing

NASA Technical Reports Server (NTRS)

Choi, Benjamin B.

2002-01-01

Linear-Quadratic-Gaussian (LQG) control is a modern state-space technique for designing optimal dynamic regulators. It enables us to trade off regulation performance and control effort, and to take into account process and measurement noise. The Structural Mechanics and Dynamics Branch at the NASA Glenn Research Center has developed an LQG control for a fault-tolerant magnetic bearing suspension rig to optimize system performance and to reduce the sensor and processing noise. The LQG regulator consists of an optimal state-feedback gain and a Kalman state estimator. The first design step is to seek a state-feedback law that minimizes the cost function of regulation performance, which is measured by a quadratic performance criterion with user-specified weighting matrices, and to define the tradeoff between regulation performance and control effort. The next design step is to derive a state estimator using a Kalman filter because the optimal state feedback cannot be implemented without full state measurement. Since the Kalman filter is an optimal estimator when dealing with Gaussian white noise, it minimizes the asymptotic covariance of the estimation error.

A rational model for maximizing the effects of therapeutic relationship regulation in personality disorders with poor metacognition and over-regulation of affects.

PubMed

Dimaggio, Giancarlo; Carcione, Antonino; Salvatore, Giampaolo; Semerari, Antonio; Nicolò, Giuseppe

2010-11-01

The therapeutic relationship plays a key role in personality disorder (PD) psychotherapy. Some aspects of therapeutic relationship regulation appear important for treatment of PD clients, including those with constricted relational schemas, poor metacognition, and over-regulation of affects described here. AIM.: To propose a rational model for how and when to work on the therapeutic relationship by treating the underlying personality pathology. Formalize a step-by-step procedure for performing operations such as validation of clients' experiences, creating a sense of sharedness, assessing the quality of the therapeutic relationship in order to prevent and repair ruptures in the alliance, self-disclosing by the therapist, and metacommunication on the basis of clients' responses to treatment. We discuss the implications of this model for further research into the PD therapy process. 2010 The British Psychological Society.

Temporomandibular joint formation requires two distinct hedgehog-dependent steps.

PubMed

Purcell, Patricia; Joo, Brian W; Hu, Jimmy K; Tran, Pamela V; Calicchio, Monica L; O'Connell, Daniel J; Maas, Richard L; Tabin, Clifford J

2009-10-27

We conducted a genetic analysis of the developing temporo-mandibular or temporomandi-bular joint (TMJ), a highly specialized synovial joint that permits movement and function of the mammalian jaw. First, we used laser capture microdissection to perform a genome-wide expression analysis of each of its developing components. The expression patterns of genes identified in this screen were examined in the TMJ and compared with those of other synovial joints, including the shoulder and the hip joints. Striking differences were noted, indicating that the TMJ forms via a distinct molecular program. Several components of the hedgehog (Hh) signaling pathway are among the genes identified in the screen, including Gli2, which is expressed specifically in the condyle and in the disk of the developing TMJ. We found that mice deficient in Gli2 display aberrant TMJ development such that the condyle loses its growth-plate-like cellular organization and no disk is formed. In addition, we used a conditional strategy to remove Smo, a positive effector of the Hh signaling pathway, from chondrocyte progenitors. This cell autonomous loss of Hh signaling allows for disk formation, but the resulting structure fails to separate from the condyle. Thus, these experiments establish that Hh signaling acts at two distinct steps in disk morphogenesis, condyle initiation, and disk-condyle separation and provide a molecular framework for future studies of the TMJ.

Temporomandibular joint formation requires two distinct hedgehog-dependent steps

PubMed Central

Purcell, Patricia; Joo, Brian W.; Hu, Jimmy K.; Tran, Pamela V.; Calicchio, Monica L.; O'Connell, Daniel J.; Maas, Richard L.; Tabin, Clifford J.

2009-01-01

We conducted a genetic analysis of the developing temporo-mandibular or temporomandi-bular joint (TMJ), a highly specialized synovial joint that permits movement and function of the mammalian jaw. First, we used laser capture microdissection to perform a genome-wide expression analysis of each of its developing components. The expression patterns of genes identified in this screen were examined in the TMJ and compared with those of other synovial joints, including the shoulder and the hip joints. Striking differences were noted, indicating that the TMJ forms via a distinct molecular program. Several components of the hedgehog (Hh) signaling pathway are among the genes identified in the screen, including Gli2, which is expressed specifically in the condyle and in the disk of the developing TMJ. We found that mice deficient in Gli2 display aberrant TMJ development such that the condyle loses its growth-plate-like cellular organization and no disk is formed. In addition, we used a conditional strategy to remove Smo, a positive effector of the Hh signaling pathway, from chondrocyte progenitors. This cell autonomous loss of Hh signaling allows for disk formation, but the resulting structure fails to separate from the condyle. Thus, these experiments establish that Hh signaling acts at two distinct steps in disk morphogenesis, condyle initiation, and disk–condyle separation and provide a molecular framework for future studies of the TMJ. PMID:19815519

Distinct pH regulation of slow and rapid anion channels at the plasma membrane of Arabidopsis thaliana hypocotyl cells.

PubMed

Colcombet, Jean; Lelièvre, Françoise; Thomine, Sébastien; Barbier-Brygoo, Hélène; Frachisse, Jean-Marie

2005-07-01

Variations in both intracellular and extracellular pH are known to be involved in a wealth of physiological responses. Using the patch-clamp technique on Arabidopsis hypocotyl cells, it is shown that rapid-type and slow-type anion channels at the plasma membrane are both regulated by pH via distinct mechanisms. Modifications of pH modulate the voltage-dependent gating of the rapid channel. While intracellular alkalinization facilitates channel activation by shifting the voltage gate towards negative potentials, extracellular alkalinization shifts the activation threshold to more positive potentials, away from physiological resting membrane potentials. By contrast, pH modulates slow anion channel activity in a voltage-independent manner. Intracellular acidification and extracellular alkalinization increase slow anion channel currents. The possible role of these distinct modulations in physiological processes involving anion efflux and modulation of extracellular and/or intracellular pH, such as elicitor and ABA signalling, are discussed.

Non-additive interactions involving two distinct elements mediate sloppy-paired regulation by pair-rule transcription factors

PubMed Central

Prazak, Lisa; Fujioka, Miki; Gergen, J. Peter

2010-01-01

The relatively simple combinatorial rules responsible for establishing the initial metameric expression of sloppy-paired-1 (slp1) in the Drosophila blastoderm embryo make this system an attractive model for investigating the mechanism of regulation by pair rule transcription factors. This investigation of slp1 cis-regulatory architecture identifies two distinct elements, a proximal early stripe element (PESE) and a distal early stripe element (DESE) located from −3.1 kb to −2.5 kb and from −8.1 kb to −7.1 kb upstream of the slp1 promoter, respectively, that mediate this early regulation. The proximal element expresses only even-numbered stripes and mediates repression by Even-skipped (Eve) as well as by the combination of Runt and Fushi-tarazu (Ftz). A 272 basepair sub-element of PESE retains Eve-dependent repression, but is expressed throughout the even-numbered parasegments due to the loss of repression by Runt and Ftz. In contrast, the distal element expresses both odd and even-numbered stripes and also drives inappropriate expression in the anterior half of the odd-numbered parasegments due to an inability to respond to repression by Eve. Importantly, a composite reporter gene containing both early stripe elements recapitulates pair-rule gene-dependent regulation in a manner beyond what is expected from combining their individual patterns. These results indicate interactions involving distinct cis-elements contribute to the proper integration of pair-rule regulatory information. A model fully accounting for these results proposes that metameric slp1 expression is achieved through the Runt-dependent regulation of interactions between these two pair-rule response elements and the slp1 promoter. PMID:20435028

Promoter analysis reveals globally differential regulation of human long non-coding RNA and protein-coding genes

DOE PAGES

Alam, Tanvir; Medvedeva, Yulia A.; Jia, Hui; ...

