Kinesin Khc-73/KIF13B modulates retrograde BMP signaling by influencing endosomal dynamics at the Drosophila neuromuscular junction.

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Liao, Edward H; Gray, Lindsay; Tsurudome, Kazuya; El-Mounzer, Wassim; Elazzouzi, Fatima; Baim, Christopher; Farzin, Sarah; Calderon, Mario R; Kauwe, Grant; Haghighi, A Pejmun

2018-01-01

Retrograde signaling is essential for neuronal growth, function and survival; however, we know little about how signaling endosomes might be directed from synaptic terminals onto retrograde axonal pathways. We have identified Khc-73, a plus-end directed microtubule motor protein, as a regulator of sorting of endosomes in Drosophila larval motor neurons. The number of synaptic boutons and the amount of neurotransmitter release at the Khc-73 mutant larval neuromuscular junction (NMJ) are normal, but we find a significant decrease in the number of presynaptic release sites. This defect in Khc-73 mutant larvae can be genetically enhanced by a partial genetic loss of Bone Morphogenic Protein (BMP) signaling or suppressed by activation of BMP signaling in motoneurons. Consistently, activation of BMP signaling that normally enhances the accumulation of phosphorylated form of BMP transcription factor Mad in the nuclei, can be suppressed by genetic removal of Khc-73. Using a number of assays including live imaging in larval motor neurons, we show that loss of Khc-73 curbs the ability of retrograde-bound endosomes to leave the synaptic area and join the retrograde axonal pathway. Our findings identify Khc-73 as a regulator of endosomal traffic at the synapse and modulator of retrograde BMP signaling in motoneurons.

The Endosome-associated Deubiquitinating Enzyme USP8 Regulates BACE1 Enzyme Ubiquitination and Degradation.

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Yeates, Eniola Funmilayo Aduke; Tesco, Giuseppina

2016-07-22

The β-site amyloid precursor protein-cleaving enzyme (BACE1) is the rate-limiting enzyme in the production of amyloid-β, the toxic peptide that accumulates in the brain of subjects affected by Alzheimer disease. Our previous studies have shown that BACE1 is degraded via the lysosomal pathway and that that depletion of the trafficking molecule Golgi-localized γ-ear-containing ARF-binding protein 3 (GGA3) results in increased BACE1 levels and activity because of impaired lysosomal degradation. We also determined that GGA3 regulation of BACE1 levels requires its ability to bind ubiquitin. Accordingly, we reported that BACE1 is ubiquitinated at lysine 501 and that lack of ubiquitination at lysine 501 produces BACE1 stabilization. Ubiquitin conjugation is a reversible process mediated by deubiquitinating enzymes. The ubiquitin-specific peptidase 8 (USP8), an endosome-associated deubiquitinating enzyme, regulates the ubiquitination, trafficking, and lysosomal degradation of several plasma membrane proteins. Here, we report that RNAi-mediated depletion of USP8 reduced levels of both ectopically expressed and endogenous BACE1 in H4 human neuroglioma cells. Moreover, USP8 depletion increased BACE1 ubiquitination, promoted BACE1 accumulation in the early endosomes and late endosomes/lysosomes, and decreased levels of BACE1 in the recycling endosomes. We also found that decreased BACE1 protein levels were accompanied by a decrease in BACE1-mediated amyloid precursor protein cleavage and amyloid-β levels. Our findings demonstrate that USP8 plays a key role in the trafficking and degradation of BACE1 by deubiquitinating lysine 501. These studies suggest that therapies able to accelerate BACE1 degradation (e.g. by increasing BACE1 ubiquitination) may represent a potential treatment for Alzheimer disease. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

The Endosome-associated Deubiquitinating Enzyme USP8 Regulates BACE1 Enzyme Ubiquitination and Degradation*

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Yeates, Eniola Funmilayo Aduke; Tesco, Giuseppina

2016-01-01

The β-site amyloid precursor protein-cleaving enzyme (BACE1) is the rate-limiting enzyme in the production of amyloid-β, the toxic peptide that accumulates in the brain of subjects affected by Alzheimer disease. Our previous studies have shown that BACE1 is degraded via the lysosomal pathway and that that depletion of the trafficking molecule Golgi-localized γ-ear-containing ARF-binding protein 3 (GGA3) results in increased BACE1 levels and activity because of impaired lysosomal degradation. We also determined that GGA3 regulation of BACE1 levels requires its ability to bind ubiquitin. Accordingly, we reported that BACE1 is ubiquitinated at lysine 501 and that lack of ubiquitination at lysine 501 produces BACE1 stabilization. Ubiquitin conjugation is a reversible process mediated by deubiquitinating enzymes. The ubiquitin-specific peptidase 8 (USP8), an endosome-associated deubiquitinating enzyme, regulates the ubiquitination, trafficking, and lysosomal degradation of several plasma membrane proteins. Here, we report that RNAi-mediated depletion of USP8 reduced levels of both ectopically expressed and endogenous BACE1 in H4 human neuroglioma cells. Moreover, USP8 depletion increased BACE1 ubiquitination, promoted BACE1 accumulation in the early endosomes and late endosomes/lysosomes, and decreased levels of BACE1 in the recycling endosomes. We also found that decreased BACE1 protein levels were accompanied by a decrease in BACE1-mediated amyloid precursor protein cleavage and amyloid-β levels. Our findings demonstrate that USP8 plays a key role in the trafficking and degradation of BACE1 by deubiquitinating lysine 501. These studies suggest that therapies able to accelerate BACE1 degradation (e.g. by increasing BACE1 ubiquitination) may represent a potential treatment for Alzheimer disease. PMID:27302062

Neurotrophin signaling endosomes; biogenesis, regulation, and functions

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Yamashita, Naoya; Kuruvilla, Rejji

2016-01-01

In the nervous system, communication between neurons and their post-synaptic target cells is critical for the formation, refinement and maintenance of functional neuronal connections. Diffusible signals secreted by target tissues, exemplified by the family of neurotrophins, impinge on nerve terminals to influence diverse developmental events including neuronal survival and axonal growth. Key mechanisms of action of target-derived neurotrophins include the cell biological processes of endocytosis and retrograde trafficking of their Trk receptors from growth cones to cell bodies. In this review, we summarize the molecular mechanisms underlying this endosome-mediated signaling, focusing on the instructive role of neurotrophin signaling itself in directing its own trafficking. Recent studies have linked impaired neurotrophin trafficking to neurodevelopmental disorders, highlighting the relevance of neurotrophin endosomes in human health. PMID:27327126

PIKfyve mediates the motility of late endosomes and lysosomes in neuronal dendrites.

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Tsuruta, Fuminori; Dolmetsch, Ricardo E

2015-09-25

The endosome/lysosome system in the nervous system is critically important for a variety of neuronal functions such as neurite outgrowth, retrograde transport, and synaptic plasticity. In neurons, the endosome/lysosome system is crucial for the activity-dependent internalization of membrane proteins and contributes to the regulation of lipid level on the plasma membrane. Although homeostasis of membrane dynamics plays important roles in the properties of central nervous systems, it has not been elucidated how endosome/lysosome system is regulated. Here, we report that phosphatidylinositol 3-phosphate 5-kinase (PIKfyve) mediates the motility of late endosomes and lysosomes in neuronal dendrites. Endosomes and lysosomes are highly motile in resting neurons, however knockdown of PIKfyve led to a significant reduction in late endosomes and lysosomes motility. We also found that vesicle acidification is crucial for their motility and PIKfyve is associated with this process indirectly. These data suggest that PIKfyve mediates vesicle motility through the regulation of vesicle integrity in neurons. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Heterotrimeric G protein subunits are located on rat liver endosomes

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Van Dyke Rebecca W

2004-01-01

Full Text Available Abstract Background Rat liver endosomes contain activated insulin receptors and downstream signal transduction molecules. We undertook these studies to determine whether endosomes also contain heterotrimeric G proteins that may be involved in signal transduction from G protein-coupled receptors. Results By Western blotting Gsα, Giα1,2, Giα3 and Gβ were enriched in both canalicular (CM and basolateral (BLM membranes but also readily detectable on three types of purified rat liver endosomes in the order recycling receptor compartment (RRC > compartment for uncoupling of receptor and ligand (CURL > multivesicular bodies (MVB >> purified secondary lysosomes. Western blotting with antibodies to Na, K-ATPase and to other proteins associated with plasma membranes and intracellular organelles indicated this was not due to contamination of endosome preparations by CM or BLM. Adenylate cyclase (AC was also identified on purified CM, BLM, RRC, CURL and MVB. Percoll gradient fractionation of liver postnuclear supernatants demonstrated co-occurrence of endosomes and heterotrimeric G protein subunits in fractions with little plasma membrane markers. By confocal microscopy, punctate staining for Gsα, Giα3 and Gβ corresponded to punctate areas of endocytosed Texas red-dextran in hepatocytes from control and cholera toxin-treated livers. Conclusion We conclude that heterotrimeric G protein subunits as well as AC likely traffic into hepatocytes on endosome membranes, possibly generating downstream signals spatially separate from signalling generated at the plasma membrane, analogous to the role(s of internalized insulin receptors.

Tetraspanin CD63 Bridges Autophagic and Endosomal Processes To Regulate Exosomal Secretion and Intracellular Signaling of Epstein-Barr Virus LMP1

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Hurwitz, Stephanie N; Cheerathodi, Mujeeb R; Nkosi, Dingani; York, Sara B; Meckes, David G

2018-03-01

The tetraspanin protein CD63 has been recently described as a key factor in extracellular vesicle (EV) production and endosomal cargo sorting. In the context of Epstein-Barr virus (EBV) infection, CD63 is required for the efficient packaging of the major viral oncoprotein latent membrane protein 1 (LMP1) into exosomes and other EV populations and acts as a negative regulator of LMP1 intracellular signaling. Accumulating evidence has also pointed to intersections of the endosomal and autophagy pathways in maintaining cellular secretory processes and as sites for viral assembly and replication. Indeed, LMP1 can activate the mammalian target of rapamycin (mTOR) pathway to suppress host cell autophagy and facilitate cell growth and proliferation. Despite the growing recognition of cross talk between endosomes and autophagosomes and its relevance to viral infection, little is understood about the molecular mechanisms governing endosomal and autophagy convergence. Here, we demonstrate that CD63-dependent vesicle protein secretion directly opposes intracellular signaling activation downstream of LMP1, including mTOR-associated proteins. Conversely, disruption of normal autolysosomal processes increases LMP1 secretion and dampens signal transduction by the viral protein. Increases in mTOR activation following CD63 knockout are coincident with the development of serum-dependent autophagic vacuoles that are acidified in the presence of high LMP1 levels. Altogether, these findings suggest a key role of CD63 in regulating the interactions between endosomal and autophagy processes and limiting cellular signaling activity in both noninfected and virally infected cells. IMPORTANCE The close connection between extracellular vesicles and viruses is becoming rapidly and more widely appreciated. EBV, a human gamma herpesvirus that contributes to the progression of a multitude of lymphomas and carcinomas in immunocompromised or genetically susceptible populations, packages its major

Virtual Reality Game Education to Learn Traffic Regulation

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Andru Deva Lukito

2017-10-01

Full Text Available Abstract – Traffic accident has become number 3 of children death cause in the world according to WHO[1]. Traffic accident that involve children often caused by their own by breaking the law or regulation. Therefor education about traffic regulation and law including traffic sign and its meaning must be given to children early. Because education means process to change a person or a group attitude and behavior in order to make them mature through teaching and training [2]. One of them that can be used is digital media.  One of interactive digital media is digital game, various form of digital game start from 2D, 2.5D, 3D with many point of view and new technology. VR (Virtual Reality as new digital media where alternate reality exist to test various theory without any real consequences, according to Greenbaum “Virtual Reality is an alternate world filled with computer-generated images that respond to human movements. These simulated environments are usually visited with the aid of an expensive data suit which features stereophonic video goggles and fiber-optic data gloves”[3]. Greenbaum statement before were make VR suitable to test traffic law and regulation and educate kid to obey the traffic sign and regulation without real consequences from real world. This Journal contain the result of using virtual reality as traffic regulation education media. Education material that arranged consisting traffic sign that appear on the road and safety riding gear. Keywords – Virtual Reality, Traffic sign, Road traffic, children, education

Protein complexes and cholesterol in the control of late endosomal dynamicsCholesterol and multi-protein complexes in the control of late endosomal dynamics

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Kant, Rik Henricus Nicolaas van der

2013-01-01

Late endosomal transport is disrupted in several diseases such as Niemann-Pick type C, ARC syndrome and Alzheimer’s disease. This thesis describes the regulation of late endosomal dynamics by cholesterol and multi-protein complexes. We find that cholesterol acts as a cellular tomtom that steers the

Erythroid cell mitochondria receive endosomal iron by a "kiss-and-run" mechanism.

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Hamdi, Amel; Roshan, Tariq M; Kahawita, Tanya M; Mason, Anne B; Sheftel, Alex D; Ponka, Prem

2016-12-01

In erythroid cells, more than 90% of transferrin-derived iron enters mitochondria where ferrochelatase inserts Fe 2+ into protoporphyrin IX. However, the path of iron from endosomes to mitochondrial ferrochelatase remains elusive. The prevailing opinion is that, after its export from endosomes, the redox-active metal spreads into the cytosol and mysteriously finds its way into mitochondria through passive diffusion. In contrast, this study supports the hypothesis that the highly efficient transport of iron toward ferrochelatase in erythroid cells requires a direct interaction between transferrin-endosomes and mitochondria (the "kiss-and-run" hypothesis). Using a novel method (flow sub-cytometry), we analyze lysates of reticulocytes after labeling these organelles with different fluorophores. We have identified a double-labeled population definitively representing endosomes interacting with mitochondria, as demonstrated by confocal microscopy. Moreover, we conclude that this endosome-mitochondrion association is reversible, since a "chase" with unlabeled holotransferrin causes a time-dependent decrease in the size of the double-labeled population. Importantly, the dissociation of endosomes from mitochondria does not occur in the absence of holotransferrin. Additionally, mutated recombinant holotransferrin, that cannot release iron, significantly decreases the uptake of 59 Fe by reticulocytes and diminishes 59 Fe incorporation into heme. This suggests that endosomes, which are unable to provide iron to mitochondria, cause a "traffic jam" leading to decreased endocytosis of holotransferrin. Altogether, our results suggest that a molecular mechanism exists to coordinate the iron status of endosomal transferrin with its trafficking. Besides its contribution to the field of iron metabolism, this study provides evidence for a new intracellular trafficking pathway of organelles. Copyright © 2016 Elsevier B.V. All rights reserved.

Virtual Reality Game Education to Learn Traffic Regulation

OpenAIRE

Andru Deva Lukito

2017-01-01

Abstract – Traffic accident has become number 3 of children death cause in the world according to WHO[1]. Traffic accident that involve children often caused by their own by breaking the law or regulation. Therefor education about traffic regulation and law including traffic sign and its meaning must be given to children early. Because education means process to change a person or a group attitude and behavior in order to make them mature through teaching and training [2]. One of them that ca...

Gαs regulates Glucagon-Like Peptide 1 Receptor-mediated cyclic AMP generation at Rab5 endosomal compartment.

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Girada, Shravan Babu; Kuna, Ramya S; Bele, Shilpak; Zhu, Zhimeng; Chakravarthi, N R; DiMarchi, Richard D; Mitra, Prasenjit

2017-10-01

Upon activation, G protein coupled receptors (GPCRs) associate with heterotrimeric G proteins at the plasma membrane to initiate second messenger signaling. Subsequently, the activated receptor experiences desensitization, internalization, and recycling back to the plasma membrane, or it undergoes lysosomal degradation. Recent reports highlight specific cases of persistent cyclic AMP generation by internalized GPCRs, although the functional significance and mechanistic details remain to be defined. Cyclic AMP generation from internalized Glucagon-Like Peptide-1 Receptor (GLP-1R) has previously been reported from our laboratory. This study aimed at deciphering the molecular mechanism by which internalized GLP-R supports sustained cyclic AMP generation upon receptor activation in pancreatic beta cells. We studied the time course of cyclic AMP generation following GLP-1R activation with particular emphasis on defining the location where cyclic AMP is generated. Detection involved a novel GLP-1 conjugate coupled with immunofluorescence using specific endosomal markers. Finally, we employed co-immunoprecipitation as well as immunofluorescence to assess the protein-protein interactions that regulate GLP-1R mediated cyclic AMP generation at endosomes. Our data reveal that prolonged association of G protein α subunit Gαs with activated GLP-1R contributed to sustained cyclic AMP generation at Rab 5 endosomal compartment. The findings provide the mechanism of endosomal cyclic AMP generation following GLP-1R activation. We identified the specific compartment that serves as an organizing center to generate endosomal cyclic AMP by internalized activated receptor complex. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

Characterization of the Mammalian CORVET and HOPS Complexes and Their Modular Restructuring for Endosome Specificity

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van der Kant, Rik; Jonker, Caspar T. H.; Wijdeven, Ruud H.; Bakker, Jeroen; Janssen, Lennert; Klumperman, Judith; Neefjes, Jacques

2015-01-01

Trafficking of cargo through the endosomal system depends on endosomal fusion events mediated by SNARE proteins, Rab-GTPases, and multisubunit tethering complexes. The CORVET and HOPS tethering complexes, respectively, regulate early and late endosomal tethering and have been characterized in detail

Late endosomal cholesterol accumulation leads to impaired intra-endosomal trafficking.

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Komla Sobo

Full Text Available BACKGROUND: Pathological accumulation of cholesterol in late endosomes is observed in lysosomal storage diseases such as Niemann-Pick type C. We here analyzed the effects of cholesterol accumulation in NPC cells, or as phenocopied by the drug U18666A, on late endosomes membrane organization and dynamics. METHODOLOGY/PRINCIPAL FINDINGS: Cholesterol accumulation did not lead to an increase in the raft to non-raft membrane ratio as anticipated. Strikingly, we observed a 2-3 fold increase in the size of the compartment. Most importantly, properties and dynamics of late endosomal intralumenal vesicles were altered as revealed by reduced late endosomal vacuolation induced by the mutant pore-forming toxin ASSP, reduced intoxication by the anthrax lethal toxin and inhibition of infection by the Vesicular Stomatitis Virus. CONCLUSIONS/SIGNIFICANCE: These results suggest that back fusion of intralumenal vesicles with the limiting membrane of late endosomes is dramatically perturbed upon cholesterol accumulation.

First-Generation Antipsychotic Haloperidol Alters the Functionality of the Late Endosomal/Lysosomal Compartment in Vitro.

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Canfrán-Duque, Alberto; Barrio, Luis C; Lerma, Milagros; de la Peña, Gema; Serna, Jorge; Pastor, Oscar; Lasunción, Miguel A; Busto, Rebeca

2016-03-18

First- and second-generation antipsychotics (FGAs and SGAs, respectively), have the ability to inhibit cholesterol biosynthesis and also to interrupt the intracellular cholesterol trafficking, interfering with low-density lipoprotein (LDL)-derived cholesterol egress from late endosomes/lysosomes. In the present work, we examined the effects of FGA haloperidol on the functionality of late endosomes/lysosomes in vitro. In HepG2 hepatocarcinoma cells incubated in the presence of 1,1'-dioctadecyl-3,3,3,3'-tetramethylindocarbocyanineperchlorate (DiI)-LDL, treatment with haloperidol caused the enlargement of organelles positive for late endosome markers lysosome-associated membrane protein 2 (LAMP-2) and LBPA (lysobisphosphatidic acid), which also showed increased content of both free-cholesterol and DiI derived from LDL. This indicates the accumulation of LDL-lipids in the late endosomal/lysosomal compartment caused by haloperidol. In contrast, LDL traffic through early endosomes and the Golgi apparatus appeared to be unaffected by the antipsychotic as the distribution of both early endosome antigen 1 (EEA1) and coatomer subunit β (β-COP) were not perturbed. Notably, treatment with haloperidol significantly increased the lysosomal pH and decreased the activities of lysosomal protease and β-d-galactosidase in a dose-dependent manner. We conclude that the alkalinization of the lysosomes' internal milieu induced by haloperidol affects lysosomal functionality.

Regulation of P2Y1 receptor traffic by sorting Nexin 1 is retromer independent.

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Nisar, Shaista; Kelly, Eamonn; Cullen, Pete J; Mundell, Stuart J

2010-04-01

The activity and traffic of G-protein coupled receptors (GPCRs) is tightly controlled. Recent work from our laboratory has shown that P2Y(1) and P2Y(12) responsiveness is rapidly and reversibly modulated in human platelets and that the underlying mechanism requires receptor trafficking as an essential part of this process. However, little is known about the molecular mechanisms underlying P2Y receptor traffic. Sorting nexin 1 (SNX1) has been shown to regulate the endosomal sorting of cell surface receptors either to lysosomes where they are downregulated or back to the cell surface. These functions may in part be due to interactions of SNX1 with the mammalian retromer complex. In this study, we investigated the role of SNX1 in P2Y receptor trafficking. We show that P2Y(1) receptors recycle via a slow recycling pathway that is regulated by SNX1, whereas P2Y(12) receptors return to the cell surface via a rapid route that is SNX1 independent. SNX1 inhibition caused a dramatic increase in the rate of P2Y(1) receptor recycling, whereas inhibition of Vps26 and Vps35 known to be present in retromer had no effect, indicating that SNX1 regulation of P2Y(1) receptor recycling is retromer independent. In addition, inhibition of SNX4, 6 and 17 proteins did not affect P2Y(1) receptor recycling. SNX1 has also been implicated in GPCR degradation; however, we provide evidence that P2Y receptor degradation is SNX1 independent. These data describe a novel function of SNX1 in the regulation of P2Y(1) receptor recycling and suggest that SNX1 plays multiple roles in endocytic trafficking of GPCRs.

Endosomal protein sorting and autophagy genes contribute to the regulation of yeast life span.

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Longo, Valter D; Nislow, Corey; Fabrizio, Paola

2010-11-01

Accumulating evidence from various organisms points to a role for autophagy in the regulation of life span. By performing a genome-wide screen to identify novel life span determinants in Saccharomyces cerevisiae, we have obtained further insights into the autophagy-related and -unrelated degradation processes that may be important for preventing cellular senescence. The generation of multivesicular bodies and their fusion with the vacuole in the endosomal pathway emerged as novel cell functions involved in yeast chronological survival and longevity extension.

ER network homeostasis is critical for plant endosome streaming and endocytosis

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Stefano, Giovanni; Renna, Luciana; Lai, YaShiuan; Slabaugh, Erin; Mannino, Nicole; Buono, Rafael A; Otegui, Marisa S; Brandizzi, Federica

2015-01-01

Eukaryotic cells internalize cargo at the plasma membrane via endocytosis, a vital process that is accomplished through a complex network of endosomal organelles. In mammalian cells, the ER is in close association with endosomes and regulates their fission. Nonetheless, the physiological role of such interaction on endocytosis is yet unexplored. Here, we probed the existence of ER–endosome association in plant cells and assayed its physiological role in endocytosis. Through live-cell imaging and electron microscopy studies, we established that endosomes are extensively associated with the plant ER, supporting conservation of interaction between heterotypic organelles in evolutionarily distant kingdoms. Furthermore, by analyzing ER–endosome dynamics in genetic backgrounds with defects in ER structure and movement, we also established that the ER network integrity is necessary for homeostasis of the distribution and streaming of various endosome populations as well as for efficient endocytosis. These results support a novel model that endocytosis homeostasis depends on a spatiotemporal control of the endosome dynamics dictated by the ER membrane network. PMID:27462431

Basolateral Endocytic Recycling Requires RAB-10 and AMPH-1 Mediated Recruitment of RAB-5 GAP TBC-2 to Endosomes

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Liu, Ou; Grant, Barth D.

2015-01-01

The small GTPase RAB-5/Rab5 is a master regulator of the early endosome, required for a myriad of coordinated activities, including the degradation and recycling of internalized cargo. Here we focused on the recycling function of the early endosome and the regulation of RAB-5 by GAP protein TBC-2 in the basolateral C. elegans intestine. We demonstrate that downstream basolateral recycling regulators, GTPase RAB-10/Rab10 and BAR domain protein AMPH-1/Amphiphysin, bind to TBC-2 and help to recruit it to endosomes. In the absence of RAB-10 or AMPH-1 binding to TBC-2, RAB-5 membrane association is abnormally high and recycling cargo is trapped in early endosomes. Furthermore, the loss of TBC-2 or AMPH-1 leads to abnormally high spatial overlap of RAB-5 and RAB-10. Taken together our results indicate that RAB-10 and AMPH-1 mediated down-regulation of RAB-5 is an important step in recycling, required for cargo exit from early endosomes and regulation of early endosome–recycling endosome interactions. PMID:26393361

An Essential Role of Hrs/Vps27 in Endosomal Cholesterol Trafficking

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Ximing Du

2012-01-01

Full Text Available The endosomal sorting complex required for transport (ESCRT plays a crucial role in the degradation of ubiquitinated endosomal membrane proteins. Here, we report that Hrs, a key protein of the ESCRT-0 complex, is required for the transport of low-density lipoprotein-derived cholesterol from endosomes to the endoplasmic reticulum. This function of Hrs in cholesterol transport is distinct from its previously defined role in lysosomal sorting and downregulation of membrane receptors via the ESCRT pathway. In line with this, knocking down other ESCRT proteins does not cause prominent endosomal cholesterol accumulation. Importantly, the localization and biochemical properties of key cholesterol-sorting proteins, NPC1 and NPC2, appear to be unchanged upon Hrs knockdown. Our data identify Hrs as a regulator of endosomal cholesterol trafficking and provide additional insights into the budding of intralumenal vesicles.

Cholesterol Regulates Syntaxin 6 Trafficking at trans-Golgi Network Endosomal Boundaries

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Meritxell Reverter

2014-05-01

Full Text Available Inhibition of cholesterol export from late endosomes causes cellular cholesterol imbalance, including cholesterol depletion in the trans-Golgi network (TGN. Here, using Chinese hamster ovary (CHO Niemann-Pick type C1 (NPC1 mutant cell lines and human NPC1 mutant fibroblasts, we show that altered cholesterol levels at the TGN/endosome boundaries trigger Syntaxin 6 (Stx6 accumulation into VAMP3, transferrin, and Rab11-positive recycling endosomes (REs. This increases Stx6/VAMP3 interaction and interferes with the recycling of αVβ3 and α5β1 integrins and cell migration, possibly in a Stx6-dependent manner. In NPC1 mutant cells, restoration of cholesterol levels in the TGN, but not inhibition of VAMP3, restores the steady-state localization of Stx6 in the TGN. Furthermore, elevation of RE cholesterol is associated with increased amounts of Stx6 in RE. Hence, the fine-tuning of cholesterol levels at the TGN-RE boundaries together with a subset of cholesterol-sensitive SNARE proteins may play a regulatory role in cell migration and invasion.

First-Generation Antipsychotic Haloperidol Alters the Functionality of the Late Endosomal/Lysosomal Compartment in Vitro

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Alberto Canfrán-Duque

2016-03-01

Full Text Available First- and second-generation antipsychotics (FGAs and SGAs, respectively, have the ability to inhibit cholesterol biosynthesis and also to interrupt the intracellular cholesterol trafficking, interfering with low-density lipoprotein (LDL-derived cholesterol egress from late endosomes/lysosomes. In the present work, we examined the effects of FGA haloperidol on the functionality of late endosomes/lysosomes in vitro. In HepG2 hepatocarcinoma cells incubated in the presence of 1,1′-dioctadecyl-3,3,3,3′-tetramethylindocarbocyanineperchlorate (DiI-LDL, treatment with haloperidol caused the enlargement of organelles positive for late endosome markers lysosome-associated membrane protein 2 (LAMP-2 and LBPA (lysobisphosphatidic acid, which also showed increased content of both free-cholesterol and DiI derived from LDL. This indicates the accumulation of LDL-lipids in the late endosomal/lysosomal compartment caused by haloperidol. In contrast, LDL traffic through early endosomes and the Golgi apparatus appeared to be unaffected by the antipsychotic as the distribution of both early endosome antigen 1 (EEA1 and coatomer subunit β (β-COP were not perturbed. Notably, treatment with haloperidol significantly increased the lysosomal pH and decreased the activities of lysosomal protease and β-d-galactosidase in a dose-dependent manner. We conclude that the alkalinization of the lysosomes’ internal milieu induced by haloperidol affects lysosomal functionality.

First-Generation Antipsychotic Haloperidol Alters the Functionality of the Late Endosomal/Lysosomal Compartment in Vitro

Science.gov (United States)

Canfrán-Duque, Alberto; Barrio, Luis C.; Lerma, Milagros; de la Peña, Gema; Serna, Jorge; Pastor, Oscar; Lasunción, Miguel A.; Busto, Rebeca

2016-01-01

First- and second-generation antipsychotics (FGAs and SGAs, respectively), have the ability to inhibit cholesterol biosynthesis and also to interrupt the intracellular cholesterol trafficking, interfering with low-density lipoprotein (LDL)-derived cholesterol egress from late endosomes/lysosomes. In the present work, we examined the effects of FGA haloperidol on the functionality of late endosomes/lysosomes in vitro. In HepG2 hepatocarcinoma cells incubated in the presence of 1,1′-dioctadecyl-3,3,3,3′-tetramethylindocarbocyanineperchlorate (DiI)-LDL, treatment with haloperidol caused the enlargement of organelles positive for late endosome markers lysosome-associated membrane protein 2 (LAMP-2) and LBPA (lysobisphosphatidic acid), which also showed increased content of both free-cholesterol and DiI derived from LDL. This indicates the accumulation of LDL-lipids in the late endosomal/lysosomal compartment caused by haloperidol. In contrast, LDL traffic through early endosomes and the Golgi apparatus appeared to be unaffected by the antipsychotic as the distribution of both early endosome antigen 1 (EEA1) and coatomer subunit β (β-COP) were not perturbed. Notably, treatment with haloperidol significantly increased the lysosomal pH and decreased the activities of lysosomal protease and β-d-galactosidase in a dose-dependent manner. We conclude that the alkalinization of the lysosomes’ internal milieu induced by haloperidol affects lysosomal functionality. PMID:26999125

TLR2 ligands induce NF-κB activation from endosomal compartments of human monocytes.

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Karim J Brandt

Full Text Available Localization of Toll-like receptors (TLR in subcellular organelles is a major strategy to regulate innate immune responses. While TLR4, a cell-surface receptor, signals from both the plasma membrane and endosomal compartments, less is known about the functional role of endosomal trafficking upon TLR2 signaling. Here we show that the bacterial TLR2 ligands Pam3CSK4 and LTA activate NF-κB-dependent signaling from endosomal compartments in human monocytes and in a NF-κB sensitive reporter cell line, despite the expression of TLR2 at the cell surface. Further analyses indicate that TLR2-induced NF-κB activation is controlled by a clathrin/dynamin-dependent endocytosis mechanism, in which CD14 serves as an important upstream regulator. These findings establish that internalization of cell-surface TLR2 into endosomal compartments is required for NF-κB activation. These observations further demonstrate the need of endocytosis in the activation and regulation of TLR2-dependent signaling pathways.

Cholesterol transfer at endosomal-organelle membrane contact sites.

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Ridgway, Neale D; Zhao, Kexin

2018-06-01

Cholesterol is delivered to the limiting membrane of late endosomes by Niemann-Pick Type C1 and C2 proteins. This review summarizes recent evidence that cholesterol transfer from endosomes to the endoplasmic reticulum and other organelles is mediated by lipid-binding proteins that localize to membrane contact sites (MCS). LDL-cholesterol in the late endosomal/lysosomes is exported to the plasma membrane, where most cholesterol resides, and the endoplasmic reticulum, which harbors the regulatory complexes and enzymes that control the synthesis and esterification of cholesterol. A major advance in dissecting these cholesterol transport pathways was identification of frequent and dynamic MCS between endosomes and the endoplasmic reticulum, peroxisomes and plasma membrane. Positioned at these MCS are members of the oxysterol-binding protein (OSBP) and steroidogenic acute regulatory protein-related lipid-transfer family of lipid transfer proteins that bridge the opposing membranes and directly or indirectly mediate cholesterol transfer. OSBP-related protein 1L (ORP1L), ORP5 and ORP6 mediate cholesterol transfer to the endoplasmic reticulum that regulates cholesterol homeostasis. ORP1L and STARD3 also move cholesterol from the endoplasmic reticulum-to-late endosomal/lysosomes under low-cholesterol conditions to facilitate intraluminal vesicle formation. Cholesterol transport also occurs at MCS with peroxisomes and possibly the plasma membrane. Frequent contacts between organelles and the endo-lysosomal vesicles are sites for bidirectional transfer of cholesterol.