2014-10-02

Transcriptional regulation of protein-coding genes is increasingly well-understood on a global scale, yet no comparable information exists for long non-coding RNA (lncRNA) genes, which were recently recognized to be as numerous as protein-coding genes in mammalian genomes. We performed a genome-wide comparative analysis of the promoters of human lncRNA and protein-coding genes, finding global differences in specific genetic and epigenetic features relevant to transcriptional regulation. These two groups of genes are hence subject to separate transcriptional regulatory programs, including distinct transcription factor (TF) proteins that significantly favor lncRNA, rather than coding-gene, promoters. We report a specific signature of promoter-proximal transcriptionalmore » regulation of lncRNA genes, including several distinct transcription factor binding sites (TFBS). Experimental DNase I hypersensitive site profiles are consistent with active configurations of these lncRNA TFBS sets in diverse human cell types. TFBS ChIP-seq datasets confirm the binding events that we predicted using computational approaches for a subset of factors. For several TFs known to be directly regulated by lncRNAs, we find that their putative TFBSs are enriched at lncRNA promoters, suggesting that the TFs and the lncRNAs may participate in a bidirectional feedback loop regulatory network. Accordingly, cells may be able to modulate lncRNA expression levels independently of mRNA levels via distinct regulatory pathways. Our results also raise the possibility that, given the historical reliance on protein-coding gene catalogs to define the chromatin states of active promoters, a revision of these chromatin signature profiles to incorporate expressed lncRNA genes is warranted in the future.« less

Promoter analysis reveals globally differential regulation of human long non-coding RNA and protein-coding genes

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

Alam, Tanvir; Medvedeva, Yulia A.; Jia, Hui

Transcriptional regulation of protein-coding genes is increasingly well-understood on a global scale, yet no comparable information exists for long non-coding RNA (lncRNA) genes, which were recently recognized to be as numerous as protein-coding genes in mammalian genomes. We performed a genome-wide comparative analysis of the promoters of human lncRNA and protein-coding genes, finding global differences in specific genetic and epigenetic features relevant to transcriptional regulation. These two groups of genes are hence subject to separate transcriptional regulatory programs, including distinct transcription factor (TF) proteins that significantly favor lncRNA, rather than coding-gene, promoters. We report a specific signature of promoter-proximal transcriptionalmore » regulation of lncRNA genes, including several distinct transcription factor binding sites (TFBS). Experimental DNase I hypersensitive site profiles are consistent with active configurations of these lncRNA TFBS sets in diverse human cell types. TFBS ChIP-seq datasets confirm the binding events that we predicted using computational approaches for a subset of factors. For several TFs known to be directly regulated by lncRNAs, we find that their putative TFBSs are enriched at lncRNA promoters, suggesting that the TFs and the lncRNAs may participate in a bidirectional feedback loop regulatory network. Accordingly, cells may be able to modulate lncRNA expression levels independently of mRNA levels via distinct regulatory pathways. Our results also raise the possibility that, given the historical reliance on protein-coding gene catalogs to define the chromatin states of active promoters, a revision of these chromatin signature profiles to incorporate expressed lncRNA genes is warranted in the future.« less

O-GlcNAc transferase regulates transcriptional activity of human Oct4.

PubMed

Constable, Sandii; Lim, Jae-Min; Vaidyanathan, Krithika; Wells, Lance

2017-10-01

O-linked β-N-acetylglucosamine (O-GlcNAc) is a single sugar modification found on many different classes of nuclear and cytoplasmic proteins. Addition of this modification, by the enzyme O-linked N-acetylglucosamine transferase (OGT), is dynamic and inducible. One major class of proteins modified by O-GlcNAc is transcription factors. O-GlcNAc regulates transcription factor properties through a variety of different mechanisms including localization, stability and transcriptional activation. Maintenance of embryonic stem (ES) cell pluripotency requires tight regulation of several key transcription factors, many of which are modified by O-GlcNAc. Octamer-binding protein 4 (Oct4) is one of the key transcription factors required for pluripotency of ES cells and more recently, the generation of induced pluripotent stem (iPS) cells. The action of Oct4 is modulated by the addition of several post-translational modifications, including O-GlcNAc. Previous studies in mice found a single site of O-GlcNAc addition responsible for transcriptional regulation. This study was designed to determine if this mechanism is conserved in humans. We mapped 10 novel sites of O-GlcNAc attachment on human Oct4, and confirmed a role for OGT in transcriptional activation of Oct4 at a site distinct from that found in mouse that allows distinction between different Oct4 target promoters. Additionally, we uncovered a potential new role for OGT that does not include its catalytic function. These results confirm that human Oct4 activity is being regulated by OGT by a mechanism that is distinct from mouse Oct4. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

CEF1/CDC5 alleles modulate transitions between catalytic conformations of the spliceosome

PubMed Central

Query, Charles C.; Konarska, Maria M.

2012-01-01

Conformational change within the spliceosome is required between the first and second catalytic steps of pre-mRNA splicing. A prior genetic screen for suppressors of an intron mutant that stalls between the two steps yielded both prp8 and non-prp8 alleles that suppressed second-step splicing defects. We have now identified the strongest non-prp8 suppressors as alleles of the NTC (Prp19 complex) component, CEF1. These cef1 alleles generally suppress second-step defects caused by a variety of intron mutations, mutations in U6 snRNA, or deletion of the second-step protein factor Prp17, and they can activate alternative 3′ splice sites. Genetic and functional interactions between cef1 and prp8 alleles suggest that they modulate the same event(s) in the first-to-second-step transition, most likely by stabilization of the second-step spliceosome; in contrast, alleles of U6 snRNA that also alter this transition modulate a distinct event, most likely by stabilization of the first-step spliceosome. These results implicate a myb-like domain of Cef1/CDC5 in interactions that modulate conformational states of the spliceosome and suggest that alteration of these events affects splice site use, resulting in alternative splicing-like patterns in yeast. PMID:22408182

STEP activation by Gαq coupled GPCRs opposes Src regulation of NMDA receptors containing the GluN2A subunit

PubMed Central

Tian, Meng; Xu, Jian; Lei, Gang; Lombroso, Paul J.; Jackson, Michael F.; MacDonald, John F.

2016-01-01

N-methyl-D-aspartate receptors (NMDARs) are necessary for the induction of synaptic plasticity and for the consolidation of learning and memory. NMDAR function is tightly regulated by functionally opposed families of kinases and phosphatases. Herein we show that the striatal-enriched protein tyrosine phosphatase (STEP) is recruited by Gαq-coupled receptors, including the M1 muscarinic acetylcholine receptor (M1R), and opposes the Src tyrosine kinase-mediated increase in the function of NMDARs composed of GluN2A. STEP activation by M1R stimulation requires IP3Rs and can depress NMDA-evoked currents with modest intracellular Ca2+ buffering. Src recruitment by M1R stimulation requires coincident NMDAR activation and can augment NMDA-evoked currents with high intracellular Ca2+ buffering. Our findings suggest that Src and STEP recruitment is contingent on differing intracellular Ca2+ dynamics that dictate whether NMDAR function is augmented or depressed following M1R stimulation. PMID:27857196

Specialisation within the DWARF14 protein family confers distinct responses to karrikins and strigolactones in Arabidopsis.

PubMed

Waters, Mark T; Nelson, David C; Scaffidi, Adrian; Flematti, Gavin R; Sun, Yueming K; Dixon, Kingsley W; Smith, Steven M

2012-04-01

Karrikins are butenolides derived from burnt vegetation that stimulate seed germination and enhance seedling responses to light. Strigolactones are endogenous butenolide hormones that regulate shoot and root architecture, and stimulate the branching of arbuscular mycorrhizal fungi. Thus, karrikins and strigolactones are structurally similar but physiologically distinct plant growth regulators. In Arabidopsis thaliana, responses to both classes of butenolides require the F-box protein MAX2, but it remains unclear how discrete responses to karrikins and strigolactones are achieved. In rice, the DWARF14 protein is required for strigolactone-dependent inhibition of shoot branching. Here, we show that the Arabidopsis DWARF14 orthologue, AtD14, is also necessary for normal strigolactone responses in seedlings and adult plants. However, the AtD14 paralogue KARRIKIN INSENSITIVE 2 (KAI2) is specifically required for responses to karrikins, and not to strigolactones. Phylogenetic analysis indicates that KAI2 is ancestral and that AtD14 functional specialisation has evolved subsequently. Atd14 and kai2 mutants exhibit distinct subsets of max2 phenotypes, and expression patterns of AtD14 and KAI2 are consistent with the capacity to respond to either strigolactones or karrikins at different stages of plant development. We propose that AtD14 and KAI2 define a class of proteins that permit the separate regulation of karrikin and strigolactone signalling by MAX2. Our results support the existence of an endogenous, butenolide-based signalling mechanism that is distinct from the strigolactone pathway, providing a molecular basis for the adaptive response of plants to smoke.

Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

PubMed

Le Billan, Florian; Amazit, Larbi; Bleakley, Kevin; Xue, Qiong-Yao; Pussard, Eric; Lhadj, Christophe; Kolkhof, Peter; Viengchareun, Say; Fagart, Jérôme; Lombès, Marc

2018-05-07

Mineralocorticoid receptors (MRs) and glucocorticoid receptors (GRs) are two closely related hormone-activated transcription factors that regulate major pathophysiologic functions. High homology between these receptors accounts for the crossbinding of their corresponding ligands, MR being activated by both aldosterone and cortisol and GR essentially activated by cortisol. Their coexpression and ability to bind similar DNA motifs highlight the need to investigate their respective contributions to overall corticosteroid signaling. Here, we decipher the transcriptional regulatory mechanisms that underlie selective effects of MRs and GRs on shared genomic targets in a human renal cellular model. Kinetic, serial, and sequential chromatin immunoprecipitation approaches were performed on the period circadian protein 1 ( PER1) target gene, providing evidence that both receptors dynamically and cyclically interact at the same target promoter in a specific and distinct transcriptional signature. During this process, both receptors regulate PER1 gene by binding as homo- or heterodimers to the same promoter region. Our results suggest a novel level of MR-GR target gene regulation, which should be considered for a better and integrated understanding of corticosteroid-related pathophysiology.-Le Billan, F., Amazit, L., Bleakley, K., Xue, Q.-Y., Pussard, E., Lhadj, C., Kolkhof, P., Viengchareun, S., Fagart, J., Lombès, M. Corticosteroid receptors adopt distinct cyclical transcriptional signatures.

Dictyostelium LvsB has a regulatory role in endosomal vesicle fusion

PubMed Central

Falkenstein, Kristin; De Lozanne, Arturo

2014-01-01

ABSTRACT Defects in human lysosomal-trafficking regulator (Lyst) are associated with the lysosomal disorder Chediak–Higashi syndrome. The absence of Lyst results in the formation of enlarged lysosome-related compartments, but the mechanism for how these compartments arise is not well established. Two opposing models have been proposed to explain Lyst function. The fission model describes Lyst as a positive regulator of fission from lysosomal compartments, whereas the fusion model identifies Lyst as a negative regulator of fusion between lysosomal vesicles. Here, we used assays that can distinguish between defects in vesicle fusion versus fission. We compared the phenotype of Dictyostelium discoideum cells defective in LvsB, the ortholog of Lyst, with that of two known fission defect mutants (μ3- and WASH-null mutants). We found that the temporal localization characteristics of the post-lysosomal marker vacuolin, as well as vesicular acidity and the fusion dynamics of LvsB-null cells are distinct from those of both μ3- and WASH-null fission defect mutants. These distinctions are predicted by the fusion defect model and implicate LvsB as a negative regulator of vesicle fusion. PMID:25086066

Regulation of Corneal Stroma Extracellular Matrix Assembly

PubMed Central

Chen, Shoujun; Mienaltowski, Michael J.; Birk, David E.

2014-01-01

The transparent cornea is the major refractive element of the eye. A finely controlled assembly of the stromal extracellular matrix is critical to corneal function, as well as in establishing the appropriate mechanical stability required to maintain corneal shape and curvature. In the stroma, homogeneous, small diameter collagen fibrils, regularly packed with a highly ordered hierarchical organization, are essential for function. This review focuses on corneal stroma assembly and the regulation of collagen fibrillogenesis. Corneal collagen fibrillogenesis involves multiple molecules interacting in sequential steps, as well as interactions between keratocytes and stroma matrix components. The stroma has the highest collagen V:I ratio in the body. Collagen V regulates the nucleation of protofibril assembly, thus controlling the number of fibrils and assembly of smaller diameter fibrils in the stroma. The corneal stroma is also enriched in small leucine-rich proteoglycans (SLRPs) that cooperate in a temporal and spatial manner to regulate linear and lateral collagen fibril growth. In addition, the fibril-associated collagens (FACITs) such as collagen XII and collagen XIV have roles in the regulation of fibril packing and inter-lamellar interactions. A communicating keratocyte network contributes to the overall and long-range regulation of stromal extracellular matrix assembly, by creating micro-domains where the sequential steps in stromal matrix assembly are controlled. Keratocytes control the synthesis of extracellular matrix components, which interact with the keratocytes dynamically to coordinate the regulatory steps into a cohesive process. Mutations or deficiencies in stromal regulatory molecules result in altered interactions and deficiencies in both transparency and refraction, leading to corneal stroma pathobiology such as stromal dystrophies, cornea plana and keratoconus. PMID:25819456

Evolution of the circuitry for conscious color vision in primates

PubMed Central

Neitz, J; Neitz, M

2017-01-01

There are many ganglion cell types and subtypes in our retina that carry color information. These have appeared at different times over the history of the evolution of the vertebrate visual system. They project to several different places in the brain and serve a variety of purposes allowing wavelength information to contribute to diverse visual functions. These include circadian photoentrainment, regulation of sleep and mood, guidance of orienting movements, detection and segmentation of objects. Predecessors to some of the circuits serving these purposes presumably arose before mammals evolved and different functions are represented by distinct ganglion cell types. However, while other animals use color information to elicit motor movements and regulate activity rhythms, as do humans, using phylogenetically ancient circuitry, the ability to appreciate color appearance may have been refined in ancestors to primates, mediated by a special set of ganglion cells that serve only that purpose. Understanding the circuitry for color vision has implications for the possibility of treating color blindness using gene therapy by recapitulating evolution. In addition, understanding how color is encoded, including how chromatic and achromatic percepts are separated is a step toward developing a complete picture of the diversity of ganglion cell types and their functions. Such knowledge could be useful in developing therapeutic strategies for blinding eye disorders that rely on stimulating elements in the retina, where more than 50 different neuron types are organized into circuits that transform signals from photoreceptors into specialized detectors many of which are not directly involved in conscious vision. PMID:27935605

Evolution of the circuitry for conscious color vision in primates.

PubMed

Neitz, J; Neitz, M

2017-02-01

There are many ganglion cell types and subtypes in our retina that carry color information. These have appeared at different times over the history of the evolution of the vertebrate visual system. They project to several different places in the brain and serve a variety of purposes allowing wavelength information to contribute to diverse visual functions. These include circadian photoentrainment, regulation of sleep and mood, guidance of orienting movements, detection and segmentation of objects. Predecessors to some of the circuits serving these purposes presumably arose before mammals evolved and different functions are represented by distinct ganglion cell types. However, while other animals use color information to elicit motor movements and regulate activity rhythms, as do humans, using phylogenetically ancient circuitry, the ability to appreciate color appearance may have been refined in ancestors to primates, mediated by a special set of ganglion cells that serve only that purpose. Understanding the circuitry for color vision has implications for the possibility of treating color blindness using gene therapy by recapitulating evolution. In addition, understanding how color is encoded, including how chromatic and achromatic percepts are separated is a step toward developing a complete picture of the diversity of ganglion cell types and their functions. Such knowledge could be useful in developing therapeutic strategies for blinding eye disorders that rely on stimulating elements in the retina, where more than 50 different neuron types are organized into circuits that transform signals from photoreceptors into specialized detectors many of which are not directly involved in conscious vision.