Snapin-regulated late endosomal transport is critical for efficient autophagy-lysosomal function in neurons.

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Cai, Qian; Lu, Li; Tian, Jin-Hua; Zhu, Yi-Bing; Qiao, Haifa; Sheng, Zu-Hang

2010-10-06

Neuron maintenance and survival require late endocytic transport from distal processes to the soma where lysosomes are predominantly localized. Here, we report a role for Snapin in attaching dynein to late endosomes through its intermediate chain (DIC). snapin(-/-) neurons exhibit aberrant accumulation of immature lysosomes, clustering and impaired retrograde transport of late endosomes along processes, reduced lysosomal proteolysis due to impaired delivery of internalized proteins and hydrolase precursors from late endosomes to lysosomes, and impaired clearance of autolysosomes, combined with reduced neuron viability and neurodegeneration. The phenotypes are rescued by expressing the snapin transgene, but not the DIC-binding-defective Snapin-L99K mutant. Snapin overexpression in wild-type neurons enhances late endocytic transport and lysosomal function, whereas expressing the mutant defective in Snapin-DIC coupling shows a dominant-negative effect. Altogether, our study highlights new mechanistic insights into how Snapin-DIC coordinates retrograde transport and late endosomal-lysosomal trafficking critical for autophagy-lysosomal function, and thus neuronal homeostasis. Copyright © 2010 Elsevier Inc. All rights reserved.

[The history of optical signals for traffic regulation].

Science.gov (United States)

Draeger, J; Harsch, V

2008-04-01

For signal transmission in traffic today, different optical, acoustic, or other physical or technical means are used for information. The different kinds of traffic (water navigation, road and rail, and, later air transport) made traffic regulation necessary early on. This regulation, from its very beginning in ancient times, began by means of optical signals; nowadays, this remains the most important method. From the very start, minimum requirements for the navigator's vision, color discrimination, dark adaptation, and even visual field were needed. For historical reasons, it was in seafaring medicine that these first developed. Besides the development of the different signals, methods for checking the requirements were soon developed. National and international requirements have been very different. Only within the last 50 years has international cooperation led to the acceptance of general standards for the different traffic modes. This article discusses the technical development of optical signals for the different kinds of traffic, from ancient times to the present, and explains the development of minimum requirements for the different visual functions.

Regulation of VEGF signaling by membrane traffic.

Science.gov (United States)

Horowitz, Arie; Seerapu, Himabindu Reddy

2012-09-01

Recent findings have drawn attention to the role of membrane traffic in the signaling of vascular endothelial growth factor (VEGF). The significance of this development stems from the pivotal function of VEGF in vasculogenesis and angiogenesis. The outline of the regulation of VEGF receptor (VEGFR) signaling by membrane traffic is similar to that of the epidermal growth factor receptor (EGFR), a prototype of the intertwining between membrane traffic and signaling. There are, however, unique features in VEGFR signaling that are conferred in part by the involvement of the co-receptor neuropilin (Nrp). Nrp1 and VEGFR2 are integrated into membrane traffic through the adaptor protein synectin, which recruits myosin VI, a molecular motor that drives inward trafficking [17,21,64]. The recent detection of only mild vascular defects in a knockin mouse model that expresses Nrp1 lacking a cytoplasmic domain [104], questions the co-receptor's role in VEGF signaling and membrane traffic. The regulation of endocytosis by ephrin-B2 is another feature unique to VEGR2/3 [18,19], but it awaits a mechanistic explanation. Current models do not fully explain how membrane traffic bridges between VEGFR and the downstream effectors that produce its functional outcome, such as cell migration. VEGF-A appears to accomplish this task in part by recruiting endocytic vesicles carrying RhoA to internalized active VEGFR2 [58]. Copyright © 2012 Elsevier Inc. All rights reserved.

Integration of two RAB5 groups during endosomal transport in plants

Science.gov (United States)

Ebine, Kazuo; Choi, Seung-won; Ichinose, Sakura; Uemura, Tomohiro; Nakano, Akihiko

2018-01-01

RAB5 is a key regulator of endosomal functions in eukaryotic cells. Plants possess two different RAB5 groups, canonical and plant-unique types, which act via unknown counteracting mechanisms. Here, we identified an effector molecule of the plant-unique RAB5 in Arabidopsis thaliana, ARA6, which we designated PLANT-UNIQUE RAB5 EFFECTOR 2 (PUF2). Preferential colocalization with canonical RAB5 on endosomes and genetic interaction analysis indicated that PUF2 coordinates vacuolar transport with canonical RAB5, although PUF2 was identified as an effector of ARA6. Competitive binding of PUF2 with GTP-bound ARA6 and GDP-bound canonical RAB5, together interacting with the shared activating factor VPS9a, showed that ARA6 negatively regulates canonical RAB5-mediated vacuolar transport by titrating PUF2 and VPS9a. These results suggest a unique and unprecedented function for a RAB effector involving the integration of two RAB groups to orchestrate endosomal trafficking in plant cells. PMID:29749929

deep-orange and carnation define distinct stages in late endosomal biogenesis in Drosophila melanogaster.

Science.gov (United States)

Sriram, V; Krishnan, K S; Mayor, Satyajit

2003-05-12

Endosomal degradation is severely impaired in primary hemocytes from larvae of eye color mutants of Drosophila. Using high resolution imaging and immunofluorescence microscopy in these cells, products of eye color genes, deep-orange (dor) and carnation (car), are localized to large multivesicular Rab7-positive late endosomes containing Golgi-derived enzymes. These structures mature into small sized Dor-negative, Car-positive structures, which subsequently fuse to form tubular lysosomes. Defective endosomal degradation in mutant alleles of dor results from a failure of Golgi-derived vesicles to fuse with morphologically arrested Rab7-positive large sized endosomes, which are, however, normally acidified and mature with wild-type kinetics. This locates the site of Dor function to fusion of Golgi-derived vesicles with the large Rab7-positive endocytic compartments. In contrast, endosomal degradation is not considerably affected in car1 mutant; fusion of Golgi-derived vesicles and maturation of large sized endosomes is normal. However, removal of Dor from small sized Car-positive endosomes is slowed, and subsequent fusion with tubular lysosomes is abolished. Overexpression of Dor in car1 mutant aggravates this defect, implicating Car in the removal of Dor from endosomes. This suggests that, in addition to an independent role in fusion with tubular lysosomes, the Sec1p homologue, Car, regulates Dor function.

Methods of analysis of the membrane trafficking pathway from recycling endosomes to lysosomes.

Science.gov (United States)

Matsui, Takahide; Fukuda, Mitsunori

2014-01-01

The transferrin receptor (TfR) is responsible for iron uptake through its trafficking between the plasma membrane and recycling endosomes, and as a result it has become a well-known marker for recycling endosomes. Although the molecular basis of the TfR recycling pathway has been thoroughly investigated, the TfR degradation mechanism has been poorly understood. Exposure of cultured cells to two drugs, the protein synthesis inhibitor cycloheximide and the V-ATPase inhibitor bafilomycin A1, recently showed that TfR is not only recycled back to the plasma membrane after endocytosis but is constitutively transported to lysosomes for degradation. The results of genome-wide screening of mouse Rab small GTPases (common regulators of membrane trafficking in all eukaryotes) have indicated that Rab12 regulates TfR trafficking to lysosomes independently of the known membrane trafficking pathways, for example, the conventional endocytic pathway and recycling pathway. This chapter summarizes the methods that the authors used to analyze the membrane trafficking pathway from recycling endosomes to lysosomes that is specifically regulated by Rab12. © 2014 Elsevier Inc. All rights reserved.

The structure and function of presynaptic endosomes

Energy Technology Data Exchange (ETDEWEB)

Jähne, Sebastian, E-mail: sebastian.jaehne1@stud.uni-goettingen.de [Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, Humboldtallee 23, 37073 Göttingen (Germany); International Max Planck Research School for Neurosciences, 37077 Göttingen (Germany); Rizzoli, Silvio O. [Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, Humboldtallee 23, 37073 Göttingen (Germany); Helm, Martin S., E-mail: martin.helm@med.uni-goettingen.de [Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, Humboldtallee 23, 37073 Göttingen (Germany); International Max Planck Research School for Molecular Biology, 37077 Göttingen (Germany)

2015-07-15

The function of endosomes and of endosome-like structures in the presynaptic compartment is still controversial. This is in part due to the absence of a consensus on definitions and markers for these compartments. Synaptic endosomes are sometimes seen as stable organelles, permanently present in the synapse. Alternatively, they are seen as short-lived intermediates in synaptic vesicle recycling, arising from the endocytosis of large vesicles from the plasma membrane, or from homotypic fusion of small vesicles. In addition, the potential function of the endosome is largely unknown in the synapse. Some groups have proposed that the endosome is involved in the sorting of synaptic vesicle proteins, albeit others have produced data that deny this possibility. In this review, we present the existing evidence for synaptic endosomes, we discuss their potential functions, and we highlight frequent technical pitfalls in the analysis of this elusive compartment. We also sketch a roadmap to definitely determine the role of synaptic endosomes for the synaptic vesicle cycle. Finally, we propose a common definition of synaptic endosome-like structures.

The structure and function of presynaptic endosomes

International Nuclear Information System (INIS)

Jähne, Sebastian; Rizzoli, Silvio O.; Helm, Martin S.

2015-01-01

The function of endosomes and of endosome-like structures in the presynaptic compartment is still controversial. This is in part due to the absence of a consensus on definitions and markers for these compartments. Synaptic endosomes are sometimes seen as stable organelles, permanently present in the synapse. Alternatively, they are seen as short-lived intermediates in synaptic vesicle recycling, arising from the endocytosis of large vesicles from the plasma membrane, or from homotypic fusion of small vesicles. In addition, the potential function of the endosome is largely unknown in the synapse. Some groups have proposed that the endosome is involved in the sorting of synaptic vesicle proteins, albeit others have produced data that deny this possibility. In this review, we present the existing evidence for synaptic endosomes, we discuss their potential functions, and we highlight frequent technical pitfalls in the analysis of this elusive compartment. We also sketch a roadmap to definitely determine the role of synaptic endosomes for the synaptic vesicle cycle. Finally, we propose a common definition of synaptic endosome-like structures

Cell polarity and patterning by PIN trafficking through early endosomal compartments in Arabidopsis thaliana.

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Hirokazu Tanaka

2013-05-01

Full Text Available PIN-FORMED (PIN proteins localize asymmetrically at the plasma membrane and mediate intercellular polar transport of the plant hormone auxin that is crucial for a multitude of developmental processes in plants. PIN localization is under extensive control by environmental or developmental cues, but mechanisms regulating PIN localization are not fully understood. Here we show that early endosomal components ARF GEF BEN1 and newly identified Sec1/Munc18 family protein BEN2 are involved in distinct steps of early endosomal trafficking. BEN1 and BEN2 are collectively required for polar PIN localization, for their dynamic repolarization, and consequently for auxin activity gradient formation and auxin-related developmental processes including embryonic patterning, organogenesis, and vasculature venation patterning. These results show that early endosomal trafficking is crucial for cell polarity and auxin-dependent regulation of plant architecture.

Shoc2 is targeted to late endosomes and required for Erk1/2 activation in EGF-stimulated cells.

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Emilia Galperin

Full Text Available Shoc2 is the putative scaffold protein that interacts with RAS and RAF, and positively regulates signaling to extracellular signal-regulated protein kinases 1 and 2 (ERK1/2. To elucidate the mechanism by which Shoc2 regulates ERK1/2 activation by the epidermal growth factor (EGF receptor (EGFR, we studied subcellular localization of Shoc2. Upon EGFR activation, endogenous Shoc2 and red fluorescent protein tagged Shoc2 were translocated from the cytosol to a subset of late endosomes containing Rab7. The endosomal recruitment of Shoc2 was blocked by overexpression of a GDP-bound H-RAS (N17S mutant and RNAi knockdown of clathrin, suggesting the requirement of RAS activity and clathrin-dependent endocytosis. RNAi depletion of Shoc2 strongly inhibited activation of ERK1/2 by low, physiological EGF concentrations, which was rescued by expression of wild-type recombinant Shoc2. In contrast, the Shoc2 (S2G mutant, that is myristoylated and found in patients with the Noonan-like syndrome, did not rescue ERK1/2 activation in Shoc2-depleted cells. Shoc2 (S2G was not located in late endosomes but was present on the plasma membrane and early endosomes. These data suggest that targeting of Shoc2 to late endosomes may facilitate EGFR-induced ERK activation under physiological conditions of cell stimulation by EGF, and therefore, may be involved in the spatiotemporal regulation of signaling through the RAS-RAF module.

Internal structure of magnetic endosomes

Science.gov (United States)

Rivière, C.; Wilhelm, C.; Cousin, F.; Dupuis, V.; Gazeau, F.; Perzynski, R.

2007-01-01

The internal structure of biological vesicles filled with magnetic nanoparticles is investigated using the following complementary analyses: electronic transmission microscopy, dynamic probing by magneto-optical birefringence and structural probing by Small Angle Neutron Scattering (SANS). These magnetic vesicles are magnetic endosomes obtained via a non-specific interaction between cells and anionic magnetic iron oxide nanoparticles. Thanks to a magnetic purification process, they are probed at two different stages of their formation within HeLa cells: (i) adsorption of nanoparticles onto the cellular membrane and (ii) their subsequent internalisation within endosomes. Differences in the microenvironment of the magnetic nanoparticles at those two different stages are highlighted here. The dynamics of magnetic nanoparticles adsorbed onto cellular membranes and confined within endosomes is respectively 3 and 5 orders of magnitude slower than for isolated magnetic nanoparticles in aqueous media. Interestingly, SANS experiments show that magnetic endosomes have an internal structure close to decorated vesicles, with magnetic nanoparticles locally decorating the endosome membrane, inside their inner-sphere. These results, important for future biomedical applications, suggest that multiple fusions of decorated vesicles are the biological processes underlying the endocytosis of that kind of nanometric materials.

Membrane Tethering Complexes in the Endosomal System

OpenAIRE

Spang, Anne

2016-01-01

Vesicles that are generated by endocytic events at the plasma membrane are destined to early endosomes. A prerequisite for proper fusion is the tethering of two membrane entities. Tethering of vesicles to early endosomes is mediated by the class C core vacuole/endosome tethering (CORVET) complex, while fusion of late endosomes with lysosomes depends on the homotypic fusion and vacuole protein sorting (HOPS) complex. Recycling through the trans-Golgi network (TGN) and to the plasma membrane is...

Gαs regulates Glucagon-Like Peptide 1 Receptor-mediated cyclic AMP generation at Rab5 endosomal compartment

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Shravan Babu Girada

2017-10-01

Conclusions: The findings provide the mechanism of endosomal cyclic AMP generation following GLP-1R activation. We identified the specific compartment that serves as an organizing center to generate endosomal cyclic AMP by internalized activated receptor complex.

Traffic

International Nuclear Information System (INIS)

Lichtblau, G.

2001-01-01

This chapter deals with passenger and freight traffic, public and private transportation, traffic related environmental impacts, future developments, traffic indicators, regional traffic planning, health costs due to road traffic related air pollution, noise pollution, measures and regulations for traffic control and fuels for traffic. In particular energy consumption, energy efficiency, pollutant emissions ( CO 2 , SO 2 , NO x , HC, CO, N 2 O, NH 3 and particulates) and environmental effects of the different types of traffic and different types of fuels are compared and studied. Legal regulations and measures for an effective traffic control are discussed. (a.n.)

The V-ATPase a2-subunit as a putative endosomal pH-sensor.

Science.gov (United States)

Marshansky, V

2007-11-01

V-ATPase (vesicular H(+)-ATPase)-driven intravesicular acidification is crucial for vesicular trafficking. Defects in vesicular acidification and trafficking have recently been recognized as essential determinants of various human diseases. An important role of endosomal acidification in receptor-ligand dissociation and in activation of lysosomal hydrolytic enzymes is well established. However, the molecular mechanisms by which luminal pH information is transmitted to the cytosolic small GTPases that control trafficking events such as budding, coat formation and fusion are unknown. Here, we discuss our recent discovery that endosomal V-ATPase is a pH-sensor regulating the degradative pathway. According to our model, V-ATPase is responsible for: (i) the generation of a pH gradient between vesicular membranes; (ii) sensing of intravesicular pH; and (iii) transmitting this information to the cytosolic side of the membrane. We also propose the hypothetical molecular mechanism involved in function of the V-ATPase a2-subunit as a putative pH-sensor. Based on extensive experimental evidence on the crucial role of histidine residues in the function of PSPs (pH-sensing proteins) in eukaryotic cells, we hypothesize that pH-sensitive histidine residues within the intra-endosomal loops and/or C-terminal luminal tail of the a2-subunit could also be involved in the pH-sensing function of V-ATPase. However, in order to identify putative pH-sensitive histidine residues and to test this hypothesis, it is absolutely essential that we increase our understanding of the folding and transmembrane topology of the a-subunit isoforms of V-ATPase. Thus the crucial role of intra-endosomal histidine residues in pH-dependent conformational changes of the V-ATPase a2-isoform, its interaction with cytosolic small GTPases and ultimately in its acidification-dependent regulation of the endosomal/lysosomal protein degradative pathway remain to be determined.

A decision support system for the maintenance of lights of traffic regulation systems

NARCIS (Netherlands)

Duyn Schouten, F.A. van der; Klusener, A.S.; Vlijmen, S.F.M. van; Vos de Wael, S.L.E.

From October 1994 to June 1995 the authors executed a project, the so called 'Lampremplace' project, in cooperation with Nederland Haarlem, a Dutch constructor and supplier of traffic regulation systems. 'Lampremplace', as it is called in traffic jargon, is the maintenance of lights of traffic

Biomechanics and Thermodynamics of Nanoparticle Interactions with Plasma and Endosomal Membrane Lipids in Cellular Uptake and Endosomal Escape

Science.gov (United States)

2015-01-01

To be effective for cytoplasmic delivery of therapeutics, nanoparticles (NPs) taken up via endocytic pathways must efficiently transport across the cell membrane and subsequently escape from the secondary endosomes. We hypothesized that the biomechanical and thermodynamic interactions of NPs with plasma and endosomal membrane lipids are involved in these processes. Using model plasma and endosomal lipid membranes, we compared the interactions of cationic NPs composed of poly(d,l-lactide-co-glycolide) modified with the dichain surfactant didodecyldimethylammonium bromide (DMAB) or the single-chain surfactant cetyltrimethylammonium bromide (CTAB) vs anionic unmodified NPs of similar size. We validated our hypothesis in doxorubicin-sensitive (MCF-7, with relatively fluid membranes) and resistant breast cancer cells (MCF-7/ADR, with rigid membranes). Despite their cationic surface charges, DMAB- and CTAB-modified NPs showed different patterns of biophysical interaction: DMAB-modified NPs induced bending of the model plasma membrane, whereas CTAB-modified NPs condensed the membrane, thereby resisted bending. Unmodified NPs showed no effects on bending. DMAB-modified NPs also induced thermodynamic instability of the model endosomal membrane, whereas CTAB-modified and unmodified NPs had no effect. Since bending of the plasma membrane and destabilization of the endosomal membrane are critical biophysical processes in NP cellular uptake and endosomal escape, respectively, we tested these NPs for cellular uptake and drug efficacy. Confocal imaging showed that in both sensitive and resistant cells DMAB-modified NPs exhibited greater cellular uptake and escape from endosomes than CTAB-modified or unmodified NPs. Further, paclitaxel-loaded DMAB-modified NPs induced greater cytotoxicity even in resistant cells than CTAB-modified or unmodified NPs or drug in solution, demonstrating the potential of DMAB-modified NPs to overcome the transport barrier in resistant cells. In

MiR-153 Regulates Amelogenesis by Targeting Endocytotic and Endosomal/lysosomal Pathways-Novel Insight into the Origins of Enamel Pathologies.

Science.gov (United States)

Yin, Kaifeng; Lin, Wenting; Guo, Jing; Sugiyama, Toshihiro; Snead, Malcolm L; Hacia, Joseph G; Paine, Michael L

2017-03-13

Amelogenesis imperfecta (AI) is group of inherited disorders resulting in enamel pathologies. The involvement of epigenetic regulation in the pathogenesis of AI is yet to be clarified due to a lack of knowledge about amelogenesis. Our previous genome-wide microRNA and mRNA transcriptome analyses suggest a key role for miR-153 in endosome/lysosome-related pathways during amelogenesis. Here we show that miR-153 is significantly downregulated in maturation ameloblasts compared with secretory ameloblasts. Within ameloblast-like cells, upregulation of miR-153 results in the downregulation of its predicted targets including Cltc, Lamp1, Clcn4 and Slc4a4, and a number of miRNAs implicated in endocytotic pathways. Luciferase reporter assays confirmed the predicted interactions between miR-153 and the 3'-UTRs of Cltc, Lamp1 (in a prior study), Clcn4 and Slc4a4. In an enamel protein intake assay, enamel cells transfected with miR-153 show a decreased ability to endocytose enamel proteins. Finally, microinjection of miR-153 in the region of mouse first mandibular molar at postnatal day 8 (PN8) induced AI-like pathologies when the enamel development reached maturity (PN12). In conclusion, miR-153 regulates maturation-stage amelogenesis by targeting key genes involved in the endocytotic and endosomal/lysosomal pathways, and disruption of miR-153 expression is a potential candidate etiologic factor contributing to the occurrence of AI.

MiR-153 Regulates Amelogenesis by Targeting Endocytotic and Endosomal/lysosomal Pathways–Novel Insight into the Origins of Enamel Pathologies

Science.gov (United States)

Yin, Kaifeng; Lin, Wenting; Guo, Jing; Sugiyama, Toshihiro; Snead, Malcolm L.; Hacia, Joseph G.; Paine, Michael L.

2017-01-01

Amelogenesis imperfecta (AI) is group of inherited disorders resulting in enamel pathologies. The involvement of epigenetic regulation in the pathogenesis of AI is yet to be clarified due to a lack of knowledge about amelogenesis. Our previous genome-wide microRNA and mRNA transcriptome analyses suggest a key role for miR-153 in endosome/lysosome-related pathways during amelogenesis. Here we show that miR-153 is significantly downregulated in maturation ameloblasts compared with secretory ameloblasts. Within ameloblast-like cells, upregulation of miR-153 results in the downregulation of its predicted targets including Cltc, Lamp1, Clcn4 and Slc4a4, and a number of miRNAs implicated in endocytotic pathways. Luciferase reporter assays confirmed the predicted interactions between miR-153 and the 3′-UTRs of Cltc, Lamp1 (in a prior study), Clcn4 and Slc4a4. In an enamel protein intake assay, enamel cells transfected with miR-153 show a decreased ability to endocytose enamel proteins. Finally, microinjection of miR-153 in the region of mouse first mandibular molar at postnatal day 8 (PN8) induced AI-like pathologies when the enamel development reached maturity (PN12). In conclusion, miR-153 regulates maturation-stage amelogenesis by targeting key genes involved in the endocytotic and endosomal/lysosomal pathways, and disruption of miR-153 expression is a potential candidate etiologic factor contributing to the occurrence of AI. PMID:28287144

L-Type Voltage-Gated Ca2+ Channels Regulate Synaptic-Activity-Triggered Recycling Endosome Fusion in Neuronal Dendrites

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Brian G. Hiester

2017-11-01

Full Text Available The repertoire and abundance of proteins displayed on the surface of neuronal dendrites are tuned by regulated fusion of recycling endosomes (REs with the dendritic plasma membrane. While this process is critical for neuronal function and plasticity, how synaptic activity drives RE fusion remains unexplored. We demonstrate a multistep fusion mechanism that requires Ca2+ from distinct sources. NMDA receptor Ca2+ initiates RE fusion with the plasma membrane, while L-type voltage-gated Ca2+ channels (L-VGCCs regulate whether fused REs collapse into the membrane or reform without transferring their cargo to the cell surface. Accordingly, NMDA receptor activation triggered AMPA-type glutamate receptor trafficking to the dendritic surface in an L-VGCC-dependent manner. Conversely, potentiating L-VGCCs enhanced AMPA receptor surface expression only when NMDA receptors were also active. Thus L-VGCCs play a role in tuning activity-triggered surface expression of key synaptic proteins by gating the mode of RE fusion.

Agonist-induced down-regulation of endogenous protein kinase c α through an endolysosomal mechanism.

Science.gov (United States)

Lum, Michelle A; Pundt, Krista E; Paluch, Benjamin E; Black, Adrian R; Black, Jennifer D

2013-05-03

Protein kinase C (PKC) isozymes undergo down-regulation upon sustained stimulation. Previous studies have pointed to the existence of both proteasome-dependent and -independent pathways of PKCα processing. Here we demonstrate that these down-regulation pathways are engaged in different subcellular compartments; proteasomal degradation occurs mainly at the plasma membrane, whereas non-proteasomal processing occurs in the perinuclear region. Using cholesterol depletion, pharmacological inhibitors, RNA interference, and dominant-negative mutants, we define the mechanisms involved in perinuclear accumulation of PKCα and identify the non-proteasomal mechanism mediating its degradation. We show that intracellular accumulation of PKCα involves at least two clathrin-independent, cholesterol/lipid raft-mediated pathways that do not require ubiquitination of the protein; one is dynamin-dependent and likely involves caveolae, whereas the other is dynamin- and small GTPase-independent. Internalized PKCα traffics through endosomes and is delivered to the lysosome for degradation. Supportive evidence includes (a) detection of the enzyme in EEA1-positive early endosomes, Rab7-positive late endosomes/multivesicular bodies, and LAMP1-positive lysosomes and (b) inhibition of its down-regulation by lysosome-disrupting agents and leupeptin. Only limited dephosphorylation of PKCα occurs during trafficking, with fully mature enzyme being the main target for lysosomal degradation. These studies define a novel and widespread mechanism of desensitization of PKCα signaling that involves endocytic trafficking and lysosome-mediated degradation of the mature, fully phosphorylated protein.

Role of the AP-5 adaptor protein complex in late endosome-to-Golgi retrieval.

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Jennifer Hirst

2018-01-01

Full Text Available The AP-5 adaptor protein complex is presumed to function in membrane traffic, but so far nothing is known about its pathway or its cargo. We have used CRISPR-Cas9 to knock out the AP-5 ζ subunit gene, AP5Z1, in HeLa cells, and then analysed the phenotype by subcellular fractionation profiling and quantitative mass spectrometry. The retromer complex had an altered steady-state distribution in the knockout cells, and several Golgi proteins, including GOLIM4 and GOLM1, were depleted from vesicle-enriched fractions. Immunolocalisation showed that loss of AP-5 led to impaired retrieval of the cation-independent mannose 6-phosphate receptor (CIMPR, GOLIM4, and GOLM1 from endosomes back to the Golgi region. Knocking down the retromer complex exacerbated this phenotype. Both the CIMPR and sortilin interacted with the AP-5-associated protein SPG15 in pull-down assays, and we propose that sortilin may act as a link between Golgi proteins and the AP-5/SPG11/SPG15 complex. Together, our findings suggest that AP-5 functions in a novel sorting step out of late endosomes, acting as a backup pathway for retromer. This provides a mechanistic explanation for why mutations in AP-5/SPG11/SPG15 cause cells to accumulate aberrant endolysosomes, and highlights the role of endosome/lysosome dysfunction in the pathology of hereditary spastic paraplegia and other neurodegenerative disorders.

Effect of diphtheria toxin T-domain on endosomal pH

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A. J. Labyntsev

2015-08-01

Full Text Available A key step in the mode of cytotoxic action of diphtheria toxin (DT is the transfer of its catalytic domain (Cd from endosomes into the cytosol. The main activity in this process is performed by the transport domain (Td, but the molecular mechanism of its action remains unknown. We have previously shown that Td can have some influence on the endosomal transport of DT. The aim of this work was to study the effect of diphtheria toxin on the toxin compartmentalization in the intracellular transporting pathway and endosomal pH. We used recombinant fragments of DT, which differed only by the presence of Td in their structure, fused with fluorescent proteins. It was shown that the toxin fragment with Td moved slower by the pathway early-late endosomes-lysosomes, and had a slightly different pattern of colocalization with endosomal markers than DT fragment without Td. In addition, endosomes containing DT fragments with Td had a constant pH of about 6.5 from the 10th to 50th minute of observation, for the same time endosomes containing DT fragments without Td demons­trated a decrease in pH from 6.3 to 5.5. These results indicate that Td inhibits acidification of endosomal medium. One of possible explanations for this may be the effect of the ion channel formed by the T-domain on the process of the endosomal acidification. This property of Td may not only inhibit maturation of endosomes but also inhibit activation of endosomal pH-dependent proteases, and this promotes successful transport of Cd into the cell cytosol.

Enhanced Endosomal Escape by Light-Fueled Liquid-Metal Transformer.

Science.gov (United States)

Lu, Yue; Lin, Yiliang; Chen, Zhaowei; Hu, Quanyin; Liu, Yang; Yu, Shuangjiang; Gao, Wei; Dickey, Michael D; Gu, Zhen

2017-04-12

Effective endosomal escape remains as the "holy grail" for endocytosis-based intracellular drug delivery. To date, most of the endosomal escape strategies rely on small molecules, cationic polymers, or pore-forming proteins, which are often limited by the systemic toxicity and lack of specificity. We describe here a light-fueled liquid-metal transformer for effective endosomal escape-facilitated cargo delivery via a chemical-mechanical process. The nanoscale transformer can be prepared by a simple approach of sonicating a low-toxicity liquid-metal. When coated with graphene quantum dots (GQDs), the resulting nanospheres demonstrate the ability to absorb and convert photoenergy to drive the simultaneous phase separation and morphological transformation of the inner liquid-metal core. The morphological transformation from nanospheres to hollow nanorods with a remarkable change of aspect ratio can physically disrupt the endosomal membrane to promote endosomal escape of payloads. This metal-based nanotransformer equipped with GQDs provides a new strategy for facilitating effective endosomal escape to achieve spatiotemporally controlled drug delivery with enhanced efficacy.

The NKG2D ligand ULBP2 is specifically regulated through an invariant chain-dependent endosomal pathway

DEFF Research Database (Denmark)

Uhlenbrock, Franziska Katharina; Hagemann-Jensen, Michael Henrik; Kehlet, Stephanie

2014-01-01

by affecting endosomal/lysosomal integrity and protein kinase C activity. The invariant chain was further essential for endosomal transport of ULBP2. This novel pathway was identified through screening experiments by which methylselenic acid was found to possess notable NKG2D ligand regulatory properties....... The protein kinase C inhibitor methylselenic acid induced MICA/B surface expression but dominantly blocked ULBP2 surface transport. Remarkably, by targeting this novel pathway we could specifically block the production of soluble ULBP2 from different, primary melanomas. Our findings strongly suggest...

The functional interplay of Rab11, FIP3 and Rho proteins on the endosomal recycling pathway controls cell shape and symmetry.

Science.gov (United States)

Bouchet, Jérôme; McCaffrey, Mary W; Graziani, Andrea; Alcover, Andrés

2018-07-04

Several families of small GTPases regulate a variety of fundamental cellular processes, encompassing growth factor signal transduction, vesicular trafficking and control of the cytoskeleton. Frequently, their action is hierarchical and complementary, but much of the detail of their functional interactions remains to be clarified. It is well established that Rab family members regulate a variety of intracellular vesicle trafficking pathways. Moreover, Rho family GTPases are pivotal for the control of the actin and microtubule cytoskeleton. However, the interplay between these 2 types of GTPases has been rarely reported. We discuss here our recent findings showing that Rab11, a key regulator of endosomal recycling, and Rac1, a central actin cytoskeleton regulator involved in lamellipodium formation and cell migration, interplay on endosomes through the Rab11 effector FIP3. In the context of the rapidly reactive T lymphocytes, Rab11-Rac1 endosomal functional interplay is important to control cell shape changes and cell symmetry during lymphocyte spreading and immunological synapse formation and ultimately modulate T cell activation.