Direct Interactions Between Gli3, Wnt8b, and Fgfs Underlie Patterning of the Dorsal Telencephalon.

PubMed

Hasenpusch-Theil, Kerstin; Watson, Julia A; Theil, Thomas

2017-02-01

A key step in the development of the cerebral cortex is a patterning process, which subdivides the telencephalon into several molecularly distinct domains and is critical for cortical arealization. This process is dependent on a complex network of interactions between signaling molecules of the Fgf and Wnt gene families and the Gli3 transcription factor gene, but a better knowledge of the molecular basis of the interplay between these factors is required to gain a deeper understanding of the genetic circuitry underlying telencephalic patterning. Using DNA-binding and reporter gene assays, we here investigate the possibility that Gli3 and these signaling molecules interact by directly regulating each other's expression. We show that Fgf signaling is required for Wnt8b enhancer activity in the cortical hem, whereas Wnt/β-catenin signaling represses Fgf17 forebrain enhancer activity. In contrast, Fgf and Wnt/β-catenin signaling cooperate to regulate Gli3 expression. Taken together, these findings indicate that mutual interactions between Gli3, Wnt8b, and Fgf17 are crucial elements of the balance between these factors thereby conferring robustness to the patterning process. Hence, our study provides a framework for understanding the genetic circuitry underlying telencephalic patterning and how defects in this process can affect the formation of cortical areas. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Pax2/8 proteins coordinate sequential induction of otic and epibranchial placodes through differential regulation of foxi1, sox3 and fgf24.

PubMed

Padanad, Mahesh S; Riley, Bruce B

2011-03-01

Vertebrate cranial placodes contribute vitally to development of sensory structures of the head. Amongst posterior placodes, the otic placode forms the inner ear whereas nearby epibranchial placodes produce sensory ganglia within branchial clefts. Though diverse in fate, these placodes show striking similarities in their early regulation. In zebrafish, both are initiated by localized Fgf signaling plus the ubiquitous competence factor Foxi1, and both express pax8 and sox3 in response. It has been suggested that Fgf initially induces a common otic/epibranchial field, which later subdivides in response to other signals. However, we find that otic and epibranchial placodes form at different times and by distinct mechanisms. Initially, Fgf from surrounding tissues induces otic expression of pax8 and sox3, which cooperate synergistically to establish otic fate. Subsequently, pax8 works with related genes pax2a/pax2b to downregulate otic expression of foxi1, a necessary step for further otic development. Additionally, pax2/8 activate otic expression of fgf24, which induces epibranchial expression of sox3. Knockdown of fgf24 or sox3 causes severe epibranchial deficiencies but has little effect on otic development. These findings clarify the roles of pax8 and sox3 and support a model whereby the otic placode forms first and induces epibranchial placodes through an Fgf-relay. Copyright © 2011 Elsevier Inc. All rights reserved.

VPS9a Activates the Rab5 GTPase ARA7 to Confer Distinct Pre- and Postinvasive Plant Innate Immunity.

PubMed

Nielsen, Mads E; Jürgens, Gerd; Thordal-Christensen, Hans

2017-08-01

Plant innate immunity can effectively prevent the proliferation of filamentous pathogens. Papilla formation at the site of attack is essential for preinvasive immunity; in postinvasive immunity, the encasement of pathogen structures inside host cells can hamper disease. Whereas papillae are highly dependent on transcytosis of premade material, little is known about encasement formation. Here, we show that endosome-associated VPS9a, the conserved guanine-nucleotide exchange factor activating Rab5 GTPases, is required for both pre- and postinvasive immunity against a nonadapted powdery mildew fungus ( Blumeria graminis f. sp hordei ) in Arabidopsis thaliana Surprisingly, VPS9a acts in addition to two previously well-described innate immunity components and thus represents an additional step in the regulation of how plants resist pathogens. We found VPS9a to be important for delivering membrane material to the encasement and VPS9a also plays a predominant role in postinvasive immunity. GTP-bound Rab5 GTPases accumulate in the encasement, but not the papillae, suggesting that two independent pathways form these defense structures. VPS9a also mediates defense to an adapted powdery mildew fungus, thus regulating a durable type of defense that works in both host and nonhost resistance. We propose that VPS9a plays a conserved role in organizing cellular endomembrane trafficking, required for delivery of defense components in response to powdery mildew fungi. © 2017 American Society of Plant Biologists. All rights reserved.

BID links ferroptosis to mitochondrial cell death pathways.

PubMed

Neitemeier, Sandra; Jelinek, Anja; Laino, Vincenzo; Hoffmann, Lena; Eisenbach, Ina; Eying, Roman; Ganjam, Goutham K; Dolga, Amalia M; Oppermann, Sina; Culmsee, Carsten

2017-08-01

Ferroptosis has been defined as an oxidative and iron-dependent pathway of regulated cell death that is distinct from caspase-dependent apoptosis and established pathways of death receptor-mediated regulated necrosis. While emerging evidence linked features of ferroptosis induced e.g. by erastin-mediated inhibition of the X c - system or inhibition of glutathione peroxidase 4 (Gpx4) to an increasing number of oxidative cell death paradigms in cancer cells, neurons or kidney cells, the biochemical pathways of oxidative cell death remained largely unclear. In particular, the role of mitochondrial damage in paradigms of ferroptosis needs further investigation. In the present study, we find that erastin-induced ferroptosis in neuronal cells was accompanied by BID transactivation to mitochondria, loss of mitochondrial membrane potential, enhanced mitochondrial fragmentation and reduced ATP levels. These hallmarks of mitochondrial demise are also established features of oxytosis, a paradigm of cell death induced by X c - inhibition by millimolar concentrations of glutamate. Bid knockout using CRISPR/Cas9 approaches preserved mitochondrial integrity and function, and mediated neuroprotective effects against both, ferroptosis and oxytosis. Furthermore, the BID-inhibitor BI-6c9 inhibited erastin-induced ferroptosis, and, in turn, the ferroptosis inhibitors ferrostatin-1 and liproxstatin-1 prevented mitochondrial dysfunction and cell death in the paradigm of oxytosis. These findings show that mitochondrial transactivation of BID links ferroptosis to mitochondrial damage as the final execution step in this paradigm of oxidative cell death. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

The structure of bradyzoite-specific enolase from Toxoplasma gondii reveals insights into its dual cytoplasmic and nuclear functions

DOE PAGES

Ruan, Jiapeng; Mouveaux, Thomas; Light, Samuel H.; ...

2015-03-01

In addition to catalyzing a central step in glycolysis, enolase assumes a remarkably diverse set of secondary functions in different organisms, including transcription regulation as documented for the oncogene c-Myc promoter-binding protein 1. The apicomplexan parasite Toxoplasma gondii differentially expresses two nuclear-localized, plant-like enolases: enolase 1 (TgENO1) in the latent bradyzoite cyst stage and enolase 2 (TgENO2) in the rapidly replicative tachyzoite stage. A 2.75 Å resolution crystal structure of bradyzoite enolase 1, the second structure to be reported of a bradyzoite-specific protein inToxoplasma, captures an open conformational state and reveals that distinctive plant-like insertions are located on surface loops.more » The enolase 1 structure reveals that a unique residue, Glu164, in catalytic loop 2 may account for the lower activity of this cyst-stage isozyme. Recombinant TgENO1 specifically binds to a TTTTCT DNA motif present in the cyst matrix antigen 1 (TgMAG1) gene promoter as demonstrated by gel retardation. Furthermore, direct physical interactions of both nuclear TgENO1 and TgENO2 with the TgMAG1 gene promoter are demonstrated n vivo using chromatin immunoprecipitation (ChIP) assays. Structural and biochemical studies reveal that T. gondii enolase functions are multifaceted, including the coordination of gene regulation in parasitic stage development. Lastly, enolase 1 provides a potential lead in the design of drugs against Toxoplasma brain cysts.« less

Parental feeding styles and adolescents' healthy eating habits. Structure and correlates of a Costa Rican questionnaire.