Unconventional secretion of tissue transglutaminase involves phospholipid-dependent delivery into recycling endosomes.

Directory of Open Access Journals (Sweden)

Evgeny A Zemskov

2011-04-01

Full Text Available Although endosomal compartments have been suggested to play a role in unconventional protein secretion, there is scarce experimental evidence for such involvement. Here we report that recycling endosomes are essential for externalization of cytoplasmic secretory protein tissue transglutaminase (tTG. The de novo synthesized cytoplasmic tTG does not follow the classical ER/Golgi-dependent secretion pathway, but is targeted to perinuclear recycling endosomes, and is delivered inside these vesicles prior to externalization. On its route to the cell surface tTG interacts with internalized β1 integrins inside the recycling endosomes and is secreted as a complex with recycled β1 integrins. Inactivation of recycling endosomes, blocking endosome fusion with the plasma membrane, or downregulation of Rab11 GTPase that controls outbound trafficking of perinuclear recycling endosomes, all abrogate tTG secretion. The initial recruitment of cytoplasmic tTG to recycling endosomes and subsequent externalization depend on its binding to phosphoinositides on endosomal membranes. These findings begin to unravel the unconventional mechanism of tTG secretion which utilizes the long loop of endosomal recycling pathway and indicate involvement of endosomal trafficking in non-classical protein secretion.

Caveolin targeting to late endosome/lysosomal membranes is induced by perturbations of lysosomal pH and cholesterol content

Science.gov (United States)

Mundy, Dorothy I.; Li, Wei Ping; Luby-Phelps, Katherine; Anderson, Richard G. W.

2012-01-01

Caveolin-1 is an integral membrane protein of plasma membrane caveolae. Here we report that caveolin-1 collects at the cytosolic surface of lysosomal membranes when cells are serum starved. This is due to an elevation of the intralysosomal pH, since ionophores and proton pump inhibitors that dissipate the lysosomal pH gradient also trapped caveolin-1 on late endosome/lysosomes. Accumulation is both saturable and reversible. At least a portion of the caveolin-1 goes to the plasma membrane upon reversal. Several studies suggest that caveolin-1 is involved in cholesterol transport within the cell. Strikingly, we find that blocking cholesterol export from lysosomes with progesterone or U18666A or treating cells with low concentrations of cyclodextrin also caused caveolin-1 to accumulate on late endosome/lysosomal membranes. Under these conditions, however, live-cell imaging shows cavicles actively docking with lysosomes, suggesting that these structures might be involved in delivering caveolin-1. Targeting of caveolin-1 to late endosome/lysosomes is not observed normally, and the degradation rate of caveolin-1 is not altered by any of these conditions, indicating that caveolin-1 accumulation is not a consequence of blocked degradation. We conclude that caveolin-1 normally traffics to and from the cytoplasmic surface of lysosomes during intracellular cholesterol trafficking. PMID:22238363

The PDZ protein GIPC regulates trafficking of the LPA1 receptor from APPL signaling endosomes and attenuates the cell's response to LPA.

Directory of Open Access Journals (Sweden)

Tal Varsano

Full Text Available Lysophosphatidic acid (LPA mediates diverse cellular responses through the activation of at least six LPA receptors--LPA(1-6, but the interacting proteins and signaling pathways that mediate the specificity of these receptors are largely unknown. We noticed that LPA(1 contains a PDZ binding motif (SVV identical to that present in two other proteins that interact with the PDZ protein GIPC. GIPC is involved in endocytic trafficking of several receptors including TrkA, VEGFR2, lutropin and dopamine D2 receptors. Here we show that GIPC binds directly to the PDZ binding motif of LPA(1 but not that of other LPA receptors. LPA(1 colocalizes and coimmunoprecipitates with GIPC and its binding partner APPL, an activator of Akt signaling found on APPL signaling endosomes. GIPC depletion by siRNA disturbed trafficking of LPA(1 to EEA1 early endosomes and promoted LPA(1 mediated Akt signaling, cell proliferation, and cell motility. We propose that GIPC binds LPA(1 and promotes its trafficking from APPL-containing signaling endosomes to EEA1 early endosomes and thus attenuates LPA-mediated Akt signaling from APPL endosomes.

INFLUENCE OF PLANNING, ORGANIZATIONAL CHARACTERISTICS AND REGULATION ON ROAD TRAFFIC SAFETY OF PEDESTRIANS

Directory of Open Access Journals (Sweden)

G. M. Kuharenok

2011-01-01

Full Text Available The paper presents results of research on planning, organizational characteristics and regulation modes at  the regulated pedestrian crossings, located out of crossroads in the street and road network of Minsk. Some regularities pertaining to the influence of the investigated characteristics on road traffic safety of pedestrians are revealed in the paper. Practical offers on increase of road traffic safety of pedestrians in the Republic of Belarus have been developed on the basis of the executed investigations and cited in the paper. 

The International Traffic in Arms Regulations: An Impediment to National Security

Science.gov (United States)

2008-05-02

00-00-2007 to 00-00-2008 4. TITLE AND SUBTITLE The International Traffic in Arms Regulations: An Impediment to National Security 5a . CONTRACT...2008 2. REPORT TYPE Program Research Paper 3. DATES COVERED (From - To) 4. TITLE AND SUBTITLE 5a . CONTRACT NUMBER The International Traffic in Arms...certain level of ITAR problems, such as on Koreasat 5 with its dual civil and military uses, U.S. companies will often choose not to expend the bid and

Visual problem solving and self-regulation in training air traffic control

NARCIS (Netherlands)

Van Meeuwen, Ludo

2013-01-01

Van Meeuwen, L. W. (2013). Visual problem solving and self-regulation in training air traffic control (Unpublished doctoral dissertation). Centre for Learning Sciences and Technologies, Open Universiteit, Heerlen, The Netherlands.

Visual Problem Solving and Self‐regulation in Training Air Traffic Control

NARCIS (Netherlands)

Meeuwen van, Ludo

2015-01-01

Van Meeuwen, L. W. (2013). Visual problem solving and self-regulation in training air traffic control (Unpublished doctoral dissertation). Centre for Learning Sciences and Technologies, Open Universiteit, Heerlen, The Netherlands.

The International Traffic in Arms Regulations: An Impediment to National Security

National Research Council Canada - National Science Library

Mahon, Steven G

2008-01-01

The International Traffic in Arms Regulations (ITAR) is putting United States (U.S.) advanced technology and weapons companies at a competitive disadvantage, particularly in the fields of satellite manufacturing and technology...

The translocon protein Sec61 mediates antigen transport from endosomes in the cytosol for cross-presentation to CD8(+) T cells.

Science.gov (United States)

Zehner, Matthias; Marschall, Andrea L; Bos, Erik; Schloetel, Jan-Gero; Kreer, Christoph; Fehrenschild, Dagmar; Limmer, Andreas; Ossendorp, Ferry; Lang, Thorsten; Koster, Abraham J; Dübel, Stefan; Burgdorf, Sven

2015-05-19

The molecular mechanisms regulating antigen translocation into the cytosol for cross-presentation are under controversial debate, mainly because direct data is lacking. Here, we have provided direct evidence that the activity of the endoplasmic reticulum (ER) translocon protein Sec61 is essential for endosome-to-cytosol translocation. We generated a Sec61-specific intrabody, a crucial tool that trapped Sec61 in the ER and prevented its recruitment into endosomes without influencing Sec61 activity and antigen presentation in the ER. Expression of this ER intrabody inhibited antigen translocation and cross-presentation, demonstrating that endosomal Sec61 indeed mediates antigen transport across endosomal membranes. Moreover, we showed that the recruitment of Sec61 toward endosomes, and hence antigen translocation and cross-presentation, is dependent on dendritic cell activation by Toll-like receptor (TLR) ligands. These data shed light on a long-lasting question regarding antigen cross-presentation and point out a role of the ER-associated degradation machinery in compartments distinct from the ER. Copyright © 2015 Elsevier Inc. All rights reserved.

Rab9-dependent retrograde transport and endosomal sorting of the endopeptidase furin

Science.gov (United States)

Chia, Pei Zhi Cheryl; Gasnereau, Isabelle; Lieu, Zi Zhao; Gleeson, Paul A.

2011-01-01

The endopeptidase furin and the trans-Golgi network protein TGN38 are membrane proteins that recycle between the TGN and plasma membrane. TGN38 is transported by a retromer-dependent pathway from early endosomes to the TGN, whereas the intracellular transport of furin is poorly defined. Here we have identified the itinerary and transport requirements of furin. Using internalisation assays, we show that furin transits the early and late endosomes en route to the TGN. The GTPase Rab9 and the TGN golgin GCC185, components of the late endosome-to-TGN pathway, were required for efficient TGN retrieval of furin. By contrast, TGN38 trafficking was independent of Rab9 and GCC185. To identify the sorting signals for the early endosome-to-TGN pathway, the trafficking of furin–TGN38 chimeras was investigated. The diversion of furin from the Rab9-dependent late-endosome-to-TGN pathway to the retromer-dependent early-endosome-to-TGN pathway required both the transmembrane domain and cytoplasmic tail of TGN38. We present evidence to suggest that the length of the transmembrane domain is a contributing factor in endosomal sorting. Overall, these data show that furin uses the Rab9-dependent pathway from late endosomes and that retrograde transport directly from early endosomes is dependent on both the transmembrane domain and the cytoplasmic tail. PMID:21693586

The endosomal recycling of FgSnc1 by FgSnx41-FgSnx4 heterodimer is essential for polarized growth and pathogenicity in Fusarium graminearum.

Science.gov (United States)

Zheng, Wenhui; Lin, Yahong; Fang, Wenqin; Zhao, Xu; Lou, Yi; Wang, Guanghui; Zheng, Huawei; Liang, Qifu; Abubakar, Yakubu Saddeeq; Olsson, Stefan; Zhou, Jie; Wang, Zonghua

2018-04-20

Endosomal sorting machineries regulate the transport of their cargoes among intracellular compartments. However, the molecular nature of such intracellular trafficking processes in pathogenic fungal development and pathogenicity remains unclear. Here, we dissect the roles and molecular mechanisms of two sorting nexin proteins and their cargoes in endosomal recycling in Fusarium graminearum using high-resolution microscopy and high-throughput co-immunoprecipitation strategies. We show that the sorting nexins, FgSnx41 and FgSnx4, interact with each other and assemble into a functionally interdependent heterodimer through their respective BAR domains. Further analyses demonstrate that the dimer localizes to the early endosomal membrane and coordinates endosomal sorting. The small GTPase FgRab5 regulates the correct localization of FgSnx41-FgSnx4 and is consequently required for its trafficking function. The protein FgSnc1 is a cargo of FgSnx41-FgSnx4 and regulates the fusion of secreted vesicles with the fungal growing apex and plasma membrane. In the absence of FgSnx41 or FgSnx4, FgSnc1 is mis-sorted and degraded in the vacuole, and null deletion of either component causes defects in the fungal polarized growth and virulence. Overall, for the first time, our results reveal the mechanism of FgSnc1 endosomal recycling by FgSnx41-FgSnx4 heterodimer which is essential for polarized growth and pathogenicity in F. graminearum. © 2018 The Authors. New Phytologist © 2018 New Phytologist Trust.

HIV-1 Envelope Glycoprotein Trafficking through the Endosomal Recycling Compartment Is Required for Particle Incorporation.

Science.gov (United States)

Kirschman, Junghwa; Qi, Mingli; Ding, Lingmei; Hammonds, Jason; Dienger-Stambaugh, Krista; Wang, Jaang-Jiun; Lapierre, Lynne A; Goldenring, James R; Spearman, Paul

2018-03-01

The human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) encodes specific trafficking signals within its long cytoplasmic tail (CT) that regulate incorporation into HIV-1 particles. Rab11-family interacting protein 1C (FIP1C) and Rab14 are host trafficking factors required for Env particle incorporation, suggesting that Env undergoes sorting from the endosomal recycling compartment (ERC) to the site of particle assembly on the plasma membrane. We disrupted outward sorting from the ERC by expressing a C-terminal fragment of FIP1C (FIP1C 560-649 ) and examined the consequences on Env trafficking and incorporation into particles. FIP1C 560-649 reduced cell surface levels of Env and prevented its incorporation into HIV-1 particles. Remarkably, Env was trapped in an exaggerated perinuclear ERC in a CT-dependent manner. Mutation of either the Yxxϕ endocytic motif or the YW 795 motif in the CT prevented Env trapping in the ERC and restored incorporation into particles. In contrast, simian immunodeficiency virus SIVmac239 Env was not retained in the ERC, while substitution of the HIV-1 CT for the SIV CT resulted in SIV Env retention in this compartment. These results provide the first direct evidence that Env traffics through the ERC and support a model whereby HIV-1 Env is specifically targeted to the ERC prior to FIP1C- and CT-dependent outward sorting to the particle assembly site on the plasma membrane. IMPORTANCE The HIV envelope protein is an essential component of the viral particle. While many aspects of envelope protein structure and function have been established, the pathway it follows in the cell prior to reaching the site of particle assembly is not well understood. The envelope protein has a very long cytoplasmic tail that interacts with the host cell trafficking machinery. Here, we utilized a truncated form of the trafficking adaptor FIP1C protein to arrest the intracellular transport of the envelope protein, demonstrating that it becomes

Arf6, Rab11 and transferrin receptor define distinct populations of recycling endosomes.

Science.gov (United States)

Kobayashi, Hotaka; Fukuda, Mitsunori

2013-09-01

Recycling endosomes are key platforms for endocytic recycling that return internalized molecules back to the plasma membrane. To determine how recycling endosomes perform their functions, searching for proteins and lipids that specifically localized at recycling endosomes has often been performed by colocalization analyses between candidate molecules and conventional recycling endosome markers. However, it remains unclear whether all the conventional markers have identical localizations. Here we report finding that three well-known recycling endosome markers, i.e., Arf6, Rab11 and transferrin receptor (TfR), have different intracellular localizations in PC12 cells. The results of immunofluorescence analyses showed that the signals of endogenous Arf6, Rab11 and TfR in nerve growth factor-stimulated PC12 cells generally differed, although there was some overlapping. Our findings provide new information about recycling endosome markers, and they highlight the heterogeneity of recycling endosomes.

Pannexin2 oligomers localize into endosomal vesicles in mammalian cells while Pannexin1 channels traffic to the plasma membrane

Directory of Open Access Journals (Sweden)

Daniela eBoassa

2015-02-01

Full Text Available Pannexin2 (Panx2 is the largest of three members of the pannexin proteins. Pannexins are topologically related to connexins and innexins, but serve different functional roles than forming gap junctions. We previously showed that pannexins form oligomeric channels but unlike connexins and innexins, they form only single membrane channels. High levels of Panx2 mRNA and protein in the Central Nervous System (CNS have been documented. Whereas Pannexin1 (Panx1 is fairly ubiquitous and Pannexin3 (Panx3 is found in skin and connective tissue, both are fully glycosylated, traffic to the plasma membrane and have functions correlated with extracellular ATP release. Here, we describe trafficking and subcellular localizations of exogenous Panx2 and Panx1 protein expression in MDCK, HeLa and HEK293T cells as well as endogenous Panx1 and Panx2 patterns in the CNS. Panx2 was found in intracellular localizations, was partially N-glycosylated, and localizations were non-overlapping with Panx1. Confocal images of hippocampal sections immunolabeled for the astrocytic protein GFAP, Panx1 and Panx2 demonstrated that the two isoforms, Panx1 and Panx2, localized at different subcellular compartments in both astrocytes and neurons. Using recombinant fusions of Panx2 with appended genetic tags developed for correlated light and electron microscopy and then expressed in different cell lines, we determined that Panx2 is localized in the membrane of intracellular vesicles and not in the endoplasmic reticulum as initially indicated by calnexin colocalization experiments. Dual immunofluorescence imaging with protein markers for specific vesicle compartments showed that Panx2 vesicles are early endosomal in origin. In electron tomographic volumes, cross-sections of these vesicles displayed fine structural details and close proximity to actin filaments. Thus, pannexins expressed at different subcellular compartments likely exert distinct functional roles, particularly in the

Cooperative endocytosis of the endosomal SNARE protein syntaxin-8 and the potassium channel TASK-1

Science.gov (United States)

Renigunta, Vijay; Fischer, Thomas; Zuzarte, Marylou; Kling, Stefan; Zou, Xinle; Siebert, Kai; Limberg, Maren M.; Rinné, Susanne; Decher, Niels; Schlichthörl, Günter; Daut, Jürgen

2014-01-01

The endosomal SNARE protein syntaxin-8 interacts with the acid-sensitive potassium channel TASK-1. The functional relevance of this interaction was studied by heterologous expression of these proteins (and mutants thereof) in Xenopus oocytes and in mammalian cell lines. Coexpression of syntaxin-8 caused a fourfold reduction in TASK-1 current, a corresponding reduction in the expression of TASK-1 at the cell surface, and a marked increase in the rate of endocytosis of the channel. TASK-1 and syntaxin-8 colocalized in the early endosomal compartment, as indicated by the endosomal markers 2xFYVE and rab5. The stimulatory effect of the SNARE protein on the endocytosis of the channel was abolished when both an endocytosis signal in TASK-1 and an endocytosis signal in syntaxin-8 were mutated. A syntaxin-8 mutant that cannot assemble with other SNARE proteins had virtually the same effect as wild-type syntaxin-8. Total internal reflection fluorescence microscopy showed formation and endocytosis of vesicles containing fluorescence-tagged clathrin, TASK-1, and/or syntaxin-8. Our results suggest that the unassembled form of syntaxin-8 and the potassium channel TASK-1 are internalized via clathrin-mediated endocytosis in a cooperative manner. This implies that syntaxin-8 regulates the endocytosis of TASK-1. Our study supports the idea that endosomal SNARE proteins can have functions unrelated to membrane fusion. PMID:24743596

The Annexin A1 Receptor FPR2 Regulates the Endosomal Export of Influenza Virus

Directory of Open Access Journals (Sweden)

Fryad Rahman

2018-05-01

Full Text Available The Formyl Peptide Receptor 2 (FPR2 is a novel promising target for the treatment of influenza. During viral infection, FPR2 is activated by annexinA1, which is present in the envelope of influenza viruses; this activation promotes virus replication. Here, we investigated whether blockage of FPR2 would affect the genome trafficking of influenza virus. We found that, upon infection and cell treatment with the specific FPR2 antagonist WRW4 or the anti-FPR2 monoclonal antibody, FN-1D6-AI, influenza viruses were blocked into endosomes. This effect was independent on the strain and was observed for H1N1 and H3N2 viruses. In addition, blocking FPR2signaling in alveolar lung A549 epithelial cells with the monoclonal anti-FPR2 antibody significantly inhibited virus replication. Altogether, these results show that FPR2signaling interferes with the endosomal trafficking of influenza viruses and provides, for the first time, the proof of concept that monoclonal antibodies directed against FPR2 inhibit virus replication. Antibodies-based therapeutics have emerged as attractive reagents in infectious diseases. Thus, this study suggests that the use of anti-FPR2 antibodies against influenza hold great promise for the future.

Regulation of vesicular traffic by a GTP-binding protein on the cytoplasmic surface of secretory vesicles in yeast

International Nuclear Information System (INIS)

Novick, P.J.; Goud, B.; Salminen, A.; Walworth, N.C.; Nair, J.; Potenza, M.

1988-01-01

Vesicular transport is an important mechanism for the intracellular traffic of proteins and lipids in eukaryotic cells. Vesicles mediate the passage of proteins between the various organelles of the secretory pathway and the exocytic release of these proteins into the extracellular environment. Vesicles also mediate the uptake of proteins and fluid from the external environment, delivering them to endosomes. Despite the generality of the vesicular transport mechanism, the process is not yet understood at a molecular level. The key questions that are addressed are (1) How are vesicles formed from the membrane of the donor organelle? (2) How are these vesicles transported? (3) How do the vesicles recognize the membrane of the target (acceptor) organelle? (4) How is membrane fusion accomplished? The genetic flexibility of yeast has been exploited to identify components of the cellular machinery required for vesicular transport

The Use of Plant Growth Regulators to Improve the Traffic Tolerance and Repair of Overseeded Bermudagrass

OpenAIRE

Marshall, Christopher Scott

2007-01-01

An active football season during the fall acclimation period tests the traffic tolerance of bermudagrass. Exogenous applications of synthetic cytokinins or cytokinin-enhancing plant growth regulators (PGRs), such as trinexapac-ethyl, may improve the traffic tolerance of "Patriot" and "Tifsport" hybrid berudagrasses (Cynodon dactylon var. dactylon x Cynodon transvaalensis). This study was designed to mimic the agronomic practices and traffic stresses experienced at Virginia Tech's Worsham Fiel...

Stochastic acidification, activation of hemagglutinin and escape of influenza viruses from an endosome

Science.gov (United States)

Lagache, Thibault; Sieben, Christian; Meyer, Tim; Herrmann, Andreas; Holcman, David

2017-06-01

Influenza viruses enter the cell inside an endosome. During the endosomal journey, acidification triggers a conformational change of the virus spike protein hemagglutinin (HA) that results in escape of the viral genome from the endosome into the cytoplasm. It is still unclear how the interplay between acidification and HA conformation changes affects the kinetics of the viral endosomal escape. We develop here a stochastic model to estimate the change of conformation of HAs inside the endosome nanodomain. Using a Markov process, we model the arrival of protons to HA binding sites and compute the kinetics of their accumulation. We compute the Mean First Passage Time (MFPT) of the number of HA bound sites to a threshold, which is used to estimate the HA activation rate for a given pH concentration. The present analysis reveals that HA proton binding sites possess a high chemical barrier, ensuring a stability of the spike protein at sub-acidic pH. We predict that activating more than 3 adjacent HAs is necessary to trigger endosomal fusion and this configuration prevents premature release of viruses from early endosomes

VEGFR2 Trafficking, Signaling and Proteolysis is Regulated by the Ubiquitin Isopeptidase USP8.

Science.gov (United States)

Smith, Gina A; Fearnley, Gareth W; Abdul-Zani, Izma; Wheatcroft, Stephen B; Tomlinson, Darren C; Harrison, Michael A; Ponnambalam, Sreenivasan

2016-01-01

Vascular endothelial growth factor A (VEGF-A) regulates many aspects of vascular function. VEGF-A binding to vascular endothelial growth factor receptor 2 (VEGFR2) stimulates endothelial signal transduction and regulates multiple cellular responses. Activated VEGFR2 undergoes ubiquitination but the enzymes that regulate this post-translational modification are unclear. In this study, the de-ubiquitinating enzyme, USP8, is shown to regulate VEGFR2 trafficking, de-ubiquitination, proteolysis and signal transduction. USP8-depleted endothelial cells displayed altered VEGFR2 ubiquitination and production of a unique VEGFR2 extracellular domain proteolytic fragment caused by VEGFR2 accumulation in the endosome-lysosome system. In addition, perturbed VEGFR2 trafficking impaired VEGF-A-stimulated signal transduction in USP8-depleted cells. Thus, regulation of VEGFR2 ubiquitination and de-ubiquitination has important consequences for the endothelial cell response and vascular physiology. © 2015 The Authors. Traffic published by John Wiley & Sons Ltd.

Mouse polyomavirus enters early endosomes, requires their acidic pH for productive infection, and meets transferrin cargo in rab11-positive endosomes

Czech Academy of Sciences Publication Activity Database

Liebl, D.; Difato, F.; Horníková, L.; Mannová, P.; Štokrová, Jitka; Forstová, J.

2006-01-01

Roč. 80, č. 9 (2006), s. 4610-4622 ISSN 0022-538X R&D Projects: GA ČR(CZ) GA204/03/0593; GA MŠk(CZ) LC545 Institutional research plan: CEZ:AV0Z50520514 Keywords : Polyomavirus internalization and trafficking * Early endosomes * Dependence of infection on endosomal pH Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.341, year: 2006

Antigen processing and remodeling of the endosomal pathway: requirements for antigen cross-presentation.

Science.gov (United States)

Compeer, Ewoud Bernardus; Flinsenberg, Thijs Willem Hendrik; van der Grein, Susanna Geertje; Boes, Marianne

2012-01-01

Cross-presentation of endocytosed antigen as peptide/class I major histocompatibility complex complexes plays a central role in the elicitation of CD8(+) T cell clones that mediate anti-viral and anti-tumor immune responses. While it has been clear that there are specific subsets of professional antigen presenting cells capable of antigen cross-presentation, identification of mechanisms involved is still ongoing. Especially amongst dendritic cells (DC), there are specialized subsets that are highly proficient at antigen cross-presentation. We here present a focused survey on the cell biological processes in the endosomal pathway that support antigen cross-presentation. This review highlights DC-intrinsic mechanisms that facilitate the cross-presentation of endocytosed antigen, including receptor-mediated uptake, maturation-induced endosomal sorting of membrane proteins, dynamic remodeling of endosomal structures and cell surface-directed endosomal trafficking. We will conclude with the description of pathogen-induced deviation of endosomal processing, and discuss how immune evasion strategies pertaining endosomal trafficking may preclude antigen cross-presentation.

Aggregation of endosomal-vacuolar compartments in the Aovps24-deleted strain in the filamentous fungus Aspergillus oryzae

International Nuclear Information System (INIS)

Tatsumi, Akinori; Shoji, Jun-ya; Kikuma, Takashi; Arioka, Manabu; Kitamoto, Katsuhiko

2007-01-01

Previously, we found that deletion of Aovps24, an ortholog of Saccharomyces cerevisiae VPS24, that encodes an ESCRT (endosomal sorting complex required for transport)-III component required for late endosomal function results in fragmented and aggregated vacuoles. Although defective late endosomal function is likely responsible for this phenotype, critical lack of our knowledge on late endosomes in filamentous fungi prevented us from further characterization. In this study, we identified late endosomes of Aspergillus oryzae, by expressing a series of fusion proteins of fluorescent proteins with orthologs of late endosomal proteins. Using these fusion proteins as markers, we observed late endosomes in the wild type strain and the Aovps24 disruptant and demonstrated that late endosomes are aberrantly aggregated in the Aovps24 disruptant. Moreover, we revealed that the aggregated late endosomes have features of vacuoles as well. As deletion of another ESCRT-III component-encoding gene, Aovps2, resulted in similar phenotypes to that in the Aovps24 disruptant, phenotypes of the Aovps24 disruptant are probably due to defective late endosomal function

Structural Basis for Endosomal Targeting by the Bro1 Domain

Science.gov (United States)

Kim, Jaewon; Sitaraman, Sujatha; Hierro, Aitor; Beach, Bridgette M.; Odorizzi, Greg; Hurley, James H.

2010-01-01

Summary Proteins delivered to the lysosome or the yeast vacuole via late endosomes are sorted by the ESCRT complexes and by associated proteins, including Alix and its yeast homolog Bro1. Alix, Bro1, and several other late endosomal proteins share a conserved 160 residue Bro1 domain whose boundaries, structure, and function have not been characterized. The crystal structure of the Bro1 domain of Bro1 reveals a folded core of 367 residues. The extended Bro1 domain is necessary and sufficient for binding to the ESCRT-III subunit Snf7 and for the recruitment of Bro1 to late endosomes. The structure resembles a boomerang with its concave face filled in and contains a triple tetratricopeptide repeat domain as a substructure. Snf7 binds to a conserved hydrophobic patch on Bro1 that is required for protein complex formation and for the protein-sorting function of Bro1. These results define a conserved mechanism whereby Bro1 domain-containing proteins are targeted to endosomes by Snf7 and its orthologs. PMID:15935782

IL4/PGE2 induction of an enlarged early endosomal compartment in mouse macrophages is Rab5-dependent

International Nuclear Information System (INIS)

Wainszelbaum, Marisa J.; Proctor, Brandon M.; Pontow, Suzanne E.; Stahl, Philip D.; Barbieri, M. Alejandro

2006-01-01

The endosomal compartment and the plasma membrane form a complex partnership that controls signal transduction and trafficking of different molecules. The specificity and functionality of the early endocytic pathway are regulated by a growing number of Rab GTPases, particularly Rab5. In this study, we demonstrate that IL4 (a Th-2 cytokine) and prostaglandin E 2 (PGE 2 ) synergistically induce Rab5 and several Rab effector proteins, including Rin1 and EEA1, and promote the formation of an enlarged early endocytic (EEE) compartment. Endosome enlargement is linked to a substantial induction of the mannose receptor (MR), a well-characterized macrophage endocytic receptor. Both MR levels and MR-mediated endocytosis are enhanced approximately 7-fold. Fluid-phase endocytosis is also elevated in treated cells. Light microscopy and fractionation studies reveal that MR colocalizes predominantly with Rab5a and partially with Rab11, an endosomal recycling pathway marker. Using retroviral expression of Rab5a:S34N, a dominant negative mutant, and siRNA Rab5a silencing, we demonstrate that Rab5a is essential for the large endosome phenotype and for localization of MR in these structures. We speculate that the EEE is maintained by activated Rab5, and that the EEE phenotype is part of some macrophage developmental program such as cell fusion, a characteristic of IL4-stimulated cells

Post-Golgi anterograde transport requires GARP-dependent endosome-to-TGN retrograde transport

Science.gov (United States)

Hirata, Tetsuya; Fujita, Morihisa; Nakamura, Shota; Gotoh, Kazuyoshi; Motooka, Daisuke; Murakami, Yoshiko; Maeda, Yusuke; Kinoshita, Taroh

2015-01-01

The importance of endosome-to–trans-Golgi network (TGN) retrograde transport in the anterograde transport of proteins is unclear. In this study, genome-wide screening of the factors necessary for efficient anterograde protein transport in human haploid cells identified subunits of the Golgi-associated retrograde protein (GARP) complex, a tethering factor involved in endosome-to-TGN transport. Knockout (KO) of each of the four GARP subunits, VPS51–VPS54, in HEK293 cells caused severely defective anterograde transport of both glycosylphosphatidylinositol (GPI)-anchored and transmembrane proteins from the TGN. Overexpression of VAMP4, v-SNARE, in VPS54-KO cells partially restored not only endosome-to-TGN retrograde transport, but also anterograde transport of both GPI-anchored and transmembrane proteins. Further screening for genes whose overexpression normalized the VPS54-KO phenotype identified TMEM87A, encoding an uncharacterized Golgi-resident membrane protein. Overexpression of TMEM87A or its close homologue TMEM87B in VPS54-KO cells partially restored endosome-to-TGN retrograde transport and anterograde transport. Therefore GARP- and VAMP4-dependent endosome-to-TGN retrograde transport is required for recycling of molecules critical for efficient post-Golgi anterograde transport of cell-surface integral membrane proteins. In addition, TMEM87A and TMEM87B are involved in endosome-to-TGN retrograde transport. PMID:26157166

The p25 Subunit of the Dynactin Complex is Required for Dynein-Early Endosome Interaction

Science.gov (United States)

2011-01-01

early endosome movement. In filamentous hyphae , dynein powers the minus end–directed movement of early endosomes (Steinberg and Schuster 2011...observed in time-lapse sequences (Video 1; Abenza et al., 2009). In still images, early endosomes were seen to distribute along the hyphae (Fig. 2 A...nuclear distribution along elongated hyphae and also for the microtubule minus end–directed movement of early endosomes away from the tip (Morris

77 FR 39392 - Amendment to the International Traffic in Arms Regulations: Yemen

Science.gov (United States)

2012-07-03

... International Traffic in Arms Regulations: Yemen AGENCY: Department of State. ACTION: Final rule. SUMMARY: The... toward Yemen. Licenses or other approvals for exports or imports of defense articles and defense services destined for or originating in Yemen will be reviewed, and may be issued, on a case-by-case basis. DATES...