PubMed

Monge-Rojas, Rafael; Smith-Castro, Vanesa; Colon-Ramos, Uriyoán; Garita-Arce, Carlos; Sánchez-López, Marta; Chinnock, Anne

2010-10-01

This study designed and validated a questionnaire aimed at examining parental feeding styles to encourage healthy eating habits among Costa Rican adolescents. Adolescents (n=133; mean age 15.4 years), and their parents, participated in the study. The parents completed a parental feeding style questionnaire, and the adolescents completed 3-day food records. Confirmatory factor analyses suggest four distinct parental feeding styles, (a) verbal encouragement of healthy eating behaviors; (b) use of verbal sanctions to indirectly control the intake of healthy food; (c) direct control of access to and intake of food; and (d) use of food to regulate emotions and behavior. There were no correlations between dietary intake and the verbal encouragement of healthy eating behaviors, but there were significant negative correlations between (1) "the use of verbal sanctions to indirectly control the intake of healthy food", and the consumption of fruit and vegetable, of calcium, iron, vitamin B6 and folic acid intake, and (2) between the "direct control of access to and intake of food" and fast food consumption and total carbohydrates intake. The use of food to regulate emotions and behavior was positively correlated with high energy-dense food consumption. Stratification of the data shows significant differences by gender in the correlations between parental feeding style and dietary intake. Understanding parental feeding styles in a Latin American context is a first step in helping researchers develops culturally-appropriate parenting intervention/prevention strategies to encourage healthy eating behaviors during adolescence.

VPS9a Activates the Rab5 GTPase ARA7 to Confer Distinct Pre- and Postinvasive Plant Innate Immunity[OPEN

PubMed Central

2017-01-01

Plant innate immunity can effectively prevent the proliferation of filamentous pathogens. Papilla formation at the site of attack is essential for preinvasive immunity; in postinvasive immunity, the encasement of pathogen structures inside host cells can hamper disease. Whereas papillae are highly dependent on transcytosis of premade material, little is known about encasement formation. Here, we show that endosome-associated VPS9a, the conserved guanine-nucleotide exchange factor activating Rab5 GTPases, is required for both pre- and postinvasive immunity against a nonadapted powdery mildew fungus (Blumeria graminis f. sp hordei) in Arabidopsis thaliana. Surprisingly, VPS9a acts in addition to two previously well-described innate immunity components and thus represents an additional step in the regulation of how plants resist pathogens. We found VPS9a to be important for delivering membrane material to the encasement and VPS9a also plays a predominant role in postinvasive immunity. GTP-bound Rab5 GTPases accumulate in the encasement, but not the papillae, suggesting that two independent pathways form these defense structures. VPS9a also mediates defense to an adapted powdery mildew fungus, thus regulating a durable type of defense that works in both host and nonhost resistance. We propose that VPS9a plays a conserved role in organizing cellular endomembrane trafficking, required for delivery of defense components in response to powdery mildew fungi. PMID:28808134

G Protein-Coupled Receptor Kinase 2 Promotes Flaviviridae Entry and Replication

PubMed Central

Le Sommer, Caroline; Barrows, Nicholas J.; Bradrick, Shelton S.; Pearson, James L.; Garcia-Blanco, Mariano A.

2012-01-01

Flaviviruses cause a wide range of severe diseases ranging from encephalitis to hemorrhagic fever. Discovery of host factors that regulate the fate of flaviviruses in infected cells could provide insight into the molecular mechanisms of infection and therefore facilitate the development of anti-flaviviral drugs. We performed genome-scale siRNA screens to discover human host factors required for yellow fever virus (YFV) propagation. Using a 2×2 siRNA pool screening format and a duplicate of the screen, we identified a high confidence list of YFV host factors. To find commonalities between flaviviruses, these candidates were compared to host factors previously identified for West Nile virus (WNV) and dengue virus (DENV). This comparison highlighted a potential requirement for the G protein-coupled receptor kinase family, GRKs, for flaviviral infection. The YFV host candidate GRK2 (also known as ADRBK1) was validated both in siRNA-mediated knockdown HuH-7 cells and in GRK−/− mouse embryonic fibroblasts. Additionally, we showed that GRK2 was required for efficient propagation of DENV and Hepatitis C virus (HCV) indicating that GRK2 requirement is conserved throughout the Flaviviridae. Finally, we found that GRK2 participates in multiple distinct steps of the flavivirus life cycle by promoting both entry and RNA synthesis. Together, our findings identified GRK2 as a novel regulator of flavivirus infection and suggest that inhibition of GRK2 function may constitute a new approach for treatment of flavivirus associated diseases. PMID:23029581

G protein-coupled receptor kinase 2 promotes flaviviridae entry and replication.

PubMed

Le Sommer, Caroline; Barrows, Nicholas J; Bradrick, Shelton S; Pearson, James L; Garcia-Blanco, Mariano A

2012-01-01

Flaviviruses cause a wide range of severe diseases ranging from encephalitis to hemorrhagic fever. Discovery of host factors that regulate the fate of flaviviruses in infected cells could provide insight into the molecular mechanisms of infection and therefore facilitate the development of anti-flaviviral drugs. We performed genome-scale siRNA screens to discover human host factors required for yellow fever virus (YFV) propagation. Using a 2 × 2 siRNA pool screening format and a duplicate of the screen, we identified a high confidence list of YFV host factors. To find commonalities between flaviviruses, these candidates were compared to host factors previously identified for West Nile virus (WNV) and dengue virus (DENV). This comparison highlighted a potential requirement for the G protein-coupled receptor kinase family, GRKs, for flaviviral infection. The YFV host candidate GRK2 (also known as ADRBK1) was validated both in siRNA-mediated knockdown HuH-7 cells and in GRK(-/-) mouse embryonic fibroblasts. Additionally, we showed that GRK2 was required for efficient propagation of DENV and Hepatitis C virus (HCV) indicating that GRK2 requirement is conserved throughout the Flaviviridae. Finally, we found that GRK2 participates in multiple distinct steps of the flavivirus life cycle by promoting both entry and RNA synthesis. Together, our findings identified GRK2 as a novel regulator of flavivirus infection and suggest that inhibition of GRK2 function may constitute a new approach for treatment of flavivirus associated diseases.

Interference of two codirectional exclusion processes in the presence of a static bottleneck: A biologically motivated model

NASA Astrophysics Data System (ADS)

Mishra, Bhavya; Chowdhury, Debashish

2017-06-01

We develop a two-species exclusion process with a distinct pair of entry and exit sites for each species of rigid rods. The relatively slower forward stepping of the rods in an extended bottleneck region, located in between the two entry sites, controls the extent of interference of the codirectional flow of the two species of rods. The relative positions of the sites of entry of the two species of rods with respect to the location of the bottleneck are motivated by a biological phenomenon. However, the primary focus of the study here is to explore the effects of the interference of the flow of the two species of rods on their spatiotemporal organization and the regulations of this interference by the extended bottleneck. By a combination of mean-field theory and computer simulation, we calculate the flux of both species of rods and their density profiles as well as the composite phase diagrams of the system. If the bottleneck is sufficiently stringent, then some of the phases become practically unrealizable, although not ruled out on the basis of any fundamental physical principle. Moreover, the extent of suppression of flow of the downstream entrants by the flow of the upstream entrants can also be regulated by the strength of the bottleneck. We speculate on the possible implications of the results in the context of the biological phenomenon that motivated the formulation of the theoretical model.