Antigen processing and remodeling of the endosomal pathway: requirements for antigen cross-presentation.

Directory of Open Access Journals (Sweden)

Ewoud Bernardus Compeer

2012-03-01

Full Text Available The cross-presentation of endocytosed antigen as peptide/class I MHC complexes plays a central role in the elicitation of CD8+ T cell clones that mediate anti-viral and anti-tumor immune responses. While it has been clear that there are specific subsets of professional antigen presenting cells (APC capable of antigen cross-presentation, description of mechanisms involved is still ongoing. Especially amongst dendritic cells (DC, there are specialized subsets that are highly proficient at antigen cross-presentation. We here present a focused survey on the cell biological processes in the endosomal pathway that support antigen cross-presentation. This review highlight DC-intrinsic mechanisms that facilitate the cross-presentation of endocytosed antigen, including receptor-mediated uptake, recycling and maturation including the sorting of membrane proteins, dynamic remodeling of endosomal structures and cell-surface directed endosomal trafficking. We will conclude with description of pathogen-induced deviation of endosomal processing, and discuss how immune evasion strategies pertaining endosomal trafficking may preclude antigen cross-presentation.

Charcot-Marie-Tooth disease-linked protein SIMPLE functions with the ESCRT machinery in endosomal trafficking

OpenAIRE

Lee, Samuel M.; Chin, Lih-Shen; Li, Lian

2012-01-01

Mutations in small integral membrane protein of lysosome/late endosome (SIMPLE) cause autosomal dominant, Charcot-Marie-Tooth disease (CMT) type 1C. The cellular function of SIMPLE is unknown and the pathogenic mechanism of SIMPLE mutations remains elusive. Here, we report that SIMPLE interacted and colocalized with endosomal sorting complex required for transport (ESCRT) components STAM1, Hrs, and TSG101 on early endosomes and functioned with the ESCRT machinery in the control of endosome-to...

NEU3 Sialidase Protein Interactors in the Plasma Membrane and in the Endosomes*

Science.gov (United States)

Cirillo, Federica; Ghiroldi, Andrea; Fania, Chiara; Piccoli, Marco; Torretta, Enrica; Tettamanti, Guido; Gelfi, Cecilia; Anastasia, Luigi

2016-01-01

NEU3 sialidase has been shown to be a key player in many physio- and pathological processes, including cell differentiation, cellular response to hypoxic stress, and carcinogenesis. The enzyme, peculiarly localized on the outer leaflet of the plasma membrane, has been shown to be able to remove sialic acid residues from the gangliosides present on adjacent cells, thus creating cell to cell interactions. Nonetheless, herein we report that the enzyme localization is dynamically regulated between the plasma membrane and the endosomes, where a substantial amount of NEU3 is stored with low enzymatic activity. However, under opportune stimuli, NEU3 is shifted from the endosomes to the plasma membrane, where it greatly increases the sialidase activity. Finally, we found that NEU3 possesses also the ability to interact with specific proteins, many of which are different in each cell compartment. They were identified by mass spectrometry, and some selected ones were also confirmed by cross-immunoprecipitation with the enzyme, supporting NEU3 involvement in the cell stress response, protein folding, and intracellular trafficking. PMID:26987901

Pannexin2 oligomers localize in the membranes of endosomal vesicles in mammalian cells while Pannexin1 channels traffic to the plasma membrane.

Science.gov (United States)

Boassa, Daniela; Nguyen, Phuong; Hu, Junru; Ellisman, Mark H; Sosinsky, Gina E

2014-01-01

Pannexin2 (Panx2) is the largest of three members of the pannexin proteins. Pannexins are topologically related to connexins and innexins, but serve different functional roles than forming gap junctions. We previously showed that pannexins form oligomeric channels but unlike connexins and innexins, they form only single membrane channels. High levels of Panx2 mRNA and protein in the Central Nervous System (CNS) have been documented. Whereas Pannexin1 (Panx1) is fairly ubiquitous and Pannexin3 (Panx3) is found in skin and connective tissue, both are fully glycosylated, traffic to the plasma membrane and have functions correlated with extracellular ATP release. Here, we describe trafficking and subcellular localizations of exogenous Panx2 and Panx1 protein expression in MDCK, HeLa, and HEK 293T cells as well as endogenous Panx1 and Panx2 patterns in the CNS. Panx2 was found in intracellular localizations, was partially N-glycosylated, and localizations were non-overlapping with Panx1. Confocal images of hippocampal sections immunolabeled for the astrocytic protein GFAP, Panx1 and Panx2 demonstrated that the two isoforms, Panx1 and Panx2, localized at different subcellular compartments in both astrocytes and neurons. Using recombinant fusions of Panx2 with appended genetic tags developed for correlated light and electron microscopy and then expressed in different cell lines, we determined that Panx2 is localized in the membrane of intracellular vesicles and not in the endoplasmic reticulum as initially indicated by calnexin colocalization experiments. Dual immunofluorescence imaging with protein markers for specific vesicle compartments showed that Panx2 vesicles are early endosomal in origin. In electron tomographic volumes, cross-sections of these vesicles displayed fine structural details and close proximity to actin filaments. Thus, pannexins expressed at different subcellular compartments likely exert distinct functional roles, particularly in the nervous system.

Two barcodes encoded by the type-1 PDZ and by phospho-Ser312 regulate retromer/WASH-mediated sorting of the ß1-adrenergic receptor from endosomes to the plasma membrane.

Science.gov (United States)

Nooh, Mohammed M; Bahouth, Suleiman W

2017-01-01

Recycling of the majority of agonist-internalized GPCR is dependent on a type I-PDZ "barcode" in their C-tail. The recycling of wild-type (WT) ß 1 -AR is also dependent on its default "type-1 PDZ barcode", but trafficking of the ß 1 -AR is inhibited when PKA or its substrate serine at position 312 (Ser 312 ) are inactivated. We tested the hypothesis that phospho-Ser 312 provided a second barcode for ß 1 -AR sorting from endosomes to the plasma membrane by determining the role of retromer/WASH complexes in ß 1 -AR trafficking. Recycling of WT ß 1 -AR or WT ß 2 -AR was dependent on targeting the retromer to endosomal membranes via SNX3 and rab7a, and on complexing the retromer to the WASH pentamer via the C-tail of FAM21 (FAM21 C ). These maneuvers however, did not inhibit the recycling of a phospho-Ser 312 ß 1 -AR mimic ((S312D) ß 1 -AR). Knockdown of the trans-acting PDZ protein sorting nexin27 (SNX27) inhibited the recycling of WT ß 1 -AR and WT ß 2 -AR, but had no effect on (S312D) ß 1 -AR∆PDZ or on phosphorylation of WT ß 1 -AR by PKA at Ser 312 . However, depletion of FKBP15, a FAM21 C -binding endosomal protein, selectively inhibited WT ß 1 -AR but not ß 2 -AR recycling, suggesting divergence might exist in GPCR trafficking roadmaps. These results indicate that two barcodes are involved in sorting WT ß 1 -AR out of early endosomes. The first and antecedent "barcode" was the "type-1 PDZ", followed by a second reversible "phospho-Ser 312 " verification "barcode". This organization allows tight regulation of ß 1 -AR density to signaling intensity in conditions associated with aberrant ß 1 -AR signaling such as in hypertension and heart failure. Copyright © 2016 Elsevier Inc. All rights reserved.

Syndapin/SDPN-1 is required for endocytic recycling and endosomal actin association in the Caenorhabditis elegans intestine

Science.gov (United States)

Gleason, Adenrele M.; Nguyen, Ken C. Q.; Hall, David H.; Grant, Barth D.

2016-01-01

Syndapin/pascin-family F-BAR domain proteins bind directly to membrane lipids and are associated with actin dynamics at the plasma membrane. Previous reports also implicated mammalian syndapin 2 in endosome function during receptor recycling, but precise analysis of a putative recycling function for syndapin in mammalian systems is difficult because of its effects on the earlier step of endocytic uptake and potential redundancy among the three separate genes that encode mammalian syndapin isoforms. Here we analyze the endocytic transport function of the only Caenorhabditis elegans syndapin, SDPN-1. We find that SDPN-1 is a resident protein of the early and basolateral recycling endosomes in the C. elegans intestinal epithelium, and sdpn-1 deletion mutants display phenotypes indicating a block in basolateral recycling transport. sdpn-1 mutants accumulate abnormal endosomes positive for early endosome and recycling endosome markers that are normally separate, and such endosomes accumulate high levels of basolateral recycling cargo. Furthermore, we observed strong colocalization of endosomal SDPN-1 with the F-actin biosensor Lifeact and found that loss of SDPN-1 greatly reduced Lifeact accumulation on early endosomes. Taken together, our results provide strong evidence for an in vivo function of syndapin in endocytic recycling and suggest that syndapin promotes transport via endosomal fission. PMID:27630264

Retrogradely Transported TrkA Endosomes Signal Locally within Dendrites to Maintain Sympathetic Neuron Synapses

Directory of Open Access Journals (Sweden)

Kathryn M. Lehigh

2017-04-01

Full Text Available Sympathetic neurons require NGF from their target fields for survival, axonal target innervation, dendritic growth and formation, and maintenance of synaptic inputs from preganglionic neurons. Target-derived NGF signals are propagated retrogradely, from distal axons to somata of sympathetic neurons via TrkA signaling endosomes. We report that a subset of TrkA endosomes that are transported from distal axons to cell bodies translocate into dendrites, where they are signaling competent and move bidirectionally, in close proximity to synaptic protein clusters. Using a strategy for spatially confined inhibition of TrkA kinase activity, we found that distal-axon-derived TrkA signaling endosomes are necessary within sympathetic neuron dendrites for maintenance of synapses. Thus, TrkA signaling endosomes have unique functions in different cellular compartments. Moreover, target-derived NGF mediates circuit formation and synapse maintenance through TrkA endosome signaling within dendrites to promote aggregation of postsynaptic protein complexes.

A FYVE zinc finger domain protein specifically links mRNA transport to endosome trafficking.

Science.gov (United States)

Pohlmann, Thomas; Baumann, Sebastian; Haag, Carl; Albrecht, Mario; Feldbrügge, Michael

2015-05-18

An emerging theme in cellular logistics is the close connection between mRNA and membrane trafficking. A prominent example is the microtubule-dependent transport of mRNAs and associated ribosomes on endosomes. This coordinated process is crucial for correct septin filamentation and efficient growth of polarised cells, such as fungal hyphae. Despite detailed knowledge on the key RNA-binding protein and the molecular motors involved, it is unclear how mRNAs are connected to membranes during transport. Here, we identify a novel factor containing a FYVE zinc finger domain for interaction with endosomal lipids and a new PAM2-like domain required for interaction with the MLLE domain of the key RNA-binding protein. Consistently, loss of this FYVE domain protein leads to specific defects in mRNA, ribosome, and septin transport without affecting general functions of endosomes or their movement. Hence, this is the first endosomal component specific for mRNP trafficking uncovering a new mechanism to couple mRNPs to endosomes.

Conformational biosensors reveal GPCR signalling from endosomes

DEFF Research Database (Denmark)

Irannejad, R; Tomshine, Jin C; Tomshine, Jon R

2013-01-01

A long-held tenet of molecular pharmacology is that canonical signal transduction mediated by G-protein-coupled receptor (GPCR) coupling to heterotrimeric G proteins is confined to the plasma membrane. Evidence supporting this traditional view is based on analytical methods that provide limited...... or no subcellular resolution. It has been subsequently proposed that signalling by internalized GPCRs is restricted to G-protein-independent mechanisms such as scaffolding by arrestins, or GPCR activation elicits a discrete form of persistent G protein signalling, or that internalized GPCRs can indeed contribute...... in the early endosome membrane, and that internalized receptors contribute to the overall cellular cyclic AMP response within several minutes after agonist application. These findings provide direct support for the hypothesis that canonical GPCR signalling occurs from endosomes as well as the plasma membrane...

PTP1B targets the endosomal sorting machinery

DEFF Research Database (Denmark)

Stuible, Matthew; Abella, Jasmine V; Feldhammer, Matthew

2010-01-01

Dephosphorylation and endocytic down-regulation are distinct processes that together control the signaling output of a variety of receptor tyrosine kinases (RTKs). PTP1B can directly dephosphorylate several RTKs, but it can also promote activation of downstream pathways through largely unknown...... mechanisms. These positive signaling functions likely contribute to the tumor-promoting effect of PTP1B in mouse cancer models. Here, we have identified STAM2, an endosomal protein involved in sorting activated RTKs for lysosomal degradation, as a substrate of PTP1B. PTP1B interacts with STAM2 at defined...... phosphotyrosine sites, and knockdown of PTP1B expression augments STAM2 phosphorylation. Intriguingly, manipulating the expression and phosphorylation state of STAM2 did not have a general effect on epidermal growth factor (EGF)-induced EGF receptor trafficking, degradation, or signaling. Instead, phosphorylated...

Rabies virus co-localizes with early (Rab5) and late (Rab7) endosomal proteins in neuronal and SH-SY5Y cells.

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Ahmad, Waqas; Li, Yingying; Guo, Yidi; Wang, Xinyu; Duan, Ming; Guan, Zhenhong; Liu, Zengshan; Zhang, Maolin

2017-06-01

Rabies virus (RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and pH-dependent pathway for trafficking and invasion into endothelial cells. Early (Rab5, EEA1) and late (Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5Y cells was investigated. Immunofluorescence, TCID 50 titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein (N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.

α1B-Adrenergic Receptors Differentially Associate with Rab Proteins during Homologous and Heterologous Desensitization

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Castillo-Badillo, Jean A.; Sánchez-Reyes, Omar B.; Alfonzo-Méndez, Marco A.; Romero-Ávila, M. Teresa; Reyes-Cruz, Guadalupe; García-Sáinz, J. Adolfo

2015-01-01

Internalization of G protein-coupled receptors can be triggered by agonists or by other stimuli. The process begins within seconds of cell activation and contributes to receptor desensitization. The Rab GTPase family controls endocytosis, vesicular trafficking, and endosomal fusion. Among their remarkable properties is the differential distribution of its members on the surface of various organelles. In the endocytic pathway, Rab 5 controls traffic from the plasma membrane to early endosomes, whereas Rab 4 and Rab 11 regulate rapid and slow recycling from early endosomes to the plasma membrane, respectively. Moreover, Rab 7 and Rab 9 regulate the traffic from late endosomes to lysosomes and recycling to the trans-Golgi. We explore the possibility that α1B-adrenergic receptor internalization induced by agonists (homologous) and by unrelated stimuli (heterologous) could involve different Rab proteins. This possibility was explored by Fluorescence Resonance Energy Transfer (FRET) using cells coexpressing α1B-adrenergic receptors tagged with the red fluorescent protein, DsRed, and different Rab proteins tagged with the green fluorescent protein. It was observed that when α1B-adrenergic receptors were stimulated with noradrenaline, the receptors interacted with proteins present in early endosomes, such as the early endosomes antigen 1, Rab 5, Rab 4, and Rab 11 but not with late endosome markers, such as Rab 9 and Rab 7. In contrast, sphingosine 1-phosphate stimulation induced rapid and transient α1B-adrenergic receptor interaction of relatively small magnitude with Rab 5 and a more pronounced and sustained one with Rab 9; interaction was also observed with Rab 7. Moreover, the GTPase activity of the Rab proteins appears to be required because no FRET was observed when dominant-negative Rab mutants were employed. These data indicate that α1B-adrenergic receptors are directed to different endocytic vesicles depending on the desensitization type (homologous vs

Rapid transporter regulation prevents substrate flow traffic jams in boron transport

Science.gov (United States)

Sotta, Naoyuki; Duncan, Susan; Tanaka, Mayuki; Sato, Takafumi

2017-01-01

Nutrient uptake by roots often involves substrate-dependent regulated nutrient transporters. For robust uptake, the system requires a regulatory circuit within cells and a collective, coordinated behaviour across the tissue. A paradigm for such systems is boron uptake, known for its directional transport and homeostasis, as boron is essential for plant growth but toxic at high concentrations. In Arabidopsis thaliana, boron uptake occurs via diffusion facilitators (NIPs) and exporters (BORs), each presenting distinct polarity. Intriguingly, although boron soil concentrations are homogenous and stable, both transporters manifest strikingly swift boron-dependent regulation. Through mathematical modelling, we demonstrate that slower regulation of these transporters leads to physiologically detrimental oscillatory behaviour. Cells become periodically exposed to potentially cytotoxic boron levels, and nutrient throughput to the xylem becomes hampered. We conclude that, while maintaining homeostasis, swift transporter regulation within a polarised tissue context is critical to prevent intrinsic traffic-jam like behaviour of nutrient flow. PMID:28870285

COMMD1 is linked to the WASH complex and regulates endosomal trafficking of the copper transporter ATP7A

NARCIS (Netherlands)

Phillips-Krawczak, Christine A.; Singla, Amika; Starokadomskyy, Petro; Deng, Zhihui; Osborne, Douglas G.; Li, Haiying; Dick, Christopher J.; Gomez, Timothy S.; Koenecke, Megan; Zhang, Jin-San; Dai, Haiming; Sifuentes-Dominguez, Luis F.; Geng, Linda N.; Kaufmann, Scott H.; Hein, Marco Y.; Wallis, Mathew; McGaughran, Julie; Gecz, Jozef; De Sluis, Bart van; Billadeau, Daniel D.; Burstein, Ezra

2015-01-01

COMMD1 deficiency results in defective copper homeostasis, but the mechanism for this has remained elusive. Here we report that COMMD1 is directly linked to early endosomes through its interaction with a protein complex containing CCDC22, CCDC93, and C16orf62. This COMMD/CCDC22/CCDC93 (CCC) complex

Hepatitis C Virus Replication Depends on Endosomal Cholesterol Homeostasis.

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Stoeck, Ina Karen; Lee, Ji-Young; Tabata, Keisuke; Romero-Brey, Inés; Paul, David; Schult, Philipp; Lohmann, Volker; Kaderali, Lars; Bartenschlager, Ralf

2018-01-01

Similar to other positive-strand RNA viruses, hepatitis C virus (HCV) causes massive rearrangements of intracellular membranes, resulting in a membranous web (MW) composed of predominantly double-membrane vesicles (DMVs), the presumed sites of RNA replication. DMVs are enriched for cholesterol, but mechanistic details on the source and recruitment of cholesterol to the viral replication organelle are only partially known. Here we focused on selected lipid transfer proteins implicated in direct lipid transfer at various endoplasmic reticulum (ER)-membrane contact sites. RNA interference (RNAi)-mediated knockdown identified several hitherto unknown HCV dependency factors, such as steroidogenic acute regulatory protein-related lipid transfer domain protein 3 (STARD3), oxysterol-binding protein-related protein 1A and -B (OSBPL1A and -B), and Niemann-Pick-type C1 (NPC1), all residing at late endosome and lysosome membranes and required for efficient HCV RNA replication but not for replication of the closely related dengue virus. Focusing on NPC1, we found that knockdown or pharmacological inhibition caused cholesterol entrapment in lysosomal vesicles concomitant with decreased cholesterol abundance at sites containing the viral replicase factor NS5A. In untreated HCV-infected cells, unesterified cholesterol accumulated at the perinuclear region, partially colocalizing with NS5A at DMVs, arguing for NPC1-mediated endosomal cholesterol transport to the viral replication organelle. Consistent with cholesterol being an important structural component of DMVs, reducing NPC1-dependent endosomal cholesterol transport impaired MW integrity. This suggests that HCV usurps lipid transfer proteins, such as NPC1, at ER-late endosome/lysosome membrane contact sites to recruit cholesterol to the viral replication organelle, where it contributes to MW functionality. IMPORTANCE A key feature of the replication of positive-strand RNA viruses is the rearrangement of the host cell

A FYVE zinc finger domain protein specifically links mRNA transport to endosome trafficking

Science.gov (United States)

Pohlmann, Thomas; Baumann, Sebastian; Haag, Carl; Albrecht, Mario; Feldbrügge, Michael

2015-01-01

An emerging theme in cellular logistics is the close connection between mRNA and membrane trafficking. A prominent example is the microtubule-dependent transport of mRNAs and associated ribosomes on endosomes. This coordinated process is crucial for correct septin filamentation and efficient growth of polarised cells, such as fungal hyphae. Despite detailed knowledge on the key RNA-binding protein and the molecular motors involved, it is unclear how mRNAs are connected to membranes during transport. Here, we identify a novel factor containing a FYVE zinc finger domain for interaction with endosomal lipids and a new PAM2-like domain required for interaction with the MLLE domain of the key RNA-binding protein. Consistently, loss of this FYVE domain protein leads to specific defects in mRNA, ribosome, and septin transport without affecting general functions of endosomes or their movement. Hence, this is the first endosomal component specific for mRNP trafficking uncovering a new mechanism to couple mRNPs to endosomes. DOI: http://dx.doi.org/10.7554/eLife.06041.001 PMID:25985087

Role of the Small GTPase Rho3 in Golgi/Endosome trafficking through functional interaction with adaptin in Fission Yeast.

Directory of Open Access Journals (Sweden)

Ayako Kita

Full Text Available BACKGROUND: We had previously identified the mutant allele of apm1(+ that encodes a homolog of the mammalian µ1A subunit of the clathrin-associated adaptor protein-1 (AP-1 complex, and we demonstrated the role of Apm1 in Golgi/endosome trafficking, secretion, and vacuole fusion in fission yeast. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we isolated rho3(+, which encodes a Rho-family small GTPase, an important regulator of exocystosis, as a multicopy-suppressor of the temperature-sensitive growth of the apm1-1 mutant cells. Overexpression of Rho3 suppressed the Cl(- sensitivity and immunosuppressant sensitivity of the apm1-1 mutant cells. Overexpression of Rho3 also suppressed the fragmentation of vacuoles, and the accumulation of v-SNARE Syb1 in Golgi/endosomes and partially suppressed the defective secretion associated with apm1-deletion cells. Notably, electron microscopic observation of the rho3-deletion cells revealed the accumulation of abnormal Golgi-like structures, vacuole fragmentation, and accumulation of secretory vesicles; these phenotypes were very similar to those of the apm1-deletion cells. Furthermore, the rho3-deletion cells and apm1-deletion cells showed very similar phenotypic characteristics, including the sensitivity to the immunosuppressant FK506, the cell wall-damaging agent micafungin, Cl(-, and valproic acid. Green fluorescent protein (GFP-Rho3 was localized at Golgi/endosomes as well as the plasma membrane and division site. Finally, Rho3 was shown to form a complex with Apm1 as well as with other subunits of the clathrin-associated AP-1 complex in a GTP- and effector domain-dependent manner. CONCLUSIONS/SIGNIFICANCE: Taken together, our findings reveal a novel role of Rho3 in the regulation of Golgi/endosome trafficking and suggest that clathrin-associated adaptor protein-1 and Rho3 co-ordinate in intracellular transport in fission yeast. To the best of our knowledge, this study provides the first evidence

Overexpression of Rab22a hampers the transport between endosomes and the Golgi apparatus

International Nuclear Information System (INIS)

Mesa, Rosana; Magadan, Javier; Barbieri, Alejandro; Lopez, Cecilia; Stahl, Philip D.; Mayorga, Luis S.

2005-01-01

The transport and sorting of soluble and membrane-associated macromolecules arriving at endosomal compartments require a complex set of Rab proteins. Rab22a has been localized to the endocytic compartment; however, very little is known about the function of Rab22a and inconsistent results have been reported in studies performed in different cell lines. To characterize the function of Rab22a in endocytic transport, the wild-type protein (Rab22a WT), a hydrolysis-deficient mutant (Rab22a Q64L), and a mutant with reduced affinity for GTP (Rab22a S19N) were expressed in CHO cells. None of the three Rab22a constructs affected the transport of rhodamine-dextran to lysosomes, the digestion of internalized proteins, or the lysosomal localization of cathepsin D. In contrast with the mild effect of Rab22a on the endosome-lysosome route, cells expressing Rab22a WT and Rab22a Q64L presented a strong delay in the retrograde transport of cholera toxin from endosomes to the Golgi apparatus. Moreover, these cells accumulated the cation independent mannose 6-phosphate receptor in endosomes. These observations indicate that Rab22a can affect the trafficking from endosomes to the Golgi apparatus probably by promoting fusion among endosomes and impairing the proper segregation of membrane domains required for targeting to the trans-Golgi network (TGN)

Traffic Light Options

DEFF Research Database (Denmark)

Jørgensen, Peter Løchte

This paper introduces, prices, and analyzes traffic light options. The traffic light option is an innovative structured OTC derivative developed independently by several London-based investment banks to suit the needs of Danish life and pension (L&P) companies, which must comply with the traffic...... 2006, and supervisory authorities in many other European countries have implemented similar regulation. Traffic light options are therefore likely to attract the attention of a wider audience of pension fund managers in the future. Focusing on the valuation of the traffic light option we set up a Black...... light scenarios. These stress scenarios entail drops in interest rates as well as in stock prices, and traffic light options are thus designed to pay off and preserve sufficient capital when interest rates and stock prices fall simultaneously. Sweden's FSA implemented a traffic light system in January...

Traffic Light Options

DEFF Research Database (Denmark)

Jørgensen, Peter Løchte

2007-01-01

This paper introduces, prices, and analyzes traffic light options. The traffic light option is an innovative structured OTC derivative developed independently by several London-based investment banks to suit the needs of Danish life and pension (L&P) companies, which must comply with the traffic...... 2006, and supervisory authorities in many other European countries have implemented similar regulation. Traffic light options are therefore likely to attract the attention of a wider audience of pension fund managers in the future. Focusing on the valuation of the traffic light option we set up a Black...... light scenarios. These stress scenarios entail drops in interest rates as well as in stock prices, and traffic light options are thus designed to pay off and preserve sufficient capital when interest rates and stock prices fall simultaneously. Sweden's FSA implemented a traffic light system in January...

The late endosome/lysosome-anchored p18-mTORC1 pathway controls terminal maturation of lysosomes

Energy Technology Data Exchange (ETDEWEB)

Takahashi, Yusuke; Nada, Shigeyuki; Mori, Shunsuke; Soma-Nagae, Taeko; Oneyama, Chitose [Department of Oncogene Research, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871 (Japan); Okada, Masato, E-mail: okadam@biken.osaka-u.ac.jp [Department of Oncogene Research, Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871 (Japan)

2012-01-27

Highlights: Black-Right-Pointing-Pointer p18 is a membrane adaptor that anchors mTORC1 to late endosomes/lysosomes. Black-Right-Pointing-Pointer We examine the role of the p18-mTORC1 pathway in lysosome biogenesis. Black-Right-Pointing-Pointer The loss of p18 causes accumulation of intact late endosomes by arresting lysosome maturation. Black-Right-Pointing-Pointer Inhibition of mTORC1 activity with rapamycin phenocopies the defects of p18 loss. Black-Right-Pointing-Pointer The p18-mTORC1 pathway plays crucial roles in the terminal maturation of lysosomes. -- Abstract: The late endosome/lysosome membrane adaptor p18 (or LAMTOR1) serves as an anchor for the mammalian target of rapamycin complex 1 (mTORC1) and is required for its activation on lysosomes. The loss of p18 causes severe defects in cell growth as well as endosome dynamics, including membrane protein transport and lysosome biogenesis. However, the mechanisms underlying these effects on lysosome biogenesis remain unknown. Here, we show that the p18-mTORC1 pathway is crucial for terminal maturation of lysosomes. The loss of p18 causes aberrant intracellular distribution and abnormal sizes of late endosomes/lysosomes and an accumulation of late endosome specific components, including Rab7, RagC, and LAMP1; this suggests that intact late endosomes accumulate in the absence of p18. These defects are phenocopied by inhibiting mTORC1 activity with rapamycin. Loss of p18 also suppresses the integration of late endosomes and lysosomes, resulting in the defective degradation of tracer proteins. These results suggest that the p18-mTORC1 pathway plays crucial roles in the late stages of lysosomal maturation, potentially in late endosome-lysosome fusion, which is required for processing of various macromolecules.

Huntingtin coordinates the dynein-mediated dynamic positioning of endosomes and lysosomes

Science.gov (United States)

Caviston, Juliane P.; Zajac, Allison L.; Tokito, Mariko; Holzbaur, Erika L.F.

2011-01-01

Huntingtin (Htt) is a membrane-associated scaffolding protein that interacts with microtubule motors as well as actin-associated adaptor molecules. We examined a role for Htt in the dynein-mediated intracellular trafficking of endosomes and lysosomes. In HeLa cells depleted of either Htt or dynein, early, recycling, and late endosomes (LE)/lysosomes all become dispersed. Despite altered organelle localization, kinetic assays indicate only minor defects in intracellular trafficking. Expression of full-length Htt is required to restore organelle localization in Htt-depleted cells, supporting a role for Htt as a scaffold that promotes functional interactions along its length. In dynein-depleted cells, LE/lysosomes accumulate in tight patches near the cortex, apparently enmeshed by cortactin-positive actin filaments; Latrunculin B-treatment disperses these patches. Peripheral LE/lysosomes in dynein-depleted cells no longer colocalize with microtubules. Htt may be required for this off-loading, as the loss of microtubule association is not seen in Htt-depleted cells or in cells depleted of both dynein and Htt. Inhibition of kinesin-1 relocalizes peripheral LE/lysosomes induced by Htt depletion but not by dynein depletion, consistent with their detachment from microtubules upon dynein knockdown. Together, these data support a model of Htt as a facilitator of dynein-mediated trafficking that may regulate the cytoskeletal association of dynamic organelles. PMID:21169558

Endosomal gene expression: a new indicator for prostate cancer patient prognosis?

LENUS (Irish Health Repository)

Johnson, Ian R D

2015-11-10

Prostate cancer continues to be a major cause of morbidity and mortality in men, but a method for accurate prognosis in these patients is yet to be developed. The recent discovery of altered endosomal biogenesis in prostate cancer has identified a fundamental change in the cell biology of this cancer, which holds great promise for the identification of novel biomarkers that can predict disease outcomes. Here we have identified significantly altered expression of endosomal genes in prostate cancer compared to non-malignant tissue in mRNA microarrays and confirmed these findings by qRT-PCR on fresh-frozen tissue. Importantly, we identified endosomal gene expression patterns that were predictive of patient outcomes. Two endosomal tri-gene signatures were identified from a previously published microarray cohort and had a significant capacity to stratify patient outcomes. The expression of APPL1, RAB5A, EEA1, PDCD6IP, NOX4 and SORT1 were altered in malignant patient tissue, when compared to indolent and normal prostate tissue. These findings support the initiation of a case-control study using larger cohorts of prostate tissue, with documented patient outcomes, to determine if different combinations of these new biomarkers can accurately predict disease status and clinical progression in prostate cancer patients.

Increased expression of endosomal members of toll-like receptor family abrogates wound healing in patients with type 2 diabetes mellitus.

Science.gov (United States)

Singh, Kanhaiya; Agrawal, Neeraj K; Gupta, Sanjeev K; Mohan, Gyanendra; Chaturvedi, Sunanda; Singh, Kiran

2016-10-01

The inflammatory phase of wound healing cascade is an important determinant of the fate of the wound. Acute inflammation is necessary to initiate proper wound healing, while chronic inflammation abrogates wound healing. Different endosomal members of toll-like receptor (TLR) family initiate inflammatory signalling via a range of different inflammatory mediators such as interferons, internal tissue damaged-associated molecular patterns (DAMPs) and hyperactive effector T cells. Sustained signalling of TLR9 and TLR7 contributes to chronic inflammation by activating the plasmacytoid dendritic cells. Diabetic wounds are also characterised by sustained inflammatory phase. The objective of this study was to analyse the differential expression of endosomal TLRs in human diabetic wounds compared with control wounds. We analysed the differential expression of TLR7 and TLR9 both at transcriptional and translational levels in wounds of 84 patients with type 2 diabetes mellitus (T2DM) and 6 control subjects without diabetes using quantitative real-time polymerase chain reaction (RT-PCR), western blot and immunohistochemistry. TLR7 and TLR9 were significantly up-regulated in wounds of the patients with T2DM compared with the controls and were dependent on the infection status of the diabetic wounds, and wounds with microbial infection exhibited lower expression levels of endosomal TLRs. Altered endosomal TLR expression in T2DM subjects might be associated with wound healing impairment. © 2015 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

Human Papillomavirus 16 Infection Induces VAP-Dependent Endosomal Tubulation.