Controlled free radical attack in the apoplast: a hypothesis for roles of O, N and S species in regulatory and polysaccharide cleavage events during rapid abscission by Azolla.

PubMed

Cohen, Michael F; Gurung, Sushma; Fukuto, Jon M; Yamasaki, Hideo

2014-03-01

Shedding of organs by abscission is a key terminal step in plant development and stress responses. Cell wall (CW) loosening at the abscission zone can occur through a combination chain breakage of apoplastic polysaccharides and tension release of cellulose microfibrils. Two distinctly regulated abscission cleavage events are amenable to study in small water ferns of the genus Azolla; one is a rapid abscission induced by environmental stimuli such as heat or chemicals, and the other is an ethylene-induced process occurring more slowly through the action of hydrolytic enzymes. Although free radicals are suggested to be involved in the induction of rapid root abscission, its mechanism is not fully understood. The apoplast contains peroxidases, metal-binding proteins and phenolic compounds that potentially generate free radicals from H2O2 to cleave polysaccharides in the CW and middle lamella. Effects of various thiol-reactive agents implicate the action of apoplastic peroxidases having accessible cysteine thiols in rapid abscission. The Ca(2+) dependency of rapid abscission may reflect the stabilization Ca(2+) confers to peroxidase structure and binding to pectin. To spur further investigation, we present a hypothetical model for small signaling molecules H2O2 and NO and their derivatives in regulating, via modification of putative protein thiols, free radical attack of apoplastic polysaccharides. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Expression Patterns, Activities and Carbohydrate-Metabolizing Regulation of Sucrose Phosphate Synthase, Sucrose Synthase and Neutral Invertase in Pineapple Fruit during Development and Ripening

PubMed Central

Zhang, Xiu-Mei; Wang, Wei; Du, Li-Qing; Xie, Jiang-Hui; Yao, Yan-Li; Sun, Guang-Ming

2012-01-01

Differences in carbohydrate contents and metabolizing-enzyme activities were monitored in apical, medial, basal and core sections of pineapple (Ananas comosus cv. Comte de paris) during fruit development and ripening. Fructose and glucose of various sections in nearly equal amounts were the predominant sugars in the fruitlets, and had obvious differences until the fruit matured. The large rise of sucrose/hexose was accompanied by dramatic changes in sucrose phosphate synthase (SPS) and sucrose synthase (SuSy) activities. By contrast, neutral invertase (NI) activity may provide a mechanism to increase fruit sink strength by increasing hexose concentrations. Furthermore, two cDNAs of Ac-sps (accession no. GQ996582) and Ac-ni (accession no. GQ996581) were first isolated from pineapple fruits utilizing conserved amino-acid sequences. Homology alignment reveals that the amino acid sequences contain some conserved function domains. Transcription expression analysis of Ac-sps, Ac-susy and Ac-ni also indicated distinct patterns related to sugar accumulation and composition of pineapple fruits. It suggests that differential expressions of multiple gene families are necessary for sugar metabolism in various parts and developmental stages of pineapple fruit. A cycle of sucrose breakdown in the cytosol of sink tissues could be mediated through both Ac-SuSy and Ac-NI, and Ac-NI could be involved in regulating crucial steps by generating sugar signals to the cells in a temporally and spatially restricted fashion. PMID:22949808

Mapping the co-localization of the circadian proteins PER2 and BMAL1 with enkephalin and substance P throughout the rodent forebrain.

PubMed

Frederick, Ariana; Goldsmith, Jory; de Zavalia, Nuria; Amir, Shimon

2017-01-01

Despite rhythmic expression of clock genes being found throughout the central nervous system, very little is known about their function outside of the suprachiasmatic nucleus. Determining the pattern of clock gene expression across neuronal subpopulations is a key step in understanding their regulation and how they may influence the functions of various brain structures. Using immunofluorescence and confocal microscopy, we quantified the co-expression of the clock proteins BMAL1 and PER2 with two neuropeptides, Substance P (SubP) and Enkephalin (Enk), expressed in distinct neuronal populations throughout the forebrain. Regions examined included the limbic forebrain (dorsal striatum, nucleus accumbens, amygdala, stria terminalis), thalamus medial habenula of the thalamus, paraventricular nucleus and arcuate nucleus of the hypothalamus and the olfactory bulb. In most regions examined, BMAL1 was homogeneously expressed in nearly all neurons (~90%), and PER2 was expressed in a slightly lower proportion of cells. There was no specific correlation to SubP- or Enk- expressing subpopulations. The olfactory bulb was unique in that PER2 and BMAL1 were expressed in a much smaller percentage of cells, and Enk was rarely found in the same cells that expressed the clock proteins (SubP was undetectable). These results indicate that clock genes are not unique to specific cell types, and further studies will be required to determine the factors that contribute to the regulation of clock gene expression throughout the brain.

Soybean (Glycine max) WRINKLED1 transcription factor, GmWRI1a, positively regulates seed oil accumulation.

PubMed

Chen, Liang; Zheng, Yuhong; Dong, Zhimin; Meng, Fanfan; Sun, Xingmiao; Fan, Xuhong; Zhang, Yunfeng; Wang, Mingliang; Wang, Shuming

2018-04-01

Soybean is the world's most important leguminous crop producing high-quality protein and oil. Elevating oil accumulation in soybean seed is always many researchers' goal. WRINKLED1 (WRI1) encodes a transcription factor of the APETALA2/ethylene responsive element-binding protein (AP2/EREBP) family that plays important roles during plant seed oil accumulation. In this study, we isolated and characterized three distinct orthologues of WRI1 in soybean (Glycine max) that display different organ-specific expression patterns, among which GmWRI1a was highly expressed in maturing soybean seed. Electrophoretic mobility shift assays and yeast one-hybrid experiments demonstrated that the GmWRI1a protein was capable of binding to AW-box, a conserved sequence in the proximal upstream regions of many genes involved in various steps of oil biosynthesis. Transgenic soybean seeds overexpressing GmWRI1a under the control of the seed-specific napin promoter showed the increased total oil and fatty acid content and the changed fatty acid composition. Furthermore, basing on the activated expressions in transgenic soybean seeds and existence of AW-box element in the promoter regions, direct downstream genes of GmWRI1a were identified, and their products were responsible for fatty acid production, elongation, desaturation and export from plastid. We conclude that GmWRI1a transcription factor can positively regulate oil accumulation in soybean seed by a complex gene expression network related to fatty acid biosynthesis.

Gene regulatory and signaling networks exhibit distinct topological distributions of motifs

NASA Astrophysics Data System (ADS)

Ferreira, Gustavo Rodrigues; Nakaya, Helder Imoto; Costa, Luciano da Fontoura

2018-04-01

The biological processes of cellular decision making and differentiation involve a plethora of signaling pathways and gene regulatory circuits. These networks in turn exhibit a multitude of motifs playing crucial parts in regulating network activity. Here we compare the topological placement of motifs in gene regulatory and signaling networks and observe that it suggests different evolutionary strategies in motif distribution for distinct cellular subnetworks.

Development of a high-throughput screening system for identification of novel reagents regulating DNA damage in human dermal fibroblasts.