Science.gov (United States)

Siddiqa, Abida; Massimi, Paola; Pim, David; Broniarczyk, Justyna; Banks, Lawrence

2018-03-15

Human papillomavirus (HPV) infection involves complex interactions with the endocytic transport machinery, which ultimately facilitates the entry of the incoming viral genomes into the trans -Golgi network (TGN) and their subsequent nuclear entry during mitosis. The endosomal pathway is a highly dynamic intracellular transport system, which consists of vesicular compartments and tubular extensions, although it is currently unclear whether incoming viruses specifically alter the endocytic machinery. In this study, using MICAL-L1 as a marker for tubulating endosomes, we show that incoming HPV-16 virions induce a profound alteration in global levels of endocytic tubulation. In addition, we also show a critical requirement for the endoplasmic reticulum (ER)-anchored protein VAP in this process. VAP plays an essential role in actin nucleation and endosome-to-Golgi transport. Indeed, the loss of VAP results in a dramatic decrease in the level of endosomal tubulation induced by incoming HPV-16 virions. This is also accompanied by a marked reduction in virus infectivity. In VAP knockdown cells, we see that the defect in virus trafficking occurs after capsid disassembly but prior to localization at the trans -Golgi network, with the incoming virion-transduced DNA accumulating in Vps29/TGN46-positive hybrid vesicles. Taken together, these studies demonstrate that infection with HPV-16 virions induces marked alterations of endocytic transport pathways, some of which are VAP dependent and required for the endosome-to-Golgi transport of the incoming viral L2/DNA complex. IMPORTANCE Human papillomavirus infectious entry involves multiple interactions with the endocytic transport machinery. In this study, we show that incoming HPV-16 virions induce a dramatic increase in endocytic tubulation. This tubulation requires ER-associated VAP, which plays a critical role in ensuring the delivery of cargoes from the endocytic compartments to the trans -Golgi network. Indeed, the loss of

Human ClC-6 is a late endosomal glycoprotein that associates with detergent-resistant lipid domains.

Directory of Open Access Journals (Sweden)

Sofie Ignoul

Full Text Available BACKGROUND: The mammalian CLC protein family comprises nine members (ClC-1 to -7 and ClC-Ka, -Kb that function either as plasma membrane chloride channels or as intracellular chloride/proton antiporters, and that sustain a broad spectrum of cellular processes, such as membrane excitability, transepithelial transport, endocytosis and lysosomal degradation. In this study we focus on human ClC-6, which is structurally most related to the late endosomal/lysomal ClC-7. PRINCIPAL FINDINGS: Using a polyclonal affinity-purified antibody directed against a unique epitope in the ClC-6 COOH-terminal tail, we show that human ClC-6, when transfected in COS-1 cells, is N-glycosylated in a region that is evolutionary poorly conserved between mammalian CLC proteins and that is located between the predicted helices K and M. Three asparagine residues (N410, N422 and N432 have been defined by mutagenesis as acceptor sites for N-glycosylation, but only two of the three sites seem to be simultaneously N-glycosylated. In a differentiated human neuroblastoma cell line (SH-SY5Y, endogenous ClC-6 colocalizes with LAMP-1, a late endosomal/lysosomal marker, but not with early/recycling endosomal markers such as EEA-1 and transferrin receptor. In contrast, when transiently expressed in COS-1 or HeLa cells, human ClC-6 mainly overlaps with markers for early/recycling endosomes (transferrin receptor, EEA-1, Rab5, Rab4 and not with late endosomal/lysosomal markers (LAMP-1, Rab7. Analogously, overexpression of human ClC-6 in SH-SY5Y cells also leads to an early/recycling endosomal localization of the exogenously expressed ClC-6 protein. Finally, in transiently transfected COS-1 cells, ClC-6 copurifies with detergent-resistant membrane fractions, suggesting its partitioning in lipid rafts. Mutating a juxtamembrane string of basic amino acids (amino acids 71-75: KKGRR disturbs the association with detergent-resistant membrane fractions and also affects the segregation of ClC-6

Natural Modulators of Endosomal Toll-Like Receptor-Mediated Psoriatic Skin Inflammation

Directory of Open Access Journals (Sweden)

Chao-Yang Lai

2017-01-01

Full Text Available Psoriasis is a chronic inflammatory autoimmune disease that can be initiated by excessive activation of endosomal toll-like receptors (TLRs, particularly TLR7, TLR8, and TLR9. Therefore, inhibitors of endosomal TLR activation are being investigated for their ability to treat this disease. The currently approved biological drugs adalimumab, etanercept, infliximab, ustekinumab, ixekizumab, and secukizumab are antibodies against effector cytokines that participate in the initiation and development of psoriasis. Several immune modulatory oligonucleotides and small molecular weight compounds, including IMO-3100, IMO-8400, and CPG-52364, that block the interaction between endosomal TLRs and their ligands are under clinical investigation for their effectiveness in the treatment of psoriasis. In addition, several chemical compounds, including AS-2444697, PF-05387252, PF-05388169, PF-06650833, ML120B, and PHA-408, can inhibit TLR signaling. Although these compounds have demonstrated anti-inflammatory activity in animal models, their therapeutic potential for the treatment of psoriasis has not yet been tested. Recent studies demonstrated that natural compounds derived from plants, fungi, and bacteria, including mustard seed, Antrodia cinnamomea extract, curcumin, resveratrol, thiostrepton, azithromycin, and andrographolide, inhibited psoriasis-like inflammation induced by the TLR7 agonist imiquimod in animal models. These natural modulators employ different mechanisms to inhibit endosomal TLR activation and are administered via different routes. Therefore, they represent candidate psoriasis drugs and might lead to the development of new treatment options.

Promyelocytic leukemia bodies tether to early endosomes during mitosis.

Science.gov (United States)

Palibrk, Vuk; Lång, Emma; Lång, Anna; Schink, Kay Oliver; Rowe, Alexander D; Bøe, Stig Ove

2014-01-01

During mitosis the nuclear envelope breaks down, leading to potential interactions between cytoplasmic and nuclear components. PML bodies are nuclear structures with tumor suppressor and antiviral functions. Early endosomes, on the other hand, are cytoplasmic vesicles involved in transport and growth factor signaling. Here we demonstrate that PML bodies form stable interactions with early endosomes immediately following entry into mitosis. The 2 compartments remain stably associated throughout mitosis and dissociate in the cytoplasm of newly divided daughter cells. We also show that a minor subset of PML bodies becomes anchored to the mitotic spindle poles during cell division. The study demonstrates a stable mitosis-specific interaction between a cytoplasmic and a nuclear compartment.

The late endosome/lysosome-anchored p18-mTORC1 pathway controls terminal maturation of lysosomes

International Nuclear Information System (INIS)

Takahashi, Yusuke; Nada, Shigeyuki; Mori, Shunsuke; Soma-Nagae, Taeko; Oneyama, Chitose; Okada, Masato

2012-01-01

Highlights: ► p18 is a membrane adaptor that anchors mTORC1 to late endosomes/lysosomes. ► We examine the role of the p18-mTORC1 pathway in lysosome biogenesis. ► The loss of p18 causes accumulation of intact late endosomes by arresting lysosome maturation. ► Inhibition of mTORC1 activity with rapamycin phenocopies the defects of p18 loss. ► The p18-mTORC1 pathway plays crucial roles in the terminal maturation of lysosomes. -- Abstract: The late endosome/lysosome membrane adaptor p18 (or LAMTOR1) serves as an anchor for the mammalian target of rapamycin complex 1 (mTORC1) and is required for its activation on lysosomes. The loss of p18 causes severe defects in cell growth as well as endosome dynamics, including membrane protein transport and lysosome biogenesis. However, the mechanisms underlying these effects on lysosome biogenesis remain unknown. Here, we show that the p18-mTORC1 pathway is crucial for terminal maturation of lysosomes. The loss of p18 causes aberrant intracellular distribution and abnormal sizes of late endosomes/lysosomes and an accumulation of late endosome specific components, including Rab7, RagC, and LAMP1; this suggests that intact late endosomes accumulate in the absence of p18. These defects are phenocopied by inhibiting mTORC1 activity with rapamycin. Loss of p18 also suppresses the integration of late endosomes and lysosomes, resulting in the defective degradation of tracer proteins. These results suggest that the p18-mTORC1 pathway plays crucial roles in the late stages of lysosomal maturation, potentially in late endosome–lysosome fusion, which is required for processing of various macromolecules.

Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery

Science.gov (United States)

Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

2015-06-01

The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds’ escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds’ cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier.

Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery

Science.gov (United States)

Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

2015-01-01

The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds’ escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds’ cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier. PMID:26123532

Rapid endosomal escape of prickly nanodiamonds: implications for gene delivery.

Science.gov (United States)

Chu, Zhiqin; Miu, Kaikei; Lung, Pingsai; Zhang, Silu; Zhao, Saisai; Chang, Huan-Cheng; Lin, Ge; Li, Quan

2015-06-30

The prickly nanodiamonds easily entered cells via endocytosis followed by unique intracellular translocation characteristics—quick endosomal escape followed by stable residence in cytoplasm. Endosomal membrane rupturing is identified as the major route of nanodiamonds' escaping the vesicle confinement and to the cytoplasm. Little cytotoxicity is observed to associate with the nanodiamonds' cytosolic release. Such features enable its application for gene delivery, which requires both effective cellular uptake and cytosolic release of the gene. Taking green fluorescent protein gene as an example, we demonstrate the successful cytosolic delivery and expression of such a gene using the prickly nanodiamonds as carrier.

Enhancing endosomal escape of transduced proteins by photochemical internalisation.

Directory of Open Access Journals (Sweden)

Kevin Mellert

Full Text Available Induced internalisation of functional proteins into cultured cells has become an important aspect in a rising number of in vitro and in vivo assays. The endo-lysosomal entrapment of the transduced proteins remains the major problem in all transduction protocols. In this study we compared the efficiency, cytotoxicity and protein targeting of different commercially available transduction reagents by transducing a well-studied fluorescently labelled protein (Atto488-bovine serum albumin into cultured human sarcoma cells. The amount of internalised protein and toxicity differed between the different reagents, but the percentage of transduced cells was consistently high. Furthermore, in all protocols the signals of the transduced Atto488-BSA were predominantly punctual consistent with an endosomal localisation. To overcome the endosomal entrapment, the transduction protocols were combined with a photochemical internalisation (PCI treatment. Using this combination revealed that an endosomal disruption is highly effective in cell penetrating peptide (CPP mediated transduction, whereas lipid-mediated transductions lead to a lower signal spreading throughout the cytosol. No change in the signal distribution could be achieved in treatments using non-lipid polymers as a transduction reagent. Therefore, the combination of protein transduction protocols based on CPPs with the endosomolytic treatment PCI can facilitate protein transduction experiments in vitro.

Enhancing endosomal escape of transduced proteins by photochemical internalisation.

Science.gov (United States)

Mellert, Kevin; Lamla, Markus; Scheffzek, Klaus; Wittig, Rainer; Kaufmann, Dieter

2012-01-01

Induced internalisation of functional proteins into cultured cells has become an important aspect in a rising number of in vitro and in vivo assays. The endo-lysosomal entrapment of the transduced proteins remains the major problem in all transduction protocols. In this study we compared the efficiency, cytotoxicity and protein targeting of different commercially available transduction reagents by transducing a well-studied fluorescently labelled protein (Atto488-bovine serum albumin) into cultured human sarcoma cells. The amount of internalised protein and toxicity differed between the different reagents, but the percentage of transduced cells was consistently high. Furthermore, in all protocols the signals of the transduced Atto488-BSA were predominantly punctual consistent with an endosomal localisation. To overcome the endosomal entrapment, the transduction protocols were combined with a photochemical internalisation (PCI) treatment. Using this combination revealed that an endosomal disruption is highly effective in cell penetrating peptide (CPP) mediated transduction, whereas lipid-mediated transductions lead to a lower signal spreading throughout the cytosol. No change in the signal distribution could be achieved in treatments using non-lipid polymers as a transduction reagent. Therefore, the combination of protein transduction protocols based on CPPs with the endosomolytic treatment PCI can facilitate protein transduction experiments in vitro.

Histone deacetylase-mediated regulation of endolysosomal pH.

Science.gov (United States)

Prasad, Hari; Rao, Rajini

2018-05-04

The pH of the endolysosomal system is tightly regulated by a balance of proton pump and leak mechanisms that are critical for storage, recycling, turnover, and signaling functions in the cell. Dysregulation of endolysosomal pH has been linked to aging, amyloidogenesis, synaptic dysfunction, and various neurodegenerative disorders, including Alzheimer's disease. Therefore, understanding the mechanisms that regulate luminal pH may be key to identifying new targets for managing these disorders. Meta-analysis of yeast microarray databases revealed that nutrient-limiting conditions inhibited the histone deacetylase (HDAC) Rpd3 and thereby up-regulated transcription of the endosomal Na + /H + exchanger Nhx1, resulting in vacuolar alkalinization. Consistent with these findings, Rpd3 inhibition by the HDAC inhibitor and antifungal drug trichostatin A induced Nhx1 expression and vacuolar alkalinization. Bioinformatics analysis of Drosophila and mouse databases revealed that caloric control of the Nhx1 orthologs DmNHE3 and NHE6, respectively, is also mediated by HDACs. We show that NHE6 is a target of the transcription factor cAMP-response element-binding protein (CREB), a known regulator of cellular responses to low-nutrient conditions, providing a molecular mechanism for nutrient- and HDAC-dependent regulation of endosomal pH. Of note, pharmacological targeting of the CREB pathway to increase NHE6 expression helped regulate endosomal pH and correct defective clearance of amyloid Aβ in an apoE4 astrocyte model of Alzheimer's disease. These observations from yeast, fly, mouse, and cell culture models point to an evolutionarily conserved mechanism for HDAC-mediated regulation of endosomal NHE expression. Our insights offer new therapeutic strategies for modulation of endolysosomal pH in fungal infection and human disease. © 2018 Prasad and Rao.

75 FR 48625 - Amendment to the International Traffic in Arms Regulations: Dual Nationals and Third-Country...

Science.gov (United States)

2010-08-11

... previous policy regarding the treatment of dual nationals and foreign nationals was reconsidered. The... technology. Regulatory Analysis and Notices Administrative Procedure Act This amendment involves a foreign... amend the International Traffic in Arms Regulations (ITAR) to update the policies regarding end-user...

Integrin-linked kinase and ELMO2 modulate recycling endosomes in keratinocytes.

Science.gov (United States)

Ho, Ernest; Ivanova, Iordanka A; Dagnino, Lina

2016-12-01

The formation of tight cell-cell junctions is essential in the epidermis for its barrier properties. In this tissue, keratinocytes follow a differentiation program tightly associated with their movement from the innermost basal to the outer suprabasal layers, and with changes in their cell-cell adhesion profile. Intercellular adhesion in keratinocytes is mediated through cell-cell contacts, including E-cadherin-based adherens junctions. Although the mechanisms that mediate E-cadherin delivery to the plasma membrane have been widely studied in simple epithelia, this process is less well understood in the stratified epidermis. In this study, we have investigated the role of Engulfment and Cell Motility 2 (ELMO2) and integrin-linked kinase (ILK) in the positioning of E-cadherin-containing recycling endosomes during establishment of cell-cell contacts in differentiating keratinocytes. We now show that induction of keratinocyte differentiation by Ca 2+ is accompanied by localization of ELMO2 and ILK to Rab4- and Rab11a-containing recycling endosomes. The positioning of long-loop Rab11a-positive endosomes at areas adjacent to cell-cell contacts is disrupted in ELMO2- or ILK-deficient keratinocytes, and is associated with impaired localization of E-cadherin to cell borders. Our studies show a previously unrecognized role for ELMO2 and ILK in modulation of endosomal positioning, which may play key roles in epidermal sheet maintenance and permeability barrier function. Copyright © 2016 Elsevier B.V. All rights reserved.

Regulatory measures for traffic safety

International Nuclear Information System (INIS)

Veerapur, R.D.; Bharambe, S.D.; Patnaik, S.K.; Tandle, A.K.; Sonawane, K.A.; Kumar, Rajesh; Venkat Subramanian, K.

2017-01-01

Traffic safety is an issue related to occupational safety not restricted alone to the transportation but extends beyond. BARC has many facilities spread across large area in Mumbai and outside Mumbai. BARC deploys large number of buses, mini buses, jeeps and cars for commuting its employees to reach BARC and for commuting within BARC premises. Additionally, trucks, fire tenders, trailers etc. are also deployed for transportation of materials. No moving vehicle is ever free of the possibility of involvement in an accident. Vehicular accidents and the fatalities on road are the result of inter-play of a number of factors. The vehicle population has been steadily increasing with the pace picking up significantly in recent past. Increase in vehicle population in the face of limited road space used by a large variety of traffic has heightened the need and urgency for a well-thought-out road safety. Therefore, existence of regulatory authority to regulate traffic and vehicles to ensure safety of its employees and vehicles is very essential. BARC Traffic Safety Committee (BTSC), which is the regulating body for traffic safety is responsible for ensuring overall traffic safety. (author)

Thermoresponsive pegylated bubble liposome nanovectors for efficient siRNA delivery via endosomal escape

KAUST Repository

Alamoudi, Kholod

2017-05-19

Improving the delivery of siRNA into cancer cells via bubble liposomes. Designing a thermoresponsive pegylated liposome through the introduction of ammonium bicarbonate salt into liposomes so as to control their endosomal escape for gene therapy.A sub-200 nm nanovector was fully characterized and examined for cellular uptake, cytotoxicity, endosomal escape and gene silencing.The siRNA-liposomes were internalized into cancer cells within 5 min and then released siRNAs in the cytosol prior to lysosomal degradation upon external temperature elevation. This was confirmed by confocal bioimaging and gene silencing reaching up to 90% and further demonstrated by the protein inhibition of both target genes.The thermoresponsiveness of ammonium bicarbonate containing liposomes enabled the rapid endosomal escape of the particles and resulted in an efficient gene silencing.

Thermoresponsive pegylated bubble liposome nanovectors for efficient siRNA delivery via endosomal escape

KAUST Repository

Alamoudi, Kholod; Martins, Patricia; Croissant, Jonas G.; Patil, Sachin; Omar, Haneen; Khashab, Niveen M.

2017-01-01

Improving the delivery of siRNA into cancer cells via bubble liposomes. Designing a thermoresponsive pegylated liposome through the introduction of ammonium bicarbonate salt into liposomes so as to control their endosomal escape for gene therapy.A sub-200 nm nanovector was fully characterized and examined for cellular uptake, cytotoxicity, endosomal escape and gene silencing.The siRNA-liposomes were internalized into cancer cells within 5 min and then released siRNAs in the cytosol prior to lysosomal degradation upon external temperature elevation. This was confirmed by confocal bioimaging and gene silencing reaching up to 90% and further demonstrated by the protein inhibition of both target genes.The thermoresponsiveness of ammonium bicarbonate containing liposomes enabled the rapid endosomal escape of the particles and resulted in an efficient gene silencing.

Cystatin F as a regulator of immune cell cytotoxicity.

Science.gov (United States)

Kos, Janko; Nanut, Milica Perišić; Prunk, Mateja; Sabotič, Jerica; Dautović, Esmeralda; Jewett, Anahid

2018-05-10

Cysteine cathepsins are lysosomal peptidases involved in the regulation of innate and adaptive immune responses. Among the diverse processes, regulation of granule-dependent cytotoxicity of cytotoxic T-lymphocytes (CTLs) and natural killer (NK) cells during cancer progression has recently gained significant attention. The function of cysteine cathepsins is regulated by endogenous cysteine protease inhibitors-cystatins. Whereas other cystatins are generally cytosolic or extracellular proteins, cystatin F is present in endosomes and lysosomes and is thus able to regulate the activity of its target directly. It is delivered to endosomal/lysosomal vesicles as an inactive, disulphide-linked dimer. Proteolytic cleavage of its N-terminal part leads to the monomer, the only form that is a potent inhibitor of cathepsins C, H and L, involved in the activation of granzymes and perforin. In NK cells and CTLs the levels of active cathepsin C and of granzyme B are dependent on the concentration of monomeric, active cystatin F. In tumour microenvironment, inactive dimeric cystatin F can be secreted from tumour cells or immune cells and further taken up by the cytotoxic cells. Subsequent monomerization and inhibition of cysteine cathepsins within the endosomal/lysosomal vesicles impairs granzyme and perforin activation, and provokes cell anergy. Further, the glycosylation pattern has been shown to be important in controlling secretion of cystatin F from target cells, as well as internalization by cytotoxic cells and trafficking to endosomal/lysosomal vesicles. Cystatin F is therefore an important mediator used by bystander cells to reduce NK and T-cell cytotoxicity.

Endocytosis of wheat germ agglutinin binding sites from the cell surface into a tubular endosomal network.

Science.gov (United States)

Raub, T J; Koroly, M J; Roberts, R M

1990-04-01

By using fluorescence and electron microscopy, the endocytic pathway encountered by cell surface components after they had bound wheat germ agglutinin (WGA) was visualized. The majority of these components are thought to consist of sialylated glycoproteins (HMWAG) that represent a subpopulation of the total cell surface proteins but most of the externally disposed plasma membrane proteins of the cell. Examination of semi-thin sections by medium- and high-voltage electron microscopy revealed the three-dimensional organization of vesicular and tubular endosomes. Binding of either fluorescein isothiocyanate-, horseradish peroxidase-, or ferritin-conjugated WGA to cells at 4 degrees C showed that the HMWAG were distributed uniformly over the cell surface. Warming of surface-labeled cells to 37 degrees C resulted in the endocytosis of WGA into peripheral endosomes via invagination of regions of both coated and uncoated membrane. The peripheral endosome appeared as isolated complexes comprising a vesicular element (300-400 nm diam.) surrounded by and continuous with tubular cisternae (45-60 nm diam.), which did not interconnect the endosomes. After 30 min or more label also became localized in a network of anastomosing tubules (45-60 nm diam.) that were located in the centrosomal region of the cell. Endocytosed WGA-HMWAG complexes did not become associated with cisternae of the Golgi apparatus, although tubular and vesicular endosomes were noted in the vicinity of the trans-Golgi region. The accumulation of WGA-HMWAG in the endosomes within the centrosomal region was inhibited when cells were incubated at 18 degrees C. None of these compartments contained acid phosphatase activity, a result that is consistent with other data that the HMWAG do not pass through lysosomes initially. The kinetics of labeling were consistent with the interpretation that recycling of most of the WGA binding surface glycoproteins occurred rapidly from early peripheral endosomes followed by the

Ubiquitin regulates GGA3-mediated degradation of BACE1.

Science.gov (United States)

Kang, Eugene L; Cameron, Andrew N; Piazza, Fabrizio; Walker, Kendall R; Tesco, Giuseppina

2010-07-30

BACE1 (beta-site amyloid precursor protein-cleaving enzyme 1) is a membrane-tethered member of the aspartyl proteases, essential for the production of beta-amyloid, a toxic peptide that accumulates in the brain of subjects affected by Alzheimer disease. The BACE1 C-terminal fragment contains a DXXLL motif that has been shown to bind the VHS (VPS27, Hrs, and STAM) domain of GGA1-3 (Golgi-localized gamma-ear-containing ARF-binding proteins). GGAs are trafficking molecules involved in the transport of proteins containing the DXXLL signal from the Golgi complex to endosomes. Moreover, GGAs bind ubiquitin and traffic synthetic and endosomal ubiquitinated cargoes to lysosomes. We have previously shown that depletion of GGA3 results in increased BACE1 levels and activity because of impaired lysosomal degradation. Here, we report that the accumulation of BACE1 is rescued by the ectopic expression of GGA3 in H4 neuroglioma cells depleted of GGA3. Accordingly, the overexpression of GGA3 reduces the levels of BACE1 and beta-amyloid. We then established that mutations in the GGA3 VPS27, Hrs, and STAM domain (N91A) or in BACE1 di-leucine motif (L499A/L500A), able to abrogate their binding, did not affect the ability of ectopically expressed GGA3 to rescue BACE1 accumulation in cells depleted of GGA3. Instead, we found that BACE1 is ubiquitinated at lysine 501 and is mainly monoubiquitinated and Lys-63-linked polyubiquitinated. Finally, a GGA3 mutant with reduced ability to bind ubiquitin (GGA3L276A) was unable to regulate BACE1 levels both in rescue and overexpression experiments. These findings indicate that levels of GGA3 tightly and inversely regulate BACE1 levels via interaction with ubiquitin sorting machinery.

Ubiquitin Regulates GGA3-mediated Degradation of BACE1*

Science.gov (United States)

Kang, Eugene L.; Cameron, Andrew N.; Piazza, Fabrizio; Walker, Kendall R.; Tesco, Giuseppina

2010-01-01

BACE1 (β-site amyloid precursor protein-cleaving enzyme 1) is a membrane-tethered member of the aspartyl proteases, essential for the production of β-amyloid, a toxic peptide that accumulates in the brain of subjects affected by Alzheimer disease. The BACE1 C-terminal fragment contains a DXXLL motif that has been shown to bind the VHS (VPS27, Hrs, and STAM) domain of GGA1–3 (Golgi-localized γ-ear-containing ARF-binding proteins). GGAs are trafficking molecules involved in the transport of proteins containing the DXXLL signal from the Golgi complex to endosomes. Moreover, GGAs bind ubiquitin and traffic synthetic and endosomal ubiquitinated cargoes to lysosomes. We have previously shown that depletion of GGA3 results in increased BACE1 levels and activity because of impaired lysosomal degradation. Here, we report that the accumulation of BACE1 is rescued by the ectopic expression of GGA3 in H4 neuroglioma cells depleted of GGA3. Accordingly, the overexpression of GGA3 reduces the levels of BACE1 and β-amyloid. We then established that mutations in the GGA3 VPS27, Hrs, and STAM domain (N91A) or in BACE1 di-leucine motif (L499A/L500A), able to abrogate their binding, did not affect the ability of ectopically expressed GGA3 to rescue BACE1 accumulation in cells depleted of GGA3. Instead, we found that BACE1 is ubiquitinated at lysine 501 and is mainly monoubiquitinated and Lys-63-linked polyubiquitinated. Finally, a GGA3 mutant with reduced ability to bind ubiquitin (GGA3L276A) was unable to regulate BACE1 levels both in rescue and overexpression experiments. These findings indicate that levels of GGA3 tightly and inversely regulate BACE1 levels via interaction with ubiquitin sorting machinery. PMID:20484053

Navigators’ Behavior in Traffic Separation Schemes

Directory of Open Access Journals (Sweden)

Zbigniew Pietrzykowski

2015-03-01

Full Text Available One of the areas of decision support in the navigational ship conduct process is a Traffic Separation Scheme. TSSs are established in areas with high traffic density, often near the shore and in port approaches. The main purpose of these schemes is to improve maritime safety by channeling vessel traffic into streams. Traffic regulations as well as ships behavior in real conditions in chosen TSSs have been analyzed in order to develop decision support algorithms.

77 FR 76864 - Amendment to the International Traffic in Arms Regulations: Afghanistan and Change to Policy on...

Science.gov (United States)

2012-12-31

... Belarus, Cuba, Eritrea, Iran, North Korea, Syria, and Venezuela. This policy also applies to countries... International Traffic in Arms Regulations: Afghanistan and Change to Policy on Prohibited Exports AGENCY.... Goforth, Director, Office of Defense Trade Controls Policy, U.S. Department of State, telephone (202) 663...

CED-10/Rac1 regulates endocytic recycling through the RAB-5 GAP TBC-2.

Directory of Open Access Journals (Sweden)

Lin Sun

Full Text Available Rac1 is a founding member of the Rho-GTPase family and a key regulator of membrane remodeling. In the context of apoptotic cell corpse engulfment, CED-10/Rac1 acts with its bipartite guanine nucleotide exchange factor, CED-5/Dock180-CED-12/ELMO, in an evolutionarily conserved pathway to promote phagocytosis. Here we show that in the context of the Caenorhabditis elegans intestinal epithelium CED-10/Rac1, CED-5/Dock180, and CED-12/ELMO promote basolateral recycling. Furthermore, we show that CED-10 binds to the RAB-5 GTPase activating protein TBC-2, that CED-10 contributes to recruitment of TBC-2 to endosomes, and that recycling cargo is trapped in recycling endosomes in ced-12, ced-10, and tbc-2 mutants. Expression of GTPase defective RAB-5(Q78L also traps recycling cargo. Our results indicate that down-regulation of early endosome regulator RAB-5/Rab5 by a CED-5, CED-12, CED-10, TBC-2 cascade is an important step in the transport of cargo through the basolateral recycling endosome for delivery to the plasma membrane.

Characterization of PEBBLEs as a Tool for Real-Time Measurement of Dictyostelium discoideum Endosomal pH

Directory of Open Access Journals (Sweden)

Everett Moding

2009-01-01

Full Text Available The measurement of intracellular ion concentration change is important for understanding the cellular mechanisms for communication. Recently developed nanosensors, (Photonic Explorers for Biomedical use with Biologically Localized Embedding PEBBLEs, have a number of advantages for measuring ions in cells over established methods using microelectrodes, unbound fluorescent dyes, or NMR. PEBBLE sensors have been shown to work in principle for measuring dynamic ion changes, but few in vivo applications have been demonstrated. We modified the protocol for the fabrication of pH sensing PEBBLEs and developed a protocol for the utilization of these sensors for the monitoring of dynamic pH changes in the endosomes of slime mold Dictyostelium discoideum (D. discoideum. Oregon Green 514-CdSe Quantum Dot PEBBLEs were used to measure real-time pH inside D. discoideum endosomes during cAMP stimulation. Endosomal pH was shown to decrease during cAMP signaling, demonstrating a movement of protons into the endosomes of D. discoideum amoebae.

deep-orange and carnation define distinct stages in late endosomal biogenesis in Drosophila melanogaster

OpenAIRE

Sriram, V.; Krishnan, K.S.; Mayor, Satyajit

2003-01-01

Endosomal degradation is severely impaired in primary hemocytes from larvae of eye color mutants of Drosophila. Using high resolution imaging and immunofluorescence microscopy in these cells, products of eye color genes, deep-orange (dor) and carnation (car), are localized to large multivesicular Rab7-positive late endosomes containing Golgi-derived enzymes. These structures mature into small sized Dor-negative, Car-positive structures, which subsequently fuse to form tubular lysosomes. Defec...

A fluorescence resonance energy transfer-based approach for investigating late endosome-lysosome retrograde fusion events.

Science.gov (United States)

Kaufmann, A M; Goldman, S D B; Krise, J P

2009-03-01

Traditionally, lysosomes have been considered to be a terminal endocytic compartment. Recent studies suggest that lysosomes are quite dynamic, being able to fuse with other late endocytic compartments as well as with the plasma membrane. Here we describe a quantitative fluorescence energy transfer (FRET)-based method for assessing rates of retrograde fusion between terminal lysosomes and late endosomes in living cells. Late endosomes were specifically labeled with 800-nm latex beads that were conjugated with streptavidin and Alexa Fluor 555 (FRET donor). Terminal lysosomes were specifically labeled with 10,000-MW dextran polymers conjugated with biotin and Alexa Fluor 647 (FRET acceptor). Following late endosome-lysosome fusion, the strong binding affinity between streptavidin and biotin brought the donor and acceptor fluorophore molecules into close proximity, thereby facilitating the appearance of a FRET emission signal. Because apparent size restrictions in the endocytic pathway do not permit endocytosed latex beads from reaching terminal lysosomes in an anterograde fashion, the appearance of the FRET signal is consistent with retrograde transport of lysosomal cargo back to late endosomes. We assessed the efficiency of this transport step in fibroblasts affected by different lysosome storage disorders-Niemann-Pick type C, mucolipidosis type IV, and Sandhoff's disease, all of which have a similar lysosomal lipid accumulation phenotype. We report here, for the first time, that these disorders can be distinguished by their rate of transfer of lysosome cargos to late endosomes, and we discuss the implications of these findings for developing new therapeutic strategies.

An Inside Job: How Endosomal Na+/H+ Exchangers Link to Autism and Neurological Disease

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Kalyan C. Kondapalli

2014-06-01

Full Text Available Autism imposes a major impediment to childhood development and a huge emotional and financial burden on society. In recent years, there has been rapidly accumulating genetic evidence that links the eNHE, a subset of Na+/H+ exchangers that localize to intracellular vesicles, to a variety of neurological conditions including autism, attention deficit hyperactivity disorder, intellectual disability and epilepsy. By providing a leak pathway for protons pumped by the V-ATPase, eNHE determine luminal pH and regulate cation (Na+, K+ content in early and recycling endosomal compartments. Loss-of-function mutations in eNHE cause hyperacidification of endosomal lumen, as a result of imbalance in pump and leak pathways. Two isoforms, NHE6 and NHE9 are highly expressed in brain, including hippocampus and cortex. Here, we summarize evidence for the importance of luminal cation content and pH on processing, delivery and fate of cargo and on the surface expression and function of membrane receptors and neurotransmitter transporters, drawing upon insights from model organisms and mammalian cells. These studies lead to cellular models of eNHE activity in pre- and post-synaptic neurons and astrocytes, where they could impact synapse development and plasticity. The study of eNHE has provided new insight on the mechanism of autism and other debilitating neurological disorders and opened up new possibilities for therapeutic intervention.

Small Molecules for Early Endosome-Specific Patch Clamping.

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Chen, Cheng-Chang; Butz, Elisabeth S; Chao, Yu-Kai; Grishchuk, Yulia; Becker, Lars; Heller, Stefan; Slaugenhaupt, Susan A; Biel, Martin; Wahl-Schott, Christian; Grimm, Christian

2017-07-20

To resolve the subcellular distribution of endolysosomal ion channels, we have established a novel experimental approach to selectively patch clamp Rab5 positive early endosomes (EE) versus Rab7/LAMP1-positive late endosomes/lysosomes (LE/LY). To functionally characterize ion channels in endolysosomal membranes with the patch-clamp technique, it is important to develop techniques to selectively enlarge the respective organelles. We found here that two small molecules, wortmannin and latrunculin B, enlarge Rab5-positive EE when combined but not Rab7-, LAMP1-, or Rab11 (RE)-positive vesicles. The two compounds act rapidly, specifically, and are readily applicable in contrast to genetic approaches or previously used compounds such as vacuolin, which enlarges EE, RE, and LE/LY. We apply this approach here to measure currents mediated by TRPML channels, in particular TRPML3, which we found to be functionally active in both EE and LE/LY in overexpressing cells as well as in endogenously expressing CD11b+ lung-tissue macrophages. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rab7: roles in membrane trafficking and disease.

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Zhang, Ming; Chen, Li; Wang, Shicong; Wang, Tuanlao

2009-06-01

The endocytosis pathway controls multiple cellular and physiological events. The lysosome is the destination of newly synthesized lysosomal hydrolytic enzymes. Internalized molecules or particles are delivered to the lysosome for degradation through sequential transport along the endocytic pathway. The endocytic pathway is also emerging as a signalling platform, in addition to the well-known role of the plasma membrane for signalling. Rab7 is a late endosome-/lysosome-associated small GTPase, perhaps the only lysosomal Rab protein identified to date. Rab7 plays critical roles in the endocytic processes. Through interaction with its partners (including upstream regulators and downstream effectors), Rab7 participates in multiple regulation mechanisms in endosomal sorting, biogenesis of lysosome [or LRO (lysosome-related organelle)] and phagocytosis. These processes are closely related to substrates degradation, antigen presentation, cell signalling, cell survival and microbial pathogen infection. Consistently, mutations or dysfunctions of Rab7 result in traffic disorders, which cause various diseases, such as neuropathy, cancer and lipid metabolism disease. Rab7 also plays important roles in microbial pathogen infection and survival, as well as in participating in the life cycle of viruses. Here, we give a brief review on the central role of Rab7 in endosomal traffic and summarize the studies focusing on the participation of Rab7 in disease pathogenesis. The underlying mechanism governed by Rab7 and its partners will also be discussed.

Phosphatidylinositol 3,5-Bisphosphate-Rich Membrane Domains in Endosomes and Lysosomes.

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Takatori, Sho; Tatematsu, Tsuyako; Cheng, Jinglei; Matsumoto, Jun; Akano, Takuya; Fujimoto, Toyoshi

2016-02-01

Phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P2 ) has critical functions in endosomes and lysosomes. We developed a method to define nanoscale distribution of PtdIns(3,5)P2 using freeze-fracture electron microscopy. GST-ATG18-4×FLAG was used to label PtdIns(3,5)P2 and its binding to phosphatidylinositol 3-phosphate (PtdIns(3)P) was blocked by an excess of the p40(phox) PX domain. In yeast exposed to hyperosmotic stress, PtdIns(3,5)P2 was concentrated in intramembrane particle (IMP)-deficient domains in the vacuolar membrane, which made close contact with adjacent membranes. The IMP-deficient domain was also enriched with PtdIns(3)P, but was deficient in Vph1p, a liquid-disordered domain marker. In yeast lacking either PtdIns(3,5)P2 or its effector, Atg18p, the IMP-deficient, PtdIns(3)P-rich membranes were folded tightly to make abnormal tubular structures, thus showing where the vacuolar fragmentation process is arrested when PtdIns(3,5)P2 metabolism is defective. In HeLa cells, PtdIns(3,5)P2 was significantly enriched in the vesicular domain of RAB5- and RAB7-positive endosome/lysosomes of the tubulo-vesicular morphology. This biased distribution of PtdIns(3,5)P2 was also observed using fluorescence microscopy, which further showed enrichment of a retromer component, VPS35, in the tubular domain. This is the first report to show segregation of PtdIns(3,5)P2 -rich and -deficient domains in endosome/lysosomes, which should be important for endosome/lysosome functionality. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

G protein-coupled receptor (GPCR) signaling via heterotrimeric G proteins from endosomes.

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Tsvetanova, Nikoleta G; Irannejad, Roshanak; von Zastrow, Mark

2015-03-13

Some G protein-coupled receptors (GPCRs), in addition to activating heterotrimeric G proteins in the plasma membrane, appear to elicit a "second wave" of G protein activation after ligand-induced internalization. We briefly summarize evidence supporting this view and then discuss what is presently known about the functional significance of GPCR-G protein activation in endosomes. Endosomal activation can shape the cellular response temporally by prolonging its overall duration, and may shape the response spatially by moving the location of intracellular second messenger production relative to effectors. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

The N-terminal domains of Vps3 and Vps8 are critical for localization and function of the CORVET tethering complex on endosomes.

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Nadine Epp

Full Text Available Endosomal biogenesis depends on multiple fusion and fission events. For fusion, the heterohexameric CORVET complex as an effector of the endosomal Rab5/Vps21 GTPase has a central function in the initial tethering event. Here, we show that the CORVET-specific Vps3 and Vps8 subunits, which interact with Rab5/Vps21, require their N-terminal domains for localization and function. Surprisingly, CORVET may lack either one of the two N-terminal domains, but not both, to promote protein sorting via the endosome. The dually truncated complex mislocalizes to the cytosol and is impaired in endocytic protein sorting, but not in assembly. Furthermore, the endosomal localization can be rescued by overexpression of Vps21 or one of the truncated CORVET subunits, even though CORVET assembly is not impaired by loss of the N-terminal domains or in strains lacking all endosomal Rab5s and Ypt7. We thus conclude that CORVET requires only its C-terminal domains for assembly and has beyond its putative β-propeller domains additional binding sites for endosomes, which could be important to bind Vps21 and other endosome-specific factors for efficient endosome tethering.

Review on Driverless Traffic from Management Perspective

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Chen Tingting

2017-01-01

Full Text Available The move towards automated driving is gaining ground. This paper reviews the development process of self-driving technology and discusses the safety and efficiency advantages of autonomous vehicles. The discussion shows that the existing traffic management system, including transport infrastructures and regulations, should be changed accordingly to maximize the advantages of autonomous driving. Thus, this paper subsequently gives an insight of the traffic management from three aspects: fully self-driving traffic infrastructures, mixed traffic infrastructures and regulations. First, it is summarized in detail what should be adjusted in intersections, parking lots, pedestrian crossings, ramps, signs and markings. With the transformation of traffic infrastructures, the advantages of driverless car will be more pronounced on account of increased capacity, reduced delay and land use. Also, this paper indicates that the implementations of strict product liability for self-driving car manufacturers and no-fault tort liability for users are applicable to automated vehicle accidents.

Lysosomal and endosomal heterogeneity in the liver: A comparison of the intracellular pathways of endocytosis in rat liver cells

International Nuclear Information System (INIS)

Kindberg, G.M.; Tolleshaug, H.; Gjoen, T.; Berg, T.

1991-01-01

Air-filled albumin microspheres, asialoorosomucoid and formaldehyde-treated serum albumin are selectively taken up by endocytosis in rat liver Kupffer cells, parenchymal cells and endothelial cells, respectively. Intracellular transport and degradation of endocytosed material were studied by subcellular fractionation in sucrose and Nycodenz gradients after intravenous injection of the ligand. By using ligands labeled with 125I-tyramine-cellobiose, the subcellular distribution of labeled degradation products can be studied because they are trapped at the site of formation. The results show that the kinetics of intracellular transport are different in hepatic parenchymal, endothelial and Kupffer cells. In endothelial cells, the ligand is associated with two types of endosomes during the first minutes after internalization and then is transferred rapidly to the lysosomes. In parenchymal cells, 125I-tyramine-cellobiose-asialoorosomucoid was located in a relatively slowly sedimenting vesicle during the first minute after internalization and subsequently in denser endosomes. Degradation of 125I-tyramine-cellobiose-asialoorosomucoid in parenchymal cells started later than that of 125I-tyramine-cellobiose-formaldehyde-treated serum albumin in endothelial cells. Furthermore, the ligand seemed to be transferred relatively slowly from endosomes to lysosomes, and most of the undegraded ligand was in the endosomes. The rate-limiting step of proteolysis in parenchymal cells is probably the transport from endosomes to lysosomes. In Kupffer cells, most 125I-tyramine-cellobiose-microspheres are found as undegraded material in very dense endosomes up to 3 hr after injection. After 20 hr, most of the ligand is degraded in lysosomes distributed at a lower density than the endosomes in Nycodenz and sucrose gradients

Active transportation: do current traffic safety policies protect non-motorists?

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Mader, Emily M; Zick, Cathleen D

2014-06-01

This study investigated the impact that state traffic safety regulations have on non-motorist fatality rates. Data obtained from the National Highway Traffic Safety Administration (NHTSA), the Federal Highway Administration (FHWA), and the National Institute on Alcohol Abuse and Alcoholism (NIAAA) were analyzed through a pooled time series cross-sectional model using fixed effects regression for all 50 states from 1999 to 2009. Two dependent variables were used in separate models measuring annual state non-motorist fatalities per million population, and the natural log of state non-motorist fatalities. Independent variables measuring traffic policies included state expenditures for highway law enforcement and safety per capita; driver cell phone use regulations; graduated driver license regulations; driver blood alcohol concentration regulations; bike helmet regulations; and seat belt regulations. Other control variables included percent of all vehicle miles driven that are urban and mean per capita alcohol consumption per year. Non-motorist traffic safety was positively impacted by state highway law enforcement and safety expenditures per capita, with a decrease in non-motorist fatalities occurring with increased spending. Per capita consumption of alcohol also influenced non-motorist fatalities, with higher non-motorist fatalities occurring with higher per capita consumption of alcohol. Other traffic safety covariates did not appear to have a significant impact on non-motorist fatality rates in the models. Our research suggests that increased expenditures on state highway and traffic safety and the initiation/expansion of programs targeted at curbing both driver and non-motorist intoxication are a starting point for the implementation of traffic safety policies that reduce risks for non-motorists. Copyright © 2014 Elsevier Ltd. All rights reserved.

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

Science.gov (United States)

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

2011-01-01

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

Concerted regulation of npc2 binding to endosomal/lysosomal membranes by bis(monoacylglycero)phosphate and sphingomyelin

DEFF Research Database (Denmark)

Enkavi, Giray; Mikkolainen, Heikki; Güngör, Burçin

2017-01-01

remained elusive. Here, based on an extensive set of atomistic simulations and free energy calculations, we clarify the mechanism and energetics of npc2-membrane binding and characterize the roles of physiologically relevant key lipids associated with the binding process. Our results capture in atomistic......Niemann-Pick Protein C2 (npc2) is a small soluble protein critical for cholesterol transport within and from the lysosome and the late endosome. Intriguingly, npc2-mediated cholesterol transport has been shown to be modulated by lipids, yet the molecular mechanism of npc2-membrane interactions has......). This mode is associated with cholesterol uptake and release. On the other hand, the second mode (Supine) places the cholesterol binding pocket away from the membrane surface, but has overall higher membrane binding affinity. We determined that bis(monoacylglycero)phosphate (bmp) is specifically required...

The Use of Adaptive Traffic Signal Systems Based on Floating Car Data

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Vittorio Astarita

2017-01-01

Full Text Available This paper presents a simple concept which has not been, up to now, thoroughly explored in scientific research: the use of information coming from the network of Internet connected mobile devices (on vehicles to regulate traffic light systems. Three large-scale changes are going to shape the future of transportation and could lead to the regulation of traffic signal system based on floating car data (FCD: (i the implementation of Internet connected cars with global navigation satellite (GNSS system receivers and the autonomous car revolution; (ii the spreading of mobile cooperative Web 2.0 and the extension to connected vehicles; (iii an increasing need for sustainability of transportation in terms of energy efficiency, traffic safety, and environmental issues. Up to now, the concept of floating car data (FCD has only been extensively used to obtain traffic information and estimate traffic parameters. Traffic lights regulation based on FCD technology has not been fully researched since the implementation requires new ideas and algorithms. This paper intends to provide a seminal insight into the important issue of adaptive traffic light based on FCD by presenting ideas that can be useful to researchers and engineers in the long-term task of developing new algorithms and systems that may revolutionize the way traffic lights are regulated.

75 FR 54692 - Notice of Debarment Pursuant to Section 127.7(c) of the International Traffic in Arms Regulations

Science.gov (United States)

2010-09-08

... International Traffic in Arms Regulations Title: Bureau of Political-Military Affairs; Statutory Debarment under... Compliance, Bureau of Political-Military Affairs, Department of State (202) 663-2980. SUPPLEMENTARY.... Statutory debarment is based solely upon conviction in a criminal proceeding, conducted by a United States...

Anterograde trafficking of KCa3.1 in polarized epithelia is Rab1- and Rab8-dependent and recycling endosome-independent.

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Claudia A Bertuccio

Full Text Available The intermediate conductance, Ca2+-activated K+ channel (KCa3.1 targets to the basolateral (BL membrane in polarized epithelia where it plays a key role in transepithelial ion transport. However, there are no studies defining the anterograde and retrograde trafficking of KCa3.1 in polarized epithelia. Herein, we utilize Biotin Ligase Acceptor Peptide (BLAP-tagged KCa3.1 to address these trafficking steps in polarized epithelia, using MDCK, Caco-2 and FRT cells. We demonstrate that KCa3.1 is exclusively targeted to the BL membrane in these cells when grown on filter supports. Following endocytosis, KCa3.1 degradation is prevented by inhibition of lysosomal/proteosomal pathways. Further, the ubiquitylation of KCa3.1 is increased following endocytosis from the BL membrane and PR-619, a deubiquitylase inhibitor, prevents degradation, indicating KCa3.1 is targeted for degradation by ubiquitylation. We demonstrate that KCa3.1 is targeted to the BL membrane in polarized LLC-PK1 cells which lack the μ1B subunit of the AP-1 complex, indicating BL targeting of KCa3.1 is independent of μ1B. As Rabs 1, 2, 6 and 8 play roles in ER/Golgi exit and trafficking of proteins to the BL membrane, we evaluated the role of these Rabs in the trafficking of KCa3.1. In the presence of dominant negative Rab1 or Rab8, KCa3.1 cell surface expression was significantly reduced, whereas Rabs 2 and 6 had no effect. We also co-immunoprecipitated KCa3.1 with both Rab1 and Rab8. These results suggest these Rabs are necessary for the anterograde trafficking of KCa3.1. Finally, we determined whether KCa3.1 traffics directly to the BL membrane or through recycling endosomes in MDCK cells. For these studies, we used either recycling endosome ablation or dominant negative RME-1 constructs and determined that KCa3.1 is trafficked directly to the BL membrane rather than via recycling endosomes. These results are the first to describe the anterograde and retrograde trafficking of KCa3

Legal aspects of motor traffic trauma in Sri Lanka

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Clifford Perera

2016-12-01

Legislation plays a critical role in regulating motor traffic in a country. The Motor Traffic Act of Sri Lanka has undergone many revisions in the recent past to accommodate new provisions to regularize road traffic effectively. However implementation of such provisions is heavily dependent on the rapid and effective action of the police on road traffic offences and awareness and attitudes of courts towards penalizing offenders in view of streamlining the legislation on road traffic.

ON THE ARRIVAL TRAFFIC FLOW ORGANIZATION

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Nikita A. Assorov

2017-01-01

Full Text Available This article is about air traffic flow organization, ICAO regulations describe the organizing of traffic flow as one of the purposes of air traffic control, but they don’t state exactly at what point the flow has to be organized and metered. The flight phase, where air traffic controller interferes with his instructions in order to begin organizing of all aircraft landing at a certain airport depends on the actual traffic volume per hour, airspace capacity and design.The example of air traffic situation in Moscow Domodedovo airport is described in the article, with runway 32 right in use, no significant weather, real usage of STARs, considering all the ICAO and Russian Federation regulations regarding speed control with the restrictions mentioned in AIP of Moscow Domodedovo. The purpose of the experiment is to prove the need of metering the air traffic flow on the entry points in Moscow TMA, because in case of unorganized air traffic flow approach controllers will have additional unnecessary workload.The conducted calculations show, that only 3 aircraft entering TMA on the same distance from initial approach point can be handled using only speed control and existing standard arrival procedures, in all other cases vectoring or holding areas should be used.In order to avoid such situations and increase the number of the aircraft that can be handled by the approach controller with less instructions, all the traffic arriving on the TMA entry point has to be metered by area control centre, because the air traffic control unit has much more space and time for long term speed control modifications, e.g. ±0,02 Mach. In conclusion a simple rule comes to mind – the bigger inbound traffic is, the earlier one has to organize it, in order to do it speed control, radar vectors, miles-in-trail can be used. Also new equipment and technology can help air traffic controller with this task, e.g. AMAN (arrival manager, in addition to this, the experience of

Early to Late Endosome Trafficking Controls Secretion and Zymogen Activation in Rodent and Human Pancreatic Acinar Cells.

Science.gov (United States)

Messenger, Scott W; Thomas, Diana Dh; Cooley, Michelle M; Jones, Elaina K; Falkowski, Michelle A; August, Benjamin K; Fernandez, Luis A; Gorelick, Fred S; Groblewski, Guy E

2015-11-01

Pancreatic acinar cells have an expanded apical endosomal system, the physiological and pathophysiological significance of which is still emerging. Phosphatidylinositol-3,5-bisphosphate (PI(3,5)P 2 ) is an essential phospholipid generated by PIKfyve, which phosphorylates phosphatidylinositol-3-phosphate (PI(3)P). PI(3,5)P 2 is necessary for maturation of early endosomes (EE) to late endosomes (LE). Inhibition of EE to LE trafficking enhances anterograde endosomal trafficking and secretion at the plasma membrane by default through a recycling endosome (RE) intermediate. We assessed the effects of modulating PIKfyve activity on apical trafficking and pancreatitis responses in pancreatic acinar cells. Inhibition of EE to LE trafficking was achieved using pharmacological inhibitors of PIKfyve, expression of dominant negative PIKfyve K1877E, or constitutively active Rab5-GTP Q79L. Anterograde endosomal trafficking was manipulated by expression of constitutively active and dominant negative Rab11a mutants. The effects of these agents on secretion, endolysosomal exocytosis of lysosome associated membrane protein (LAMP1), and trypsinogen activation in response to high-dose CCK-8, bile acids and cigarette toxin was determined. PIKfyve inhibition increased basal and stimulated secretion. Adenoviral overexpression of PIKfyve decreased secretion leading to cellular death. Expression of Rab5-GTP Q79L or Rab11a-GTP Q70L enhanced secretion. Conversely, dominant-negative Rab11a-GDP S25N reduced secretion. High-dose CCK inhibited endolysosomal exocytosis that was reversed by PIKfyve inhibition. PIKfyve inhibition blocked intracellular trypsin accumulation and cellular damage responses to high CCK-8, tobacco toxin, and bile salts in both rodent and human acini. These data demonstrate that EE-LE trafficking acutely controls acinar secretion and the intracellular activation of zymogens leading to the pathogenicity of acute pancreatitis.

Traffic networks as information systems a viability approach

CERN Document Server

Aubin, Jean-Pierre

2017-01-01

This authored monograph covers a viability to approach to traffic management by advising to vehicles circulated on the network the velocity they should follow for satisfying global traffic conditions;. It presents an investigation of three structural innovations: The objective is to broadcast at each instant and at each position the advised celerity to vehicles, which could be read by auxiliary speedometers or used by cruise control devices. Namely, 1. Construct regulation feedback providing at each time and position advised velocities (celerities) for minimizing congestion or other requirements. 2. Taking into account traffic constraints of different type, the first one being to remain on the roads, to stop at junctions, etc. 3. Use information provided by the probe vehicles equipped with GPS to the traffic regulator; 4. Use other global traffic measures of vehicles provided by different types of sensors; These results are based on convex analysis, intertemporal optimization and viability theory as mathemati...

Traffic analysis and control using image processing

Science.gov (United States)

Senthilkumar, K.; Ellappan, Vijayan; Arun, A. R.

2017-11-01

This paper shows the work on traffic analysis and control till date. It shows an approach to regulate traffic the use of image processing and MATLAB systems. This concept uses computational images that are to be compared with original images of the street taken in order to determine the traffic level percentage and set the timing for the traffic signal accordingly which are used to reduce the traffic stoppage on traffic lights. They concept proposes to solve real life scenarios in the streets, thus enriching the traffic lights by adding image receivers like HD cameras and image processors. The input is then imported into MATLAB to be used. as a method for calculating the traffic on roads. Their results would be computed in order to adjust the traffic light timings on a particular street, and also with respect to other similar proposals but with the added value of solving a real, big instance.

Direct binding of retromer to human papillomavirus type 16 minor capsid protein L2 mediates endosome exit during viral infection.

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Andreea Popa

2015-02-01

Full Text Available Trafficking of human papillomaviruses to the Golgi apparatus during virus entry requires retromer, an endosomal coat protein complex that mediates the vesicular transport of cellular transmembrane proteins from the endosome to the Golgi apparatus or the plasma membrane. Here we show that the HPV16 L2 minor capsid protein is a retromer cargo, even though L2 is not a transmembrane protein. We show that direct binding of retromer to a conserved sequence in the carboxy-terminus of L2 is required for exit of L2 from the early endosome and delivery to the trans-Golgi network during virus entry. This binding site is different from known retromer binding motifs and can be replaced by a sorting signal from a cellular retromer cargo. Thus, HPV16 is an unconventional particulate retromer cargo, and retromer binding initiates retrograde transport of viral components from the endosome to the trans-Golgi network during virus entry. We propose that the carboxy-terminal segment of L2 protein protrudes through the endosomal membrane and is accessed by retromer in the cytoplasm.

An ER-Associated Pathway Defines Endosomal Architecture for Controlled Cargo Transport

NARCIS (Netherlands)

Jongsma, Marlieke L. M.; Berlin, Ilana; Wijdeven, Ruud H. M.; Janssen, Lennert; Janssen, George M. C.; Garstka, Malgorzata A.; Janssen, Hans; Mensink, Mark; van Veelen, Peter A.; Spaapen, Robbert M.; Neefjes, Jacques

2016-01-01

Through a network of progressively maturing vesicles, the endosomal system connects the cell's interior with extracellular space. Intriguingly, this network exhibits a bilateral architecture, comprised of a relatively immobile perinuclear vesicle "cloud" and a highly dynamic peripheral contingent.

Identification of Risk Factors Influencing Road Traffic Accidents

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M. Touahmia

2018-02-01

Full Text Available Road traffic accidents (RTAs are becoming a major problem around the world, incurring enormous losses of human and economic resources. Recent reports from the World Health Organization (WHO reveal that each year more than 1.25 million people are killed and 50 million are injured in road traffic accidents worldwide. In Saudi Arabia, statistics show that at least one traffic accident occurs every minute, causing up to 7,000 deaths and over 39,000 injuries annually. In this study, the main causes of RATs in the province of Hail are examined. The data was collected through the use of a survey which was developed to evaluate the effect of influencing parameters on RTA rate. The results show that 67% of RTAs result from human factors, 29% from road conditions and 4% from vehicle defects. Excessive speed and violation of traffic rules and regulations were found to be the main causes of RATs. Low rates of compliance with speed limit signs and seat-belt regulations were also observed. These findings highlight the need of strengthening effective traffic law enforcement alongside with improving traffic safety and raising public awareness.

5 CFR 842.207 - Air traffic controllers.

Science.gov (United States)

2010-01-01

... 5 Administrative Personnel 2 2010-01-01 2010-01-01 false Air traffic controllers. 842.207 Section 842.207 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT (CONTINUED) CIVIL SERVICE REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-BASIC ANNUITY Eligibility § 842.207 Air traffic controllers. (a) An employee who separates from...

Unmanned Aircraft Systems Traffic Management (UTM): Conflict Mitigation Approach

Science.gov (United States)

Johnson, Marcus

2018-01-01

This is a presentation that describes the UAS Traffic Management Approach to an alternate means of compliance with 91.113 right of way regulations. UTM is an "air traffic management" ecosystem for uncontrolled operations.

Synaptic activity regulates AMPA receptor trafficking through different recycling pathways

Science.gov (United States)

Zheng, Ning; Jeyifous, Okunola; Munro, Charlotte; Montgomery, Johanna M; Green, William N

2015-01-01

Changes in glutamatergic synaptic strength in brain are dependent on AMPA-type glutamate receptor (AMPAR) recycling, which is assumed to occur through a single local pathway. In this study, we present evidence that AMPAR recycling occurs through different pathways regulated by synaptic activity. Without synaptic stimulation, most AMPARs recycled in dynamin-independent endosomes containing the GTPase, Arf6. Few AMPARs recycled in dynamin-dependent endosomes labeled by transferrin receptors (TfRs). AMPAR recycling was blocked by alterations in the GTPase, TC10, which co-localized with Arf6 endosomes. TC10 mutants that reduced AMPAR recycling had no effect on increased AMPAR levels with long-term potentiation (LTP) and little effect on decreased AMPAR levels with long-term depression. However, internalized AMPAR levels in TfR-containing recycling endosomes increased after LTP, indicating increased AMPAR recycling through the dynamin-dependent pathway with synaptic plasticity. LTP-induced AMPAR endocytosis is inconsistent with local recycling as a source of increased surface receptors, suggesting AMPARs are trafficked from other sites. DOI: http://dx.doi.org/10.7554/eLife.06878.001 PMID:25970033

Augmenting the Efficacy of Immunotoxins and Other Targeted Protein Toxins by Endosomal Escape Enhancers

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Hendrik Fuchs

2016-07-01

Full Text Available The toxic moiety of almost all protein-based targeted toxins must enter the cytosol of the target cell to mediate its fatal effect. Although more than 500 targeted toxins have been investigated in the past decades, no antibody-targeted protein toxin has been approved for tumor therapeutic applications by the authorities to date. Missing efficacy can be attributed in many cases to insufficient endosomal escape and therefore subsequent lysosomal degradation of the endocytosed toxins. To overcome this drawback, many strategies have been described to weaken the membrane integrity of endosomes. This comprises the use of lysosomotropic amines, carboxylic ionophores, calcium channel antagonists, various cell-penetrating peptides of viral, bacterial, plant, animal, human and synthetic origin, other organic molecules and light-induced techniques. Although the efficacy of the targeted toxins was typically augmented in cell culture hundred or thousand fold, in exceptional cases more than million fold, the combination of several substances harbors new problems including additional side effects, loss of target specificity, difficulties to determine the therapeutic window and cell type-dependent variations. This review critically scrutinizes the chances and challenges of endosomal escape enhancers and their potential role in future developments.

Proteolytic processing of epidermal growth factor within endosomes

International Nuclear Information System (INIS)

Gorman, R.M.; Savage, C.R. Jr.; Poretz, R.D.; Schaudies, R.P.

1986-01-01

The authors have reported previously that EGF enters 3 biochemically distinct non-lysosomal intracellular compartments prior to detection within lysosomes. Earlier studies have demonstrated that EGF is processes by sequential removal of 1, 4 and 1 aminoacyl residues at the C-terminus. The final form, which lacks the 6 residues, accumulates in secondary lysosomes. After subcellular fractionation of fibroblasts exposed to 125 I-EGF, ligand is detected with 3 non-lysosomal endocytic compartments and is fully processed prior to entrance into secondary lysosome. Following internalization, EGF enters an early endosomal compartment (E 1 ). The composition of the ligand (60%, -1 form; 40%, native form) represents an enhancement of the -1 form relative to that on the plasma membrane following the 90 min, 0 0 binding period. The proportion of different EGF forms in E 1 remains constant through the 2 min pulse and chase periods up to 30 min. However, in the ultimate endosomal compartment, E 4 , the proportion of the -6 form increases from 25% at 15 min to greater than 75% in 30 min, with a concomitant decrease of the -1 and -5 forms. Secondary lysosomes contain an EGF composition similar to that found in E 4 at 30 min. Accordingly, it appears that EGF is processed to the -6 form following passage through E 1 and during its tenure in E 4

The Processed Amino-Terminal Fragment of Human TLR7 Acts as a Chaperone To Direct Human TLR7 into Endosomes

Science.gov (United States)

Shepherd, Dawn; Booth, Sarah; Waithe, Dominic; Reis e Sousa, Caetano

2015-01-01

TLR7 mediates innate immune responses to viral RNA in endocytic compartments. Mouse and human (h)TLR7 undergo proteolytic cleavage, resulting in the generation of a C-terminal fragment that accumulates in endosomes and associates with the signaling adaptor MyD88 upon receptor triggering by TLR7 agonists. Although mouse TLR7 is cleaved in endosomes by acidic proteases, hTLR7 processing can occur at neutral pH throughout the secretory pathway through the activity of furin-like proprotein convertases. However, the mechanisms by which cleaved hTLR7 reaches the endosomal compartment remain unclear. In this study, we demonstrate that, after hTLR7 proteolytic processing, the liberated amino (N)-terminal fragment remains bound to the C terminus through disulfide bonds and provides key trafficking information that ensures correct delivery of the complex to endosomal compartments. In the absence of the N-terminal fragment, the C-terminal fragment is redirected to the cell surface, where it is functionally inactive. Our data reveal a novel role for the N terminus of hTLR7 as a molecular chaperone that provides processed hTLR7 with the correct targeting instructions to reach the endosomal compartment, hence ensuring its biological activity and preventing inadvertent cell surface responses to self-RNA. PMID:25917086

A voltage-gated calcium channel regulates lysosomal fusion with endosomes and autophagosomes and is required for neuronal homeostasis.