PubMed

Bae, Seunghee; An, In-Sook; An, Sungkwan

2015-09-01

Ultraviolet (UV) radiation is a major inducer of skin aging and accumulated exposure to UV radiation increases DNA damage in skin cells, including dermal fibroblasts. In the present study, we developed a novel DNA repair regulating material discovery (DREAM) system for the high-throughput screening and identification of putative materials regulating DNA repair in skin cells. First, we established a modified lentivirus expressing the luciferase and hypoxanthine phosphoribosyl transferase (HPRT) genes. Then, human dermal fibroblast WS-1 cells were infected with the modified lentivirus and selected with puromycin to establish cells that stably expressed luciferase and HPRT (DREAM-F cells). The first step in the DREAM protocol was a 96-well-based screening procedure, involving the analysis of cell viability and luciferase activity after pretreatment of DREAM-F cells with reagents of interest and post-treatment with UVB radiation, and vice versa. In the second step, we validated certain effective reagents identified in the first step by analyzing the cell cycle, evaluating cell death, and performing HPRT-DNA sequencing in DREAM-F cells treated with these reagents and UVB. This DREAM system is scalable and forms a time-saving high-throughput screening system for identifying novel anti-photoaging reagents regulating DNA damage in dermal fibroblasts.

Steps in the design, development and formative evaluation of obesity prevention-related behavior change trials.

PubMed

Baranowski, Tom; Cerin, Ester; Baranowski, Janice

2009-01-21

Obesity prevention interventions through dietary and physical activity change have generally not been effective. Limitations on possible program effectiveness are herein identified at every step in the mediating variable model, a generic conceptual framework for understanding how interventions may promote behavior change. To minimize these problems, and thereby enhance likely intervention effectiveness, four sequential types of formative studies are proposed: targeted behavior validation, targeted mediator validation, intervention procedure validation, and pilot feasibility intervention. Implementing these studies would establish the relationships at each step in the mediating variable model, thereby maximizing the likelihood that an intervention would work and its effects would be detected. Building consensus among researchers, funding agencies, and journal editors on distinct intervention development studies should avoid identified limitations and move the field forward.

Steps in the design, development and formative evaluation of obesity prevention-related behavior change trials

PubMed Central

Baranowski, Tom; Cerin, Ester; Baranowski, Janice

2009-01-01

Obesity prevention interventions through dietary and physical activity change have generally not been effective. Limitations on possible program effectiveness are herein identified at every step in the mediating variable model, a generic conceptual framework for understanding how interventions may promote behavior change. To minimize these problems, and thereby enhance likely intervention effectiveness, four sequential types of formative studies are proposed: targeted behavior validation, targeted mediator validation, intervention procedure validation, and pilot feasibility intervention. Implementing these studies would establish the relationships at each step in the mediating variable model, thereby maximizing the likelihood that an intervention would work and its effects would be detected. Building consensus among researchers, funding agencies, and journal editors on distinct intervention development studies should avoid identified limitations and move the field forward. PMID:19159476

One-step generation of continuous-variable quadripartite cluster states in a circuit QED system

NASA Astrophysics Data System (ADS)

Yang, Zhi-peng; Li, Zhen; Ma, Sheng-li; Li, Fu-li

2017-07-01

We propose a dissipative scheme for one-step generation of continuous-variable quadripartite cluster states in a circuit QED setup consisting of four superconducting coplanar waveguide resonators and a gap-tunable superconducting flux qubit. With external driving fields to adjust the desired qubit-resonator and resonator-resonator interactions, we show that continuous-variable quadripartite cluster states of the four resonators can be generated with the assistance of energy relaxation of the qubit. By comparison with the previous proposals, the distinct advantage of our scheme is that only one step of quantum operation is needed to realize the quantum state engineering. This makes our scheme simpler and more feasible in experiment. Our result may have useful application for implementing quantum computation in solid-state circuit QED systems.

Self-Explaining Steps in Problem-Solving Tasks to Improve Self-Regulation in Secondary Education

ERIC Educational Resources Information Center

Baars, Martine; Leopold, Claudia; Paas, Fred

2018-01-01

The ability to learn in a self-regulated way is important for adolescents' academic achievements. Monitoring one's own learning is a prerequisite skill for successful self-regulated learning. However, accurate monitoring has been found to be difficult for adolescents, especially for learning problem-solving tasks such as can be found in math and…

The NAD/NARB System: Advertising Self-Regulation at Work.

ERIC Educational Resources Information Center

Hays, Robert

Self-regulation, as defined by the National Advertising Division/National Advertising Review Board (NAD/NARB), is a process whereby the advertising industry regulates itself and turns to the federal government only if the system fails. The NAD/NARB system involves a two-step process: complaints are initially handled by the NAD and then are either…

Self-Regulation in Three Types of Online Interaction: A Scale Development

ERIC Educational Resources Information Center

Cho, Moon-Heum; Cho, YoonJung

2017-01-01

The purpose of this study was to develop a scale with which to examine students' self-regulation (SR) in three types of online interaction. Using scale development steps, we constructed the online self-regulation questionnaire (OSRQ), a self-report survey. A total of 799 online students participated in the study. Data from 400 randomly selected…

MicroRNA-Target Network Inference and Local Network Enrichment Analysis Identify Two microRNA Clusters with Distinct Functions in Head and Neck Squamous Cell Carcinoma

PubMed Central

Sass, Steffen; Pitea, Adriana; Unger, Kristian; Hess, Julia; Mueller, Nikola S.; Theis, Fabian J.

2015-01-01

MicroRNAs represent ~22 nt long endogenous small RNA molecules that have been experimentally shown to regulate gene expression post-transcriptionally. One main interest in miRNA research is the investigation of their functional roles, which can typically be accomplished by identification of mi-/mRNA interactions and functional annotation of target gene sets. We here present a novel method “miRlastic”, which infers miRNA-target interactions using transcriptomic data as well as prior knowledge and performs functional annotation of target genes by exploiting the local structure of the inferred network. For the network inference, we applied linear regression modeling with elastic net regularization on matched microRNA and messenger RNA expression profiling data to perform feature selection on prior knowledge from sequence-based target prediction resources. The novelty of miRlastic inference originates in predicting data-driven intra-transcriptome regulatory relationships through feature selection. With synthetic data, we showed that miRlastic outperformed commonly used methods and was suitable even for low sample sizes. To gain insight into the functional role of miRNAs and to determine joint functional properties of miRNA clusters, we introduced a local enrichment analysis procedure. The principle of this procedure lies in identifying regions of high functional similarity by evaluating the shortest paths between genes in the network. We can finally assign functional roles to the miRNAs by taking their regulatory relationships into account. We thoroughly evaluated miRlastic on a cohort of head and neck cancer (HNSCC) patients provided by The Cancer Genome Atlas. We inferred an mi-/mRNA regulatory network for human papilloma virus (HPV)-associated miRNAs in HNSCC. The resulting network best enriched for experimentally validated miRNA-target interaction, when compared to common methods. Finally, the local enrichment step identified two functional clusters of miRNAs that were predicted to mediate HPV-associated dysregulation in HNSCC. Our novel approach was able to characterize distinct pathway regulations from matched miRNA and mRNA data. An R package of miRlastic was made available through: http://icb.helmholtz-muenchen.de/mirlastic. PMID:26694379

MicroRNA-Target Network Inference and Local Network Enrichment Analysis Identify Two microRNA Clusters with Distinct Functions in Head and Neck Squamous Cell Carcinoma.