Directory of Open Access Journals (Sweden)

Xuejun Tian

2015-03-01

Full Text Available Autophagy helps deliver sequestered intracellular cargo to lysosomes for proteolytic degradation and thereby maintains cellular homeostasis by preventing accumulation of toxic substances in cells. In a forward mosaic screen in Drosophila designed to identify genes required for neuronal function and maintenance, we identified multiple cacophony (cac mutant alleles. They exhibit an age-dependent accumulation of autophagic vacuoles (AVs in photoreceptor terminals and eventually a degeneration of the terminals and surrounding glia. cac encodes an α1 subunit of a Drosophila voltage-gated calcium channel (VGCC that is required for synaptic vesicle fusion with the plasma membrane and neurotransmitter release. Here, we show that cac mutant photoreceptor terminals accumulate AV-lysosomal fusion intermediates, suggesting that Cac is necessary for the fusion of AVs with lysosomes, a poorly defined process. Loss of another subunit of the VGCC, α2δ or straightjacket (stj, causes phenotypes very similar to those caused by the loss of cac, indicating that the VGCC is required for AV-lysosomal fusion. The role of VGCC in AV-lysosomal fusion is evolutionarily conserved, as the loss of the mouse homologues, Cacna1a and Cacna2d2, also leads to autophagic defects in mice. Moreover, we find that CACNA1A is localized to the lysosomes and that loss of lysosomal Cacna1a in cerebellar cultured neurons leads to a failure of lysosomes to fuse with endosomes and autophagosomes. Finally, we show that the lysosomal CACNA1A but not the plasma-membrane resident CACNA1A is required for lysosomal fusion. In summary, we present a model in which the VGCC plays a role in autophagy by regulating the fusion of AVs with lysosomes through its calcium channel activity and hence functions in maintaining neuronal homeostasis.

Decoupling Internalization, Acidification and Phagosmal-Endosomal/Iysosomal Phagocytosis of Internalin A coated Beads in epithelial cells

Energy Technology Data Exchange (ETDEWEB)

Blanchette, C D; Woo, Y; Thomas, C; Shen, N; Sulchek, T A; Hiddessen, A L

2008-12-22

Phagocytosis has been extensively examined in 'professional' phagocytic cells using pH sensitive dyes. However, in many of the previous studies, a separation between the end of internalization, beginning of acidification and completion of phagosomal-endosomal/lysosomal fusion was not clearly established, and in several cases, it was treated as a one-step process. In addition, very little work has been done to systematically examine phagosomal maturation in 'non-professional' phagocytic cells, such as epithelial cells. Therefore, in this study, we developed a simple and novel method to decouple and accurately measure particle internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in Madin-Darby Canine Kidney (MDCK) and Caco-2 epithelial cells. Our method was developed using a pathogen mimetic system consisting of polystyrene beads coated with Internalin A (InlA), a membrane surface protein from Listeria monocytogenes known to trigger receptor-mediated internalization. We achieved independent measurements of the rates of internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in epithelial cells by combining the InlA-coated beads (InlA-beads) with antibody quenching, pH sensitive dyes and endosomal/lysosomal dyes, as follows: the rate of InlA bead internalization was measured via antibody quenching of a pH independent dye (Alexa488) conjugated to InlA-beads, the rate at which phagosomes containing internalized InlA beads became acidified was measured using a pH dependent dye (FITC) conjugated to the beads and the rate of phagosomal-endosomal/lysosomal fusion was measured using a combination of unlabeled InlA-beads and an endosomal/lysosomal dye. By performing these independent measurements under identical experimental conditions, we were able to decouple the three processes and establish time scales for each. In a separate set of experiments, we also exploited the phagosomal acidification

Stargazin regulates AMPA receptor trafficking through adaptor protein complexes during long-term depression

Science.gov (United States)

Matsuda, Shinji; Kakegawa, Wataru; Budisantoso, Timotheus; Nomura, Toshihiro; Kohda, Kazuhisa; Yuzaki, Michisuke

2013-11-01

Long-term depression (LTD) underlies learning and memory in various brain regions. Although postsynaptic AMPA receptor trafficking mediates LTD, its underlying molecular mechanisms remain largely unclear. Here we show that stargazin, a transmembrane AMPA receptor regulatory protein, forms a ternary complex with adaptor proteins AP-2 and AP-3A in hippocampal neurons, depending on its phosphorylation state. Inhibiting the stargazin-AP-2 interaction disrupts NMDA-induced AMPA receptor endocytosis, and inhibiting that of stargazin-AP-3A abrogates the late endosomal/lysosomal trafficking of AMPA receptors, thereby upregulating receptor recycling to the cell surface. Similarly, stargazin’s interaction with AP-2 or AP-3A is necessary for low-frequency stimulus-evoked LTD in CA1 hippocampal neurons. Thus, stargazin has a crucial role in NMDA-dependent LTD by regulating two trafficking pathways of AMPA receptors—transport from the cell surface to early endosomes and from early endosomes to late endosomes/lysosomes—through its sequential binding to AP-2 and AP-3A.

Recycling endosomes in apical plasma membrane domain formation and epithelial cell polarity

NARCIS (Netherlands)

Golachowska, Magdalena R.; Hoekstra, Dick; van IJzendoorn, Sven C. D.

2010-01-01

Recycling endosomes have taken central stage in the intracellular sorting and polarized trafficking of apical and basolateral plasma membrane components. Molecular players in the underlying mechanisms are now emerging, including small GTPases, class V myosins and adaptor proteins. In particular,

Solo/Trio8, a membrane-associated short isoform of Trio, modulates endosome dynamics and neurite elongation.

Science.gov (United States)

Sun, Ying-Jie; Nishikawa, Kaori; Yuda, Hideki; Wang, Yu-Lai; Osaka, Hitoshi; Fukazawa, Nobuna; Naito, Akira; Kudo, Yoshihisa; Wada, Keiji; Aoki, Shunsuke

2006-09-01

With DNA microarrays, we identified a gene, termed Solo, that is downregulated in the cerebellum of Purkinje cell degeneration mutant mice. Solo is a mouse homologue of rat Trio8-one of multiple Trio isoforms recently identified in rat brain. Solo/Trio8 contains N-terminal sec14-like and spectrin-like repeat domains followed by a single guanine nucleotide exchange factor 1 (GEF1) domain, but it lacks the C-terminal GEF2, immunoglobulin-like, and kinase domains that are typical of Trio. Solo/Trio8 is predominantly expressed in Purkinje neurons of the mouse brain, and expression begins following birth and increases during Purkinje neuron maturation. We identified a novel C-terminal membrane-anchoring domain in Solo/Trio8 that is required for enhanced green fluorescent protein-Solo/Trio8 localization to early endosomes (positive for both early-endosome antigen 1 [EEA1] and Rab5) in COS-7 cells and primary cultured neurons. Solo/Trio8 overexpression in COS-7 cells augmented the EEA1-positive early-endosome pool, and this effect was abolished via mutation and inactivation of the GEF domain or deletion of the C-terminal membrane-anchoring domain. Moreover, primary cultured neurons transfected with Solo/Trio8 showed increased neurite elongation that was dependent on these domains. These results suggest that Solo/Trio8 acts as an early-endosome-specific upstream activator of Rho family GTPases for neurite elongation of developing Purkinje neurons.

Early to Late Endosome Trafficking Controls Secretion and Zymogen Activation in Rodent and Human Pancreatic Acinar CellsSummary

Directory of Open Access Journals (Sweden)

Scott W. Messenger

2015-11-01

Full Text Available Background & Aims: Pancreatic acinar cells have an expanded apical endosomal system, the physiologic and pathophysiologic significance of which is still emerging. Phosphatidylinositol-3,5-bisphosphate [PI(3,5P2] is an essential phospholipid generated by phosphatidylinositol 3-phosphate 5-kinase (PIKfyve, which phosphorylates phosphatidylinositol-3-phosphate (PI3P. PI(3,5P2 is necessary for maturation of early endosomes (EE to late endosomes (LE. Inhibition of EE to LE trafficking enhances anterograde endosomal trafficking and secretion at the plasma membrane by default through a recycling endosome (RE intermediate. We assessed the effects of modulating PIKfyve activity on apical trafficking and pancreatitis responses in pancreatic acinar cells. Methods: Inhibition of EE to LE trafficking was achieved using pharmacologic inhibitors of PIKfyve, expression of dominant negative PIKfyve K1877E, or constitutively active Rab5-GTP Q79L. Anterograde endosomal trafficking was manipulated by expression of constitutively active and dominant negative Rab11a mutants. The effects of these agents on secretion, endolysosomal exocytosis of lysosome associated membrane protein (LAMP1, and trypsinogen activation in response to supramaximal cholecystokinin (CCK-8, bile acids, and cigarette toxin was determined. Results: PIKfyve inhibition increased basal and stimulated secretion. Adenoviral overexpression of PIKfyve decreased secretion leading to cellular death. Expression of Rab5-GTP Q79L or Rab11a-GTP Q70L enhanced secretion. Conversely, dominant-negative Rab11a-GDP S25N reduced secretion. High-dose CCK inhibited endolysosomal exocytosis that was reversed by PIKfyve inhibition. PIKfyve inhibition blocked intracellular trypsin accumulation and cellular damage responses to supramaximal CCK-8, tobacco toxin, and bile salts in both rodent and human acini. Conclusions: These data demonstrate that EE-LE trafficking acutely controls acinar secretion and the intracellular

75 FR 39820 - Procedures for Abatement of Highway Traffic Noise and Construction Noise

Science.gov (United States)

2010-07-13

...-2008-0114] RIN 2125-AF26 Procedures for Abatement of Highway Traffic Noise and Construction Noise... the Federal regulations on the Procedures for Abatement of Highway Traffic Noise and Construction Noise. The final rule clarifies and adds definitions, the applicability of this regulation, certain...

Recycling endosomes in human cytotoxic T lymphocytes constitute an auxiliary intracellular trafficking pathway for newly synthesized perforin

Science.gov (United States)

Lesteberg, Kelsey E.; Orange, Jordan S.; Makedonas, George

2018-01-01

Background Although cytotoxic T lymphocytes (CTLs) store perforin within cytoplasmic secretory granules for immediate use, perforin is synthesized anew within hours of TCR stimulation. Previously, we observed new perforin protein at an immunologic synapse independent of secretory lysosomes; herein we aimed to determine how new perforin transits to the synapse if not via lytic granules. Results We analyzed antigen-specific human CTLs via imaging flow cytometry and high-resolution confocal microscopy, with attention to intracellular trafficking components and new perforin. The recycling endosome compartments identified by rab8, rab11a, rab4, and rab37 co-localized with new perforin, as well as the SNAREs vti1b and VAMP4. After ablating the function of the recycling endosome pathway, we observed a relative accumulation of new perforin in rab8 vesicles. Conclusions The recycling endosome pathway may serve as an auxiliary intracellular route for the delivery of new perforin to an immunologic synapse in order to perpetuate a cytotoxic response. PMID:28822075

ABMA, a small molecule that inhibits intracellular toxins and pathogens by interfering with late endosomal compartments.

Science.gov (United States)

Wu, Yu; Pons, Valérie; Goudet, Amélie; Panigai, Laetitia; Fischer, Annette; Herweg, Jo-Ana; Kali, Sabrina; Davey, Robert A; Laporte, Jérôme; Bouclier, Céline; Yousfi, Rahima; Aubenque, Céline; Merer, Goulven; Gobbo, Emilie; Lopez, Roman; Gillet, Cynthia; Cojean, Sandrine; Popoff, Michel R; Clayette, Pascal; Le Grand, Roger; Boulogne, Claire; Tordo, Noël; Lemichez, Emmanuel; Loiseau, Philippe M; Rudel, Thomas; Sauvaire, Didier; Cintrat, Jean-Christophe; Gillet, Daniel; Barbier, Julien

2017-11-14

Intracellular pathogenic microorganisms and toxins exploit host cell mechanisms to enter, exert their deleterious effects as well as hijack host nutrition for their development. A potential approach to treat multiple pathogen infections and that should not induce drug resistance is the use of small molecules that target host components. We identified the compound 1-adamantyl (5-bromo-2-methoxybenzyl) amine (ABMA) from a cell-based high throughput screening for its capacity to protect human cells and mice against ricin toxin without toxicity. This compound efficiently protects cells against various toxins and pathogens including viruses, intracellular bacteria and parasite. ABMA provokes Rab7-positive late endosomal compartment accumulation in mammalian cells without affecting other organelles (early endosomes, lysosomes, the Golgi apparatus, the endoplasmic reticulum or the nucleus). As the mechanism of action of ABMA is restricted to host-endosomal compartments, it reduces cell infection by pathogens that depend on this pathway to invade cells. ABMA may represent a novel class of broad-spectrum compounds with therapeutic potential against diverse severe infectious diseases.

Pedestrian Safety in Road Traffic in Poland

Science.gov (United States)

Budzynski, Marcin; Jamroz, Kazimierz; Mackun, Tomasz

2017-10-01

Every third road accident in Poland involves a pedestrian as a participant or, most of the time, a casualty. Pedestrian accidents are usually the result of complex situations and the outcome of a number of factors related to driver and pedestrian behaviour and road infrastructure. Safety depends largely on how well the traffic condition is perceived and on visibility in traffic. The paper presents the results of analyses of methodologies for systematic studies of pedestrian behaviour and pedestrian-driver relations. The effects of the location of the site, type of cross-section and other selected parameters on pedestrian and driver behaviour are demonstrated. The analyses showed that pedestrians are most often put at risk by too long pedestrian crossings, vehicles going too fast around pedestrian crossings, lack of proper sight distance and poorly lit or unlit pedestrian crossings. The reason for such defective infrastructure is that planners, designers, contractors and maintenance services are not receiving any support from design, marking and maintenance regulations for pedestrian traffic. In addition, the Road Traffic Law is not restrictive enough when it comes to drivers’ obligations towards pedestrian safety. Polish design regulations allow long pedestrian crossings up to four lanes in one direction or three lanes in two directions irrespective of traffic control and speed limits. Pedestrian crossings should be kept at a maximum of three lanes. There is nothing in the design regulations about the required driver-pedestrian sight distance. Neither does the Road Traffic Law help engineers with that. It is legal to park vehicles within 10 m of a pedestrian crossing which does not guarantee the necessary sight distance. Drivers must be able to see a pedestrian waiting or stepping onto the crossing from a distance that will help them come to a stop safely. It is safer to follow the principle of providing adequate pedestrian sight distance. Recommendations for

CpG traffic lights are markers of regulatory regions in humans

KAUST Repository

Khamis, Abdullah M.; Lioznova, Anna V.; Artemov, Artem V.; Ramensky, Vasily; Bajic, Vladimir B.; Medvedeva, Yulia A.

2016-01-01

DNA methylation is involved in regulation of gene expression. Although modern methods profile DNA methylation at single CpG sites, methylation levels are usually averaged over genomic regions in the downstream analyses. In this study we demonstrate that single CpG methylation can serve as a more accurate predictor of gene expression compared to average promoter / gene body methylation. CpG positions with significant correlation between methylation and expression of a gene nearby (named CpG traffic lights) are evolutionary conserved and enriched for exact TSS positions and active enhancers. Among all promoter types, CpG traffic lights are especially enriched in poised promoters. Genes that harbor CpG traffic lights are associated with development and signal transduction. Methylation levels of individual CpG traffic lights vary between cell types dramatically with the increased frequency of intermediate methylation levels, indicating cell population heterogeneity in CpG methylation levels. Being in line with the concept of the inherited stochastic epigenetic variation, methylation of such CpG positions might contribute to transcriptional regulation. Alternatively, one can hypothesize that traffic lights are markers of absent gene expression resulting from inactivation of their regulatory elements. The CpG traffic lights provide a promising insight into mechanisms of enhancer activity and gene regulation linking methylation of single CpG to expression.

CpG traffic lights are markers of regulatory regions in humans

KAUST Repository

Khamis, Abdullah M.

2016-12-29

DNA methylation is involved in regulation of gene expression. Although modern methods profile DNA methylation at single CpG sites, methylation levels are usually averaged over genomic regions in the downstream analyses. In this study we demonstrate that single CpG methylation can serve as a more accurate predictor of gene expression compared to average promoter / gene body methylation. CpG positions with significant correlation between methylation and expression of a gene nearby (named CpG traffic lights) are evolutionary conserved and enriched for exact TSS positions and active enhancers. Among all promoter types, CpG traffic lights are especially enriched in poised promoters. Genes that harbor CpG traffic lights are associated with development and signal transduction. Methylation levels of individual CpG traffic lights vary between cell types dramatically with the increased frequency of intermediate methylation levels, indicating cell population heterogeneity in CpG methylation levels. Being in line with the concept of the inherited stochastic epigenetic variation, methylation of such CpG positions might contribute to transcriptional regulation. Alternatively, one can hypothesize that traffic lights are markers of absent gene expression resulting from inactivation of their regulatory elements. The CpG traffic lights provide a promising insight into mechanisms of enhancer activity and gene regulation linking methylation of single CpG to expression.

Amyloid precursor protein and endosomal-lysosomal dysfunction in Alzheimer's disease: inseparable partners in a multifactorial disease.

Science.gov (United States)

Nixon, Ralph A

2017-07-01

Abnormalities of the endosomal-lysosomal network (ELN) are a signature feature of Alzheimer's disease (AD). These include the earliest known cytopathology that is specific to AD and that affects endosomes and induces the progressive failure of lysosomes, each of which are directly linked by distinct mechanisms to neurodegeneration. The origins of ELN dysfunction and β-amyloidogenesis closely overlap, which reflects their common genetic basis, the established early involvement of endosomes and lysosomes in amyloid precursor protein (APP) processing and clearance, and the pathologic effect of certain APP metabolites on ELN functions. Genes that promote β-amyloidogenesis in AD (APP, PSEN1/2, and APOE4) have primary effects on ELN function. The importance of primary ELN dysfunction to pathogenesis is underscored by the mutations in more than 35 ELN-related genes that, thus far, are known to cause familial neurodegenerative diseases even though different pathogenic proteins may be involved. In this article, I discuss growing evidence that implicates AD gene-driven ELN disruptions as not only the antecedent pathobiology that underlies β-amyloidogenesis but also as the essential partner with APP and its metabolites that drive the development of AD, including tauopathy, synaptic dysfunction, and neurodegeneration. The striking amelioration of diverse deficits in animal AD models by remediating ELN dysfunction further supports a need to integrate APP and ELN relationships, including the role of amyloid-β, into a broader conceptual framework of how AD arises, progresses, and may be effectively therapeutically targeted.-Nixon, R. A. Amyloid precursor protein and endosomal-lysosomal dysfunction in Alzheimer's disease: inseparable partners in a multifactorial disease. © FASEB.

Altered neurological function in mice immunized with early endosome antigen 1

Directory of Open Access Journals (Sweden)

Fritzler Marvin J

2004-01-01

Full Text Available Abstract Background Autoantibodies directed against the 160 kDa endosome protein early endosome antigen 1 (EEA1 are seen in patients with neurological diseases. To determine if antibodies to EEA1 have a neuropathological effect, mice from three major histocompatability haplotype backgrounds (H2q, H2b and H2d were immunized with EEA1 (amino acids 82–1411 that was previously shown to contain the target EEA1 epitopes. The mice were then subjected to five neuro-behavioural tests: grid walking, forelimb strength, open field, reaching and rotarod. Results The immunized SWR/J mice with sustained anti-EEA1 antibodies had significantly reduced forelimb strength than the control non-immune mice of the same strain, and BALB/CJ immune mice demonstrated significantly more forelimb errors on the grid walk test than the control group. Conclusions Antibodies to recombinant EEA1 in mice may mediate neurological deficits that are consistent with clinical features of some humans that spontaneously develop anti-EEA1 autoantibodies.

Spatio-temporal regulation of Hsp90-ligand complex leads to immune activation.

Directory of Open Access Journals (Sweden)

Yasuaki eTamura

2016-05-01

Full Text Available Hsp90 is the most abundant cytosolic HSP and is known to act as a molecular chaperone. We found that an Hsp90-cancer antigen peptide complex was efficiently cross-presented by human monocyte-derived dendritic cells and induced peptide-specific cytotoxic T lymphocytes. Furthermore, we observed that the internalized Hsp90-peptide complex was strictly sorted to the Rab5+, EEA1+ static early endosome and the Hsp90-chaperoned peptide was processed and bound to MHC class I molecules through a endosome-recycling pathway. We also found that extracellular Hsp90 complexed with CpG-A or self-DNA stimulates production of a large amount of IFN-α from pDCs via static early endosome targeting. Thus, extracellular Hsp90 can target the antigen or nucleic acid to a static early endosome by spatio-temporal regulation. Moreover, we showed that Hsp90 associates with and delivers TLR7/9 from the ER to early endosomes for ligand recognition. Hsp90 inhibitor, geldanamycin derivative inhibited the Hsp90 association with TLR7/9, resulting in inhibition IFN-α production, leading to improvement of SLE symptoms. Interstingly, we observed that serum Hsp90 is clearly increased in patients with active SLE compared with that in patients with inactive disease. Serum Hsp90 detected in SLE patients binds to self-DNA and/or anti-DNA Ab, thus leading to stimulation of pDCs to produce IFN-α. Thus, Hsp90 plays a crucial role in the pathogenesis of SLE and that an Hsp90 inhibitor will therefore provide a new therapeutic approach to SLE and other nucleic acid-related autoimmune diseases. We will discuss how spatio-temporal regulation of Hsp90-ligand complexes within antigen-presenting cells affects the innate immunity and adaptive immunity.

Android-based E-Traffic law enforcement system in Surakarta City

Science.gov (United States)

Yulianto, Budi; Setiono

2018-03-01

The urban advancement is always overpowered by the increasing number of vehicles as the need for movement of people and goods. This can lead to traffic problems if there is no effort on the implementation of traffic management and engineering, and traffic law enforcement. In this case, the Government of Surakarta City has implemented various policies and regulations related to traffic management and engineering in order to run traffic in an orderly, safe and comfortable manner according to the applicable law. However, conditions in the field shows that traffic violations still occurred frequently due to the weakness of traffic law enforcement in terms of human resources and the system. In this connection, a tool is needed to support traffic law enforcement, especially in relation to the reporting system of traffic violations. This study aims to develop an Android-based traffic violations reporting application (E-Traffic Law Enforcement) as part of the traffic law enforcement system in Surakarta City. The Android-apps records the location and time of the traffic violations incident along with the visual evidence of the infringement. This information will be connected to the database system to detect offenders and to do the traffic law enforcement process.

Drosophila VAMP7 regulates Wingless intracellular trafficking.

Science.gov (United States)

Gao, Han; He, Fang; Lin, Xinhua; Wu, Yihui

2017-01-01

Drosophila Wingless (Wg) is a morphogen that determines cell fate during development. Previous studies have shown that endocytic pathways regulate Wg trafficking and signaling. Here, we showed that loss of vamp7, a gene required for vesicle fusion, dramatically increased Wg levels and decreased Wg signaling. Interestingly, we found that levels of Dally-like (Dlp), a glypican that can interact with Wg to suppress Wg signaling at the dorsoventral boundary of the Drosophila wing, were also increased in vamp7 mutant cells. Moreover, Wg puncta in Rab4-dependent recycling endosomes were Dlp positive. We hypothesize that VAMP7 is required for Wg intracellular trafficking and the accumulation of Wg in Rab4-dependent recycling endosomes might affect Wg signaling.

Molecular determinants of the interaction between Doa1 and Hse1 involved in endosomal sorting.

Science.gov (United States)

Han, Seungsu; Shin, Donghyuk; Choi, Hoon; Lee, Sangho

2014-03-28

Yeast Doa1/Ufd3 is an adaptor protein for Cdc48 (p97 in mammal), an AAA type ATPase associated with endoplasmic reticulum-associated protein degradation pathway and endosomal sorting into multivesicular bodies. Doa1 functions in the endosomal sorting by its association with Hse1, a component of endosomal sorting complex required for transport (ESCRT) system. The association of Doa1 with Hse1 was previously reported to be mediated between PFU domain of Doa1 and SH3 of Hse1. However, it remains unclear which residues are specifically involved in the interaction. Here we report that Doa1/PFU interacts with Hse1/SH3 with a moderate affinity of 5 μM. Asn-438 of Doa1/PFU and Trp-254 of Hse1/SH3 are found to be critical in the interaction while Phe-434, implicated in ubiquitin binding via a hydrophobic interaction, is not. Small-angle X-ray scattering measurements combined with molecular docking and biochemical analysis yield the solution structure of the Doa1/PFU:Hse1/SH3 complex. Taken together, our results suggest that hydrogen bonding is a major determinant in the interaction of Doa1/PFU with Hse1/SH3. Copyright © 2014 Elsevier Inc. All rights reserved.

Molecular basis for SNX-BAR-mediated assembly of distinct endosomal sorting tubules

DEFF Research Database (Denmark)

van Weering, Jan R.T.; Sessions, Richard B.; Traer, Colin J.

2012-01-01

that dimerization is achieved in part through neutralization of charged residues in the hydrophobic BAR-dimerization interface. Membrane remodelling also requires functional amphipathic helices, predicted to be present in all SNX-BARs, and the formation of high order SNX-BAR oligomers through selective 'tip...... and organizes the tubular endosomal network....

Augmented internalisation of ferroportin to late endosomes impairs iron uptake by enterocyte-like IEC-6 cells.

Science.gov (United States)

Oates, Phillip S; Thomas, Carla

2005-08-01

Absorption of iron occurs by duodenal enterocytes, involving uptake by the divalent metal transporter-1 (DMT1) and release by ferroportin. Ferroportin responds to the hepatocyte-produced 25-amino-acid-peptide hepcidin-25 by undergoing internalisation to late endosomes that impair iron release. Ferroportin is also expressed on the apical membrane of polarised Caco-2 cells, rat intestinal cells and in IEC-6 cells (an intestinal epithelial cell line). A blocking antibody to ferroportin also impairs the uptake, but not the release, of iron. In this study IEC-6 cells were used to study the mechanism of impairment or recovery from impairment produced by the blocking antibody and the fate of DMT1 and ferroportin. Uptake of 1 muM Fe(II) was studied by adding the antibody from time 0 and after adding or removing the antibody once a steady state had been reached. Surface binding, maximum iron transport rate V(max) and transporter affinity (K(m)) were measured after impairment of iron uptake. Ferroportin and DMT1 distribution were assessed by immunofluorescence microscopy. Antibody-mediated impairment, or recovery from impairment, of Fe(II) uptake occurred within minutes. Impairment was lost when the antibody was combined with the immunizing peptide. DMT1 and ferroportin undergo internalisation to late endosomes and, in the presence of the antibody, augmented internalisation of DMT1 and ferroportin caused swelling of late endosomes. Surface binding of Fe(II) and iron transport V(max) were reduced by 50%, indicating that the antibody removed membrane-bound DMT1. The ferroportin antibody induced rapid turnover of membrane ferroportin and DMT1 and its internalisation to late endosomes, resulting in impaired Fe(II) uptake.

Acid-sensing ion channel (ASIC) 4 predominantly localizes to an early endosome-related organelle upon heterologous expression.

Science.gov (United States)

Schwartz, Verena; Friedrich, Katharina; Polleichtner, Georg; Gründer, Stefan

2015-12-15

Acid-sensing ion channels (ASICs) are voltage-independent proton-gated amiloride sensitive sodium channels, belonging to the DEG/ENaC gene family. Six different ASICs have been identified (ASIC1a, ASIC1b, ASIC2a, ASIC2b, ASIC3, ASIC4) that are activated by a drop in extracellular pH, either as homo- or heteromers. An exception is ASIC4, which is not activated by protons as a homomer and which does not contribute to functional heteromeric ASICs. Insensitivity of ASIC4 to protons and its comparatively low sequence identity to other ASICs (45%) raises the question whether ASIC4 may have different functions than other ASICs. In this study, we therefore investigated the subcellular localization of ASIC4 in heterologous cell lines, which revealed a surprising accumulation of the channel in early endosome-related vacuoles. Moreover, we identified an unique amino-terminal motif as important for forward-trafficking from the ER/Golgi to the early endosome-related compartment. Collectively, our results show that heterologously expressed ASIC4 predominantly resides in an intracellular endosomal compartment.

Prevalence of Low Back Pain Among Traffic Wardens in Lagos State ...

African Journals Online (AJOL)

to determine the prevalence of low back pain and its associated risk factors among traffic wardens. ... in regulating traffic also has a significant influence on the severity of LBP. ... wardens in Lagos State, who assume a standing position for.

Structure and Function of Vps15 in the Endosomal G Protein Signaling Pathway

Energy Technology Data Exchange (ETDEWEB)

Heenan, Erin J.; Vanhooke, Janeen L.; Temple, Brenda R.; Betts, Laurie; Sondek, John E.; Dohlman, Henrik G.; (UNC)

2009-09-11

G protein-coupled receptors mediate cellular responses to a wide variety of stimuli, including taste, light, and neurotransmitters. In the yeast Saccharomyces cerevisiae, activation of the pheromone pathway triggers events leading to mating. The view had long been held that the G protein-mediated signal occurs principally at the plasma membrane. Recently, it has been shown that the G protein {alpha} subunit Gpa1 can promote signaling at endosomes and requires two components of the sole phosphatidylinositol-3-kinase in yeast, Vps15 and Vps34. Vps15 contains multiple WD repeats and also binds to Gpa1 preferentially in the GDP-bound state; these observations led us to hypothesize that Vps15 may function as a G protein {beta} subunit at the endosome. Here we show an X-ray crystal structure of the Vps15 WD domain that reveals a seven-bladed propeller resembling that of typical G{beta} subunits. We show further that the WD domain is sufficient to bind Gpa1 as well as to Atg14, a potential G{gamma} protein that exists in a complex with Vps15. The Vps15 kinase domain together with the intermediate domain (linking the kinase and WD domains) also contributes to Gpa1 binding and is necessary for Vps15 to sustain G protein signaling. These findings reveal that the Vps15 G{beta}-like domain serves as a scaffold to assemble Gpa1 and Atg14, whereas the kinase and intermediate domains are required for proper signaling at the endosome.

Vehicle Routing with Traffic Congestion and Drivers' Driving and Working Rules

NARCIS (Netherlands)

Kok, A.L.; Hans, Elias W.; Schutten, Johannes M.J.; Zijm, Willem H.M.

2010-01-01

For the intensively studied vehicle routing problem (VRP), two real-life restrictions have received only minor attention in the VRP-literature: traffic congestion and driving hours regulations. Traffic congestion causes late arrivals at customers and long travel times resulting in large transport

Leishmania donovani resides in modified early endosomes by upregulating Rab5a expression via the downregulation of miR-494

Science.gov (United States)

Verma, Jitender Kumar; Rastogi, Ruchir

2017-01-01

Several intracellular pathogens arrest the phagosome maturation in the host cells to avoid transport to lysosomes. In contrast, the Leishmania containing parasitophorous vacuole (PV) is shown to recruit lysosomal markers and thus Leishmania is postulated to be residing in the phagolysosomes in macrophages. Here, we report that Leishmania donovani specifically upregulates the expression of Rab5a by degrading c-Jun via their metalloprotease gp63 to downregulate the expression of miR-494 in THP-1 differentiated human macrophages. Our results also show that miR-494 negatively regulates the expression of Rab5a in cells. Subsequently, L. donovani recruits and retains Rab5a and EEA1 on PV to reside in early endosomes and inhibits transport to lysosomes in human macrophages. Similarly, we have also observed that Leishmania PV also recruits Rab5a by upregulating its expression in human PBMC differentiated macrophages. However, the parasite modulates the endosome by recruiting Lamp1 and inactive pro-CathepsinD on PV via the overexpression of Rab5a in infected cells. Furthermore, siRNA knockdown of Rab5a or overexpression of miR-494 in human macrophages significantly inhibits the survival of the parasites. These results provide the first mechanistic insights of parasite-mediated remodeling of endo-lysosomal trafficking to reside in a specialized early endocytic compartment. PMID:28650977

Ebola Viral Glycoprotein Bound to Its Endosomal Receptor Niemann-Pick C1.

Science.gov (United States)

Wang, Han; Shi, Yi; Song, Jian; Qi, Jianxun; Lu, Guangwen; Yan, Jinghua; Gao, George F

2016-01-14

Filoviruses, including Ebola and Marburg, cause fatal hemorrhagic fever in humans and primates. Understanding how these viruses enter host cells could help to develop effective therapeutics. An endosomal protein, Niemann-Pick C1 (NPC1), has been identified as a necessary entry receptor for this process, and priming of the viral glycoprotein (GP) to a fusion-competent state is a prerequisite for NPC1 binding. Here, we have determined the crystal structure of the primed GP (GPcl) of Ebola virus bound to domain C of NPC1 (NPC1-C) at a resolution of 2.3 Å. NPC1-C utilizes two protruding loops to engage a hydrophobic cavity on head of GPcl. Upon enzymatic cleavage and NPC1-C binding, conformational change in the GPcl further affects the state of the internal fusion loop, triggering membrane fusion. Our data therefore provide structural insights into filovirus entry in the late endosome and the molecular basis for design of therapeutic inhibitors of viral entry. Copyright © 2016 Elsevier Inc. All rights reserved.

Decoupling internalization, acidification and phagosomal-endosomal/lysosomal fusion during phagocytosis of InlA coated beads in epithelial cells.

Directory of Open Access Journals (Sweden)

Craig D Blanchette

Full Text Available BACKGROUND: Phagocytosis has been extensively examined in 'professional' phagocytic cells using pH sensitive dyes. However, in many of the previous studies, a separation between the end of internalization, beginning of acidification and completion of phagosomal-endosomal/lysosomal fusion was not clearly established. In addition, very little work has been done to systematically examine phagosomal maturation in 'non-professional' phagocytic cells. Therefore, in this study, we developed a simple method to measure and decouple particle internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in Madin-Darby Canine Kidney (MDCK and Caco-2 epithelial cells. METHODOLOGY/PRINCIPAL FINDINGS: Our method was developed using a pathogen mimetic system consisting of polystyrene beads coated with Internalin A (InlA, a membrane surface protein from Listeria monocytogenes known to trigger receptor-mediated phagocytosis. We were able to independently measure the rates of internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in epithelial cells by combining the InlA-coated beads (InlA-beads with antibody quenching, a pH sensitive dye and an endosomal/lysosomal dye. By performing these independent measurements under identical experimental conditions, we were able to decouple the three processes and establish time scales for each. In a separate set of experiments, we exploited the phagosomal acidification process to demonstrate an additional, real-time method for tracking bead binding, internalization and phagosomal acidification. CONCLUSIONS/SIGNIFICANCE: Using this method, we found that the time scales for internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion ranged from 23-32 min, 3-4 min and 74-120 min, respectively, for MDCK and Caco-2 epithelial cells. Both the static and real-time methods developed here are expected to be readily and broadly applicable, as they simply

Road Transportation and Traffic Law Enforcement in Nigeria: A case ...

African Journals Online (AJOL)

Road Transportation and Traffic Law Enforcement in Nigeria was established in order to reduce the increasing road crashes and fatalities as well as making road users comply with traffic Laws and regulations as a counter measure, which remain as a great challenge in Nigeria. This paper discussed the roles of the Federal ...

Modeling the signaling endosome hypothesis: Why a drive to the nucleus is better than a (random walk

Directory of Open Access Journals (Sweden)

Howe Charles L

2005-10-01

Full Text Available Abstract Background Information transfer from the plasma membrane to the nucleus is a universal cell biological property. Such information is generally encoded in the form of post-translationally modified protein messengers. Textbook signaling models typically depend upon the diffusion of molecular signals from the site of initiation at the plasma membrane to the site of effector function within the nucleus. However, such models fail to consider several critical constraints placed upon diffusion by the cellular milieu, including the likelihood of signal termination by dephosphorylation. In contrast, signaling associated with retrogradely transported membrane-bounded organelles such as endosomes provides a dephosphorylation-resistant mechanism for the vectorial transmission of molecular signals. We explore the relative efficiencies of signal diffusion versus retrograde transport of signaling endosomes. Results Using large-scale Monte Carlo simulations of diffusing STAT-3 molecules coupled with probabilistic modeling of dephosphorylation kinetics we found that predicted theoretical measures of STAT-3 diffusion likely overestimate the effective range of this signal. Compared to the inherently nucleus-directed movement of retrogradely transported signaling endosomes, diffusion of STAT-3 becomes less efficient at information transfer in spatial domains greater than 200 nanometers from the plasma membrane. Conclusion Our model suggests that cells might utilize two distinct information transmission paradigms: 1 fast local signaling via diffusion over spatial domains on the order of less than 200 nanometers; 2 long-distance signaling via information packets associated with the cytoskeletal transport apparatus. Our model supports previous observations suggesting that the signaling endosome hypothesis is a subset of a more general hypothesis that the most efficient mechanism for intracellular signaling-at-a-distance involves the association of signaling

Road traffic safety in conjunction with in-vehicle ITS

Directory of Open Access Journals (Sweden)

Darja TOPOLŠEK

2014-06-01

Full Text Available Interest in Intelligent Transportation Systems comes from the problems caused by traffic congestion, road accidents and air pollution. Traffic congestion continues to grow worldwide as a result of increased motorization, population growth, changes in population density and urbanization. Interest in ITS can also be attributed to reducing road accidents and increasing traffic safety. The most common causes for road accidents are excessive speed, inattentive driving and ignorance of the right-of-way rules. To eliminate these causes, experience, knowledge of traffic regulations and a new car are not enough – vehicle safety systems have to take part as well. Therefore, the European Union issued a directive on the installation of intelligent systems, whose functions are active support during driving, warning the driver in dangerous situations and alerting passengers of the car in case of irregularities in motor function or actions carried out by the driver that may cause danger, such as swerving while falling asleep. These systems help drivers to avoid accidents, and in the event of a collision, an emergency call is automatically made. Furthermore, they can be used to regulate traffic patterns or to reduce engine performance, which would reduce pollution. With these benefits in mind, the EU has indicated to the automotive industry that installation of these new Intelligent Transportation Systems should be mandatory in their new vehicles.

Motor protein traffic regulation by supply–demand balance of resources

International Nuclear Information System (INIS)

Ciandrini, Luca; Dauloudet, Olivier; Parmeggiani, Andrea; Neri, Izaak; Walter, Jean Charles

2014-01-01

In cells and in in vitro assays the number of motor proteins involved in biological transport processes is far from being unlimited. The cytoskeletal binding sites are in contact with the same finite reservoir of motors (either the cytosol or the flow chamber) and hence compete for recruiting the available motors, potentially depleting the reservoir and affecting cytoskeletal transport. In this work we provide a theoretical framework in which to study, analytically and numerically, how motor density profiles and crowding along cytoskeletal filaments depend on the competition of motors for their binding sites. We propose two models in which finite processive motor proteins actively advance along cytoskeletal filaments and are continuously exchanged with the motor pool. We first look at homogeneous reservoirs and then examine the effects of free motor diffusion in the surrounding medium. We consider as a reference situation recent in vitro experimental setups of kinesin-8 motors binding and moving along microtubule filaments in a flow chamber. We investigate how the crowding of linear motor proteins moving on a filament can be regulated by the balance between supply (concentration of motor proteins in the flow chamber) and demand (total number of polymerized tubulin heterodimers). We present analytical results for the density profiles of bound motors and the reservoir depletion, and propose novel phase diagrams that present the formation of jams of motor proteins on the filament as a function of two tuneable experimental parameters: the motor protein concentration and the concentration of tubulins polymerized into cytoskeletal filaments. Extensive numerical simulations corroborate the analytical results for parameters in the experimental range and also address the effects of diffusion of motor proteins in the reservoir. We then propose experiments for validating our models and discuss how the ‘supply–demand’ effects can regulate motor traffic also in in vivo

Chimeric forms of furin and TGN38 are transported with the plasma membrane in the trans-Golgi network via distinct endosomal pathways.

Science.gov (United States)

Mallet, W G; Maxfield, F R

1999-07-26

Furin and TGN38 are menbrane proteins that cycle between the plasma membrane and the trans-Golgi network (TGN), each maintaining a predominant distribution in the TGN. We have used chimeric proteins with an extracellular Tac domain and the cytoplasmic domain of TGN38 or furin to study the trafficking of these proteins in endosomes. Previously, we demonstrated that the postendocytic trafficking of Tac-TGN38 to the TGN is via the endocytic recycling pathway (Ghosh, R.N.,W.G. Mallet,T.T. Soe,T.E.McGraw, and F.R. Maxfield.1998.J. Cell Biol.142:923-936). Here we show that internalized Tac-furin is delivered to the TGN through late endosomes, bypassing the endocytic recycling compartment. The transport of Tac-furin from late endosomes to the TGN appears to proceed via an efficient, single-pass mechanism. Delivery of Tac-furin but not Tac-TGN38 to the TGN is blocked by nocodazole, and the two pathways are also differentially affected by wortmannin. These studies demonstrate the existence of two independentpathways for endosomal transport of proteins to the TGN from the plasma membrane.

INTELLIGENT TRAFFIC-SAFETY MIRROR BY USING WIRELESS SENSOR NETWORK

Directory of Open Access Journals (Sweden)

Peter Danišovič

2014-03-01

Full Text Available This article is focused on the problematic of traffic safety, dealing with the problem of car intersections with blocked view crossing by a special wireless sensor network (WSN proposed for the traffic monitoring, concretely for vehicle’s detection at places, where it is necessary. Some ultra-low-power TI products were developed due to this reason: microcontroller MSP430F2232, 868MHz RF transceiver CC1101 and LDO voltage regulator TPS7033. The WSN consist of four network nodes supplied with the special safety lightings which serve the function of intelligent traffic safety mirror.

Involvement of Gβγ subunits of Gi protein coupled with S1P receptor on multivesicular endosomes in F-actin formation and cargo sorting into exosomes.

Science.gov (United States)

Kajimoto, Taketoshi; Mohamed, Nesma Nabil Ibrahim; Badawy, Shaymaa Mohamed Mohamed; Matovelo, Shubi Ambwene; Hirase, Mitsuhiro; Nakamura, Shunsuke; Yoshida, Daisuke; Okada, Taro; Ijuin, Takeshi; Nakamura, Shun-Ichi

2018-01-05

Exosomes play a critical role in cell-to-cell communication by delivering cargo molecules to recipient cells. However, the mechanism underlying the generation of the exosomal multivesicular endosome (MVE) is one of the mysteries in the field of endosome research. Although sphingolipid metabolites such as ceramide and sphingosine 1-phosphate (S1P) are known to play important roles in MVE formation and maturation, the detailed molecular mechanisms are still unclear. Here, we show that Rho family GTPases, including Cdc42 and Rac1, are constitutively activated on exosomal MVEs and are regulated by S1P signaling as measured by fluorescence resonance energy transfer (FRET)-based conformational changes. Moreover, we detected S1P signaling-induced filamentous actin (F-actin) formation. A selective inhibitor of Gβγ subunits, M119, strongly inhibited both F-actin formation on MVEs and cargo sorting into exosomal intralumenal vesicles of MVEs, both of which were fully rescued by the simultaneous expression of constitutively active Cdc42 and Rac1. Our results shed light on the mechanism underlying exosomal MVE maturation and inform the understanding of the physiological relevance of continuous activation of the S1P receptor and subsequent downstream G protein signaling to Gβγ subunits/Rho family GTPases-regulated F-actin formation on MVEs for cargo sorting into exosomal intralumenal vesicles. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Preparation of Biodegradable Oligo(lactides-Grafted Dextran Nanogels for Efficient Drug Delivery by Controlling Intracellular Traffic

Directory of Open Access Journals (Sweden)

Yuichi Ohya

2018-05-01

Full Text Available Nanogels, nanometer-sized hydrogel particles, have great potential as drug delivery carriers. To achieve effective drug delivery to the active sites in a cell, control of intracellular traffic is important. In this study, we prepared nanogels composed of dextran with oligolactide (OLA chains attached via disulfide bonds (Dex-g-SS-OLA that collapse under the reductive conditions of the cytosol to achieve efficient drug delivery. In addition, we introduced galactose (Gal residues on the nanogels, to enhance cellular uptake by receptor-mediated endocytosis, and secondary oligo-amine (tetraethylenepentamine groups, to aid in escape from endosomes via proton sponge effects. The obtained Dex-g-SS-OLA with attached Gal residues and tetraethylenepentamine (EI4 groups, EI4/Gal-Dex-g-SS-OLA, formed a nanogel with a hydrodynamic diameter of ca. 203 nm in phosphate-buffered solution. The collapse of the EI4/Gal-Dex-g-SS-OLA nanogels under reductive conditions was confirmed by a decrease in the hydrodynamic diameter in the presence of reductive agents. The specific uptake of the hydrogels into HepG2 cells and their intercellular behavior were investigated by flow cytometry and confocal laser scanning microscopy using fluorescence dye-labeled nanogels. Escape from the endosome and subsequent collapse in the cytosol of the EI4/Gal-Dex-g-SS-OLA were observed. These biodegradable nanogels that collapse under reductive conditions in the cytosol should have great potential as efficient drug carriers in, for example, cancer chemotherapy.

14 CFR Special Federal Aviation... - Air Traffic Control System Emergency Operation

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 2 2010-01-01 2010-01-01 false Air Traffic Control System Emergency Operation Federal Special Federal Aviation Regulation No. 60 Aeronautics and Space FEDERAL AVIATION ADMINISTRATION, DEPARTMENT OF TRANSPORTATION (CONTINUED) AIR TRAFFIC AND GENERAL OPERATING RULES GENERAL OPERATING AND FLIGHT RULES Pt. 91, SFAR No. 60...

A-RAF kinase functions in ARF6 regulated endocytic membrane traffic.

Directory of Open Access Journals (Sweden)

Elena Nekhoroshkova

Full Text Available BACKGROUND: RAF kinases direct ERK MAPK signaling to distinct subcellular compartments in response to growth factor stimulation. METHODOLOGY/PRINCIPAL FINDINGS: Of the three mammalian isoforms A-RAF is special in that one of its two lipid binding domains mediates a unique pattern of membrane localization. Specific membrane binding is retained by an N-terminal fragment (AR149 that corresponds to a naturally occurring splice variant termed DA-RAF2. AR149 colocalizes with ARF6 on tubular endosomes and has a dominant negative effect on endocytic trafficking. Moreover actin polymerization of yeast and mammalian cells is abolished. AR149/DA-RAF2 does not affect the internalization step of endocytosis, but trafficking to the recycling compartment. CONCLUSIONS/SIGNIFICANCE: A-RAF induced ERK activation is required for this step by activating ARF6, as A-RAF depletion or inhibition of the A-RAF controlled MEK-ERK cascade blocks recycling. These data led to a new model for A-RAF function in endocytic trafficking.

Adiponectin release and insulin receptor targeting share trans-Golgi-dependent endosomal trafficking routes

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Maria Rödiger

2018-02-01

Full Text Available Objective: Intracellular vesicle trafficking maintains cellular structures and functions. The assembly of cargo-laden vesicles at the trans-Golgi network is initiated by the ARF family of small GTPases. Here, we demonstrate the role of the trans-Golgi localized monomeric GTPase ARFRP1 in endosomal-mediated vesicle trafficking of mature adipocytes. Methods: Control (Arfrp1flox/flox and inducible fat-specific Arfrp1 knockout (Arfrp1iAT−/− mice were metabolically characterized. In vitro experiments on mature 3T3-L1 cells and primary mouse adipocytes were conducted to validate the impact of ARFRP1 on localization of adiponectin and the insulin receptor. Finally, secretion and transferrin-based uptake and recycling assays were performed with HeLa and HeLa M-C1 cells. Results: We identified the ARFRP1-based sorting machinery to be involved in vesicle trafficking relying on the endosomal compartment for cell surface delivery. Secretion of adiponectin from fat depots was selectively reduced in Arfrp1iAT−/− mice, and Arfrp1-depleted 3T3-L1 adipocytes revealed an accumulation of adiponectin in Rab11-positive endosomes. Plasma adiponectin deficiency of Arfrp1iAT−/− mice resulted in deteriorated hepatic insulin sensitivity, increased gluconeogenesis and elevated fasting blood glucose levels. Additionally, the insulin receptor, undergoing endocytic recycling after ligand binding, was less abundant at the plasma membrane of adipocytes lacking Arfrp1. This had detrimental effects on adipose insulin signaling, followed by insufficient suppression of basal lipolytic activity and impaired adipose tissue expansion. Conclusions: Our findings suggest that adiponectin secretion and insulin receptor surface targeting utilize the same post-Golgi trafficking pathways that are essential for an appropriate systemic insulin sensitivity and glucose homeostasis. Keywords: Adiponectin, ARFRP1, Exocytosis, Insulin receptor, trans-Golgi

Traffic theory

National Research Council Canada - National Science Library

Gazis, Denos C

2002-01-01

... of traffic signal settings The vehicle-actuated traffic signal 87 89 77 CHAPTER 3. TRAFFIC CONTROL 101 Objectives of Traffic Control 103 Single, Isolated Intersection 105 Synchronization Scheme...

Neuron specific Rab4 effector GRASP-1 coordinates membrane specialization and maturation of recycling endosomes

NARCIS (Netherlands)

C.C. Hoogenraad (Casper); I. Popa (Ioana); K. Futai (Kensuke); E. Sanchez-Martinez (Emma); P. Wulf (Phebe); T. van Vlijmen (Thijs); B.R. Dortland (Bjorn); V. Oorschot (Viola); R. Govers (Robert); M. Monti (Maria); A.J.R. Heck (Albert); M. Sheng (Morgan); J. Klumperman (Judith); H. Rehmann (Holger); D. Jaarsma (Dick); L.C. Kapitein (Lukas); P. van der Sluijs

2010-01-01

textabstractThe endosomal pathway in neuronal dendrites is essential for membrane receptor trafficking and proper synaptic function and plasticity. However, the molecular mechanisms that organize specific endocytic trafficking routes are poorly understood. Here, we identify GRIP-associated protein-1

Actin polymerization in the endosomal pathway, but not on the Coxiella-containing vacuole, is essential for pathogen growth.

Directory of Open Access Journals (Sweden)

Heather E Miller

2018-04-01

Full Text Available Coxiella burnetii is an intracellular bacterium that replicates within an expansive phagolysosome-like vacuole. Fusion between the Coxiella-containing vacuole (CCV and late endosomes/multivesicular bodies requires Rab7, the HOPS tethering complex, and SNARE proteins, with actin also speculated to play a role. Here, we investigated the importance of actin in CCV fusion. Filamentous actin patches formed around the CCV membrane that were preferred sites of vesicular fusion. Accordingly, the mediators of endolysosomal fusion Rab7, VAMP7, and syntaxin 8 were concentrated in CCV actin patches. Generation of actin patches required C. burnetii type 4B secretion and host retromer function. Patches decorated with VPS29 and VPS35, components of the retromer, FAM21 and WASH, members of the WASH complex that engage the retromer, and Arp3, a component of the Arp2/3 complex that generates branched actin filaments. Depletion by siRNA of VPS35 or VPS29 reduced CCV actin patches and caused Rab7 to uniformly distribute in the CCV membrane. C. burnetii grew normally in VPS35 or VPS29 depleted cells, as well as WASH-knockout mouse embryo fibroblasts, where CCVs are devoid of actin patches. Endosome recycling to the plasma membrane and trans-Golgi of glucose transporter 1 (GLUT1 and cationic-independent mannose-6-phosphate receptor (CI-M6PR, respectively, was normal in infected cells. However, siRNA knockdown of retromer resulted in aberrant trafficking of GLUT1, but not CI-M6PR, suggesting canonical retrograde trafficking is unaffected by retromer disruption. Treatment with the specific Arp2/3 inhibitor CK-666 strongly inhibited CCV formation, an effect associated with altered endosomal trafficking of transferrin receptor. Collectively, our results show that CCV actin patches generated by retromer, WASH, and Arp2/3 are dispensable for CCV biogenesis and stability. However, Arp2/3-mediated production of actin filaments required for cargo transport within the

Extracellular anti-angiogenic proteins augment an endosomal protein trafficking pathway to reach mitochondria and execute apoptosis in HUVECs.

Science.gov (United States)

Chen, Mo; Qiu, Tao; Wu, Jiajie; Yang, Yang; Wright, Graham D; Wu, Min; Ge, Ruowen

2018-03-09

Classic endocytosis destinations include the recycling endosome returning to the plasma membrane or the late endosome (LE) merging with lysosomes for cargo degradation. However, the anti-angiogenic proteins angiostatin and isthmin, are endocytosed and trafficked to mitochondria (Mito) to execute apoptosis of endothelial cells. How these extracellular proteins reach mitochondria remains a mystery. Through confocal and super-resolution fluorescent microscopy, we demonstrate that angiostatin and isthmin are trafficked to mitochondria through the interaction between LE and Mito. Using purified organelles, the LE-Mito interaction is confirmed through in vitro lipid-fusion assay, as well as single vesicle total internal reflection fluorescent microscopy. LE-Mito interaction enables the transfer of not only lipids but also proteins from LE to Mito. Angiostatin and isthmin augment this endosomal protein trafficking pathway and make use of it to reach mitochondria to execute apoptosis. Cell fractionation and biochemical analysis identified that the cytosolic scaffold protein Na+/H+ exchanger regulatory factor 1 (NHERF1) associated with LE and the t-SNARE protein synaptosome-associated protein 25 kDa (SNAP25) associated with Mito form an interaction complex to facilitate LE-Mito interaction. Proximity ligation assay coupled with fluorescent microscopy showed that both NHERF1 and SNAP25 are located at the contacting face between LE and Mito. RNAi knockdown of either NHERF1 or SNAP25 suppressed not only the mitochondrial trafficking of angiostatin and isthmin but also their anti-angiogenic and pro-apoptotic functions. Hence, this study reveals a previously unrealized endosomal protein trafficking pathway from LE to Mito that allows extracellular proteins to reach mitochondria and execute apoptosis.

Effective photo-enhancement of cellular activity of fluorophore-octaarginine antisense PNA conjugates correlates with singlet oxygen formation, endosomal escape and chromophore lipophilicity

DEFF Research Database (Denmark)

Yarani, Reza; Shiraishi, Takehiko; Nielsen, Peter E.

2018-01-01

Photochemical internalization (PCI) is a cellular drug delivery method based on the generation of light-induced reactive oxygen species (ROS) causing damage to the endosomal membrane and thereby resulting in drug release to the cytoplasm. In our study a series of antisense fluorophore octaarginin...... indicate that efficient photodynamic endosomal escape is strongly dependent on the quantum yield for photochemical singlet oxygen formation, photostability as well as the lipophilicity of the chromophore....

Bicaudal-D1 regulates the intracellular sorting and signalling of neurotrophin receptors.

Science.gov (United States)

Terenzio, Marco; Golding, Matthew; Russell, Matthew R G; Wicher, Krzysztof B; Rosewell, Ian; Spencer-Dene, Bradley; Ish-Horowicz, David; Schiavo, Giampietro

2014-07-17

We have identified a new function for the dynein adaptor Bicaudal D homolog 1 (BICD1) by screening a siRNA library for genes affecting the dynamics of neurotrophin receptor-containing endosomes in motor neurons (MNs). Depleting BICD1 increased the intracellular accumulation of brain-derived neurotrophic factor (BDNF)-activated TrkB and p75 neurotrophin receptor (p75(NTR)) by disrupting the endosomal sorting, reducing lysosomal degradation and increasing the co-localisation of these neurotrophin receptors with retromer-associated sorting nexin 1. The resulting re-routing of active receptors increased their recycling to the plasma membrane and altered the repertoire of signalling-competent TrkB isoforms and p75(NTR) available for ligand binding on the neuronal surface. This resulted in attenuated, but more sustained, AKT activation in response to BDNF stimulation. These data, together with our observation that Bicd1 expression is restricted to the developing nervous system when neurotrophin receptor expression peaks, indicate that BICD1 regulates neurotrophin signalling by modulating the endosomal sorting of internalised ligand-activated receptors. © 2014 The Authors.

Traffic at risk in Mediterranean

International Nuclear Information System (INIS)

Bilardo, U.; Mureddu, G.

1993-01-01

The Mediterranean Sea represents only about 0.7% of the planet's total water surface area, yet it is host to as much as one-quarter of the world's total maritime oil traffic. Statistics indicate that from 47 to 77,000 tonnes of crude oil are now being released annually into the Sea through accidental spills; and over the last decade, its tourism dependent coastlines have been fouled by the highest levels of tar contamination in the world. Oil carrier traffic, routed within the Sea's already overcrowded shipping lanes, is intense and this traffic is expected to increase, as a result of rises in world energy demand, to levels of from 7 to 8 million barrels a day. It has been estimated that, at the end of 1992, 90% of all large tankers operating in this area, will have reached a service life of 15-16 years which is very close to the average recommended life cycle limit of 15-20 years. Only 20% of the world's 3,000 tankers are currently equipped with double bottomed hulls. This paper uses these and other facts and figures to argue that the risks of future severe oil tanker accidents in the Mediterranean Sea are high, and that these must be countered with the development of a new set of stricter marine traffic safety regulations at the Italian, national, as well as, European level

Strategies for the Activation and Release of the Membranolytic Peptide Melittin from Liposomes Using Endosomal pH as a Trigger.

Science.gov (United States)

Oude Blenke, E; Sleszynska, M; Evers, M J W; Storm, G; Martin, N I; Mastrobattista, E

2017-02-15

Endosomolytic peptides are often coupled to drug delivery systems to enhance endosomal escape, which is crucial for the delivery of macromolecular drugs that are vulnerable to degradation in the endolysosomal pathway. Melittin is a 26 amino acid peptide derived from bee venom that has a very high membranolytic activity. However, such lytic peptides also impose a significant safety risk when applied in vivo as they often have similar activity against red blood cells and other nontarget cell membranes. Our aim is to control the membrane-disrupting capacity of these peptides in time and space by physically constraining them to a nanocarrier surface in such a way that they only become activated when delivered inside acidic endosomes. To this end, a variety of chemical approaches for the coupling of lytic peptides to liposomes via functionalized PEG-lipids were explored, including maleimide-thiol chemistry, click-chemistry, and aldehyde-hydrazide chemistry. The latter enables reversible conjugation via a hydrazone bond, allowing for release of the peptide under endosomal conditions. By carefully choosing the conjugation site and by using a pH activated analog of the melittin peptide, lytic activity toward a model membrane is completely inhibited at physiological pH. At endosomal pH the activity is restored by hydrolysis of the acid-labile hydrazone bond, releasing the peptide in its most active, free form. Furthermore, using an analogue containing a nonhydrolyzable bond as a control, it was shown that the activity observed can be completely attributed to release of the peptide, validating dynamic covalent conjugation as a suitable strategy to maintain safety during circulation.

Structural Studies of Adeno-Associated Virus Serotype 8 Capsid Transitions Associated with Endosomal Trafficking

Energy Technology Data Exchange (ETDEWEB)

Nam, Hyun-Joo; Gurda, Brittney L.; McKenna, Robert; Potter, Mark; Byrne, Barry; Salganik, Maxim; Muzyczka, Nicholas; Agbandje-McKenna, Mavis (Florida)

2012-09-17

The single-stranded DNA (ssDNA) parvoviruses enter host cells through receptor-mediated endocytosis, and infection depends on processing in the early to late endosome as well as in the lysosome prior to nuclear entry for replication. However, the mechanisms of capsid endosomal processing, including the effects of low pH, are poorly understood. To gain insight into the structural transitions required for this essential step in infection, the crystal structures of empty and green fluorescent protein (GFP) gene-packaged adeno-associated virus serotype 8 (AAV8) have been determined at pH values of 6.0, 5.5, and 4.0 and then at pH 7.5 after incubation at pH 4.0, mimicking the conditions encountered during endocytic trafficking. While the capsid viral protein (VP) topologies of all the structures were similar, significant amino acid side chain conformational rearrangements were observed on (i) the interior surface of the capsid under the icosahedral 3-fold axis near ordered nucleic acid density that was lost concomitant with the conformational change as pH was reduced and (ii) the exterior capsid surface close to the icosahedral 2-fold depression. The 3-fold change is consistent with DNA release from an ordering interaction on the inside surface of the capsid at low pH values and suggests transitions that likely trigger the capsid for genome uncoating. The surface change results in disruption of VP-VP interface interactions and a decrease in buried surface area between VP monomers. This disruption points to capsid destabilization which may (i) release VP1 amino acids for its phospholipase A2 function for endosomal escape and nuclear localization signals for nuclear targeting and (ii) trigger genome uncoating.

Phosphatidylinositol 3-phosphates-at the interface between cell signalling and membrane traffic.

Science.gov (United States)

Marat, Andrea L; Haucke, Volker

2016-03-15

Phosphoinositides (PIs) form a minor class of phospholipids with crucial functions in cell physiology, ranging from cell signalling and motility to a role as signposts of compartmental membrane identity. Phosphatidylinositol 3-phosphates are present at the plasma membrane and within the endolysosomal system, where they serve as key regulators of both cell signalling and of intracellular membrane traffic. Here, we provide an overview of the metabolic pathways that regulate cellular synthesis of PI 3-phosphates at distinct intracellular sites and discuss the mechanisms by which these lipids regulate cell signalling and membrane traffic. Finally, we provide a framework for how PI 3-phosphate metabolism is integrated into the cellular network. © 2016 The Authors.

Real-Time Corrected Traffic Correlation Model for Traffic Flow Forecasting

Directory of Open Access Journals (Sweden)

Hua-pu Lu

2015-01-01

Full Text Available This paper focuses on the problems of short-term traffic flow forecasting. The main goal is to put forward traffic correlation model and real-time correction algorithm for traffic flow forecasting. Traffic correlation model is established based on the temporal-spatial-historical correlation characteristic of traffic big data. In order to simplify the traffic correlation model, this paper presents correction coefficients optimization algorithm. Considering multistate characteristic of traffic big data, a dynamic part is added to traffic correlation model. Real-time correction algorithm based on Fuzzy Neural Network is presented to overcome the nonlinear mapping problems. A case study based on a real-world road network in Beijing, China, is implemented to test the efficiency and applicability of the proposed modeling methods.

The Cdc42 guanine nucleotide exchange factor FGD6 coordinates cell polarity and endosomal membrane recycling in osteoclasts.

Science.gov (United States)

Steenblock, Charlotte; Heckel, Tobias; Czupalla, Cornelia; Espírito Santo, Ana Isabel; Niehage, Christian; Sztacho, Martin; Hoflack, Bernard

2014-06-27

The initial step of bone digestion is the adhesion of osteoclasts onto bone surfaces and the assembly of podosomal belts that segregate the bone-facing ruffled membrane from other membrane domains. During bone digestion, membrane components of the ruffled border also need to be recycled after macropinocytosis of digested bone materials. How osteoclast polarity and membrane recycling are coordinated remains unknown. Here, we show that the Cdc42-guanine nucleotide exchange factor FGD6 coordinates these events through its Src-dependent interaction with different actin-based protein networks. At the plasma membrane, FGD6 couples cell adhesion and actin dynamics by regulating podosome formation through the assembly of complexes comprising the Cdc42-interactor IQGAP1, the Rho GTPase-activating protein ARHGAP10, and the integrin interactors Talin-1/2 or Filamin A. On endosomes and transcytotic vesicles, FGD6 regulates retromer-dependent membrane recycling through its interaction with the actin nucleation-promoting factor WASH. These results provide a mechanism by which a single Cdc42-exchange factor controlling different actin-based processes coordinates cell adhesion, cell polarity, and membrane recycling during bone degradation. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Impact of traffic noise on railway traffic safety

Directory of Open Access Journals (Sweden)

Zdravko TOŠ

2008-01-01

Full Text Available Traffic noise is one of the dominant factors of ergo-assessment. The harmful impact of traffic noise on the engine driver as target group can be studied in isolation from other ergo-assessment factors only in the initial phase of research. The simultaneous action of several related factors in the system of ergo-assessment factors has cumulative effect on the perception and psychomotoric status of the railway traffic participants in the appropriate traffic situation. The initial partial research of traffic noise by a combination of several scientific methods needs to be eventually upgraded by studying the relations among several concurrent important or dominant ergo-assessment factors.

An antibody toolkit for the study of membrane traffic in Drosophila melanogaster

Directory of Open Access Journals (Sweden)

Falko Riedel

2016-07-01

Full Text Available The use of Drosophila melanogaster as a model organism has been pivotal to understanding the developmental processes