PubMed

Sass, Steffen; Pitea, Adriana; Unger, Kristian; Hess, Julia; Mueller, Nikola S; Theis, Fabian J

2015-12-18

MicroRNAs represent ~22 nt long endogenous small RNA molecules that have been experimentally shown to regulate gene expression post-transcriptionally. One main interest in miRNA research is the investigation of their functional roles, which can typically be accomplished by identification of mi-/mRNA interactions and functional annotation of target gene sets. We here present a novel method "miRlastic", which infers miRNA-target interactions using transcriptomic data as well as prior knowledge and performs functional annotation of target genes by exploiting the local structure of the inferred network. For the network inference, we applied linear regression modeling with elastic net regularization on matched microRNA and messenger RNA expression profiling data to perform feature selection on prior knowledge from sequence-based target prediction resources. The novelty of miRlastic inference originates in predicting data-driven intra-transcriptome regulatory relationships through feature selection. With synthetic data, we showed that miRlastic outperformed commonly used methods and was suitable even for low sample sizes. To gain insight into the functional role of miRNAs and to determine joint functional properties of miRNA clusters, we introduced a local enrichment analysis procedure. The principle of this procedure lies in identifying regions of high functional similarity by evaluating the shortest paths between genes in the network. We can finally assign functional roles to the miRNAs by taking their regulatory relationships into account. We thoroughly evaluated miRlastic on a cohort of head and neck cancer (HNSCC) patients provided by The Cancer Genome Atlas. We inferred an mi-/mRNA regulatory network for human papilloma virus (HPV)-associated miRNAs in HNSCC. The resulting network best enriched for experimentally validated miRNA-target interaction, when compared to common methods. Finally, the local enrichment step identified two functional clusters of miRNAs that were predicted to mediate HPV-associated dysregulation in HNSCC. Our novel approach was able to characterize distinct pathway regulations from matched miRNA and mRNA data. An R package of miRlastic was made available through: http://icb.helmholtz-muenchen.de/mirlastic.

Epididymal secreted protein Crisp-1 and sperm function.

PubMed

Roberts, Kenneth P; Ensrud, Kathy M; Wooters, Joseph L; Nolan, Michael A; Johnston, Daniel S; Hamilton, David W

2006-05-16

Crisp-1 is a member of the cysteine-rich secretory protein family. This family of proteins is characterized by the presence of 16 conserved cysteine residues, the characteristic from which the family name is derived. Members of the Crisp protein family are found in the secretions of the reproductive tract and salivary glands, including venom toxins from several species of snakes and lizards. The Crisp proteins are modular, each containing an amino terminal pathogenesis-related (PR)-like domain and a carboxyl terminal cysteine-rich domain (CRD) connected by a hinge region. Sequence and structural similarities to proteins with known functions suggest that the Crisp family of proteins may act by regulating cellular ion channels. Rat Crisp-1 is synthesized as two distinct isoforms (referred to as Proteins D and E) by the epididymal epithelium and both are secreted into the luminal fluid where they interact with spermatozoa. Our laboratory has correlated Crisp-1 binding to sperm with inhibiting the signaling cascades that initiate capacitation while others have shown that blocking Crisp-1 binding sites on oocytes interferes with sperm-egg fusion. We hypothesize that the D and E populations of rat Crisp-1 have different interactions with sperm that modulate these distinct biological activities. Through tandem mass spectrometry (MS/MS) and monosaccharide composition analyses, we have identified at least one difference between the D and E forms as an additional single O-linked N-acetyl galactosamine on an amino terminal threonine residue in Protein E. This post-translational modification appears to account for the unique 'E' epitope bound by monoclonal antibody 4E9 developed in our laboratory, and may also lead to differential processing and localization of Protein E on sperm, when compared to Protein D. These findings are the first step in distinguishing the molecular basis of the biological activities of the D and E forms of rat Crisp-1. The epididymal-specific expression of Crisp-1, combined with its role in regulation of sperm capacitation and oocyte interaction, make it an attractive target for post-testicular contraceptive development.

A Comprehensive Approach to Sequence-oriented IsomiR annotation (CASMIR): demonstration with IsomiR profiling in colorectal neoplasia.

PubMed

Wu, Chung Wah; Evans, Jared M; Huang, Shengbing; Mahoney, Douglas W; Dukek, Brian A; Taylor, William R; Yab, Tracy C; Smyrk, Thomas C; Jen, Jin; Kisiel, John B; Ahlquist, David A

2018-05-25

MicroRNA (miRNA) profiling is an important step in studying biological associations and identifying marker candidates. miRNA exists in isoforms, called isomiRs, which may exhibit distinct properties. With conventional profiling methods, limitations in assay and analysis platforms may compromise isomiR interrogation. We introduce a comprehensive approach to sequence-oriented isomiR annotation (CASMIR) to allow unbiased identification of global isomiRs from small RNA sequencing data. In this approach, small RNA reads are maintained as independent sequences instead of being summarized under miRNA names. IsomiR features are identified through step-wise local alignment against canonical forms and precursor sequences. Through customizing the reference database, CASMIR is applicable to isomiR annotation across species. To demonstrate its application, we investigated isomiR profiles in normal and neoplastic human colorectal epithelia. We also ran miRDeep2, a popular miRNA analysis algorithm to validate isomiRs annotated by CASMIR. With CASMIR, specific and biologically relevant isomiR patterns could be identified. We note that specific isomiRs are often more abundant than their canonical forms. We identify isomiRs that are commonly up-regulated in both colorectal cancer and advanced adenoma, and illustrate advantages in targeting isomiRs as potential biomarkers over canonical forms. Studying miRNAs at the isomiR level could reveal new insight into miRNA biology and inform assay design for specific isomiRs. CASMIR facilitates comprehensive annotation of isomiR features in small RNA sequencing data for isomiR profiling and differential expression analysis.

Venous-derived angioblasts generate organ-specific vessels during zebrafish embryonic development.

PubMed

Hen, Gideon; Nicenboim, Julian; Mayseless, Oded; Asaf, Lihee; Shin, Masahiro; Busolin, Giorgia; Hofi, Roy; Almog, Gabriella; Tiso, Natascia; Lawson, Nathan D; Yaniv, Karina

2015-12-15

Formation and remodeling of vascular beds are complex processes orchestrated by multiple signaling pathways. Although it is well accepted that vessels of a particular organ display specific features that enable them to fulfill distinct functions, the embryonic origins of tissue-specific vessels and the molecular mechanisms regulating their formation are poorly understood. The subintestinal plexus of the zebrafish embryo comprises vessels that vascularize the gut, liver and pancreas and, as such, represents an ideal model in which to investigate the early steps of organ-specific vessel formation. Here, we show that both arterial and venous components of the subintestinal plexus originate from a pool of specialized angioblasts residing in the floor of the posterior cardinal vein (PCV). Using live imaging of zebrafish embryos, in combination with photoconvertable transgenic reporters, we demonstrate that these angioblasts undergo two phases of migration and differentiation. Initially, a subintestinal vein forms and expands ventrally through a Bone Morphogenetic Protein-dependent step of collective migration. Concomitantly, a Vascular Endothelial Growth Factor-dependent shift in the directionality of migration, coupled to the upregulation of arterial markers, is observed, which culminates with the generation of the supraintestinal artery. Together, our results establish the zebrafish subintestinal plexus as an advantageous model for the study of organ-specific vessel development and provide new insights into the molecular mechanisms controlling its formation. More broadly, our findings suggest that PCV-specialized angioblasts contribute not only to the formation of the early trunk vasculature, but also to the establishment of late-forming, tissue-specific vascular beds. © 2015. Published by The Company of Biologists Ltd.

A Systems Approach towards an Intelligent and Self-Controlling Platform for Integrated Continuous Reaction Sequences**

PubMed Central

Ingham, Richard J; Battilocchio, Claudio; Fitzpatrick, Daniel E; Sliwinski, Eric; Hawkins, Joel M; Ley, Steven V

2015-01-01

Performing reactions in flow can offer major advantages over batch methods. However, laboratory flow chemistry processes are currently often limited to single steps or short sequences due to the complexity involved with operating a multi-step process. Using new modular components for downstream processing, coupled with control technologies, more advanced multi-step flow sequences can be realized. These tools are applied to the synthesis of 2-aminoadamantane-2-carboxylic acid. A system comprising three chemistry steps and three workup steps was developed, having sufficient autonomy and self-regulation to be managed by a single operator. PMID:25377747

29 CFR 100.560 - Communications.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 29 Labor 2 2012-07-01 2012-07-01 false Communications. 100.560 Section 100.560 Labor Regulations Relating to Labor NATIONAL LABOR RELATIONS BOARD ADMINISTRATIVE REGULATIONS Enforcement of... § 100.560 Communications. (a) The agency shall take appropriate steps to ensure effective communication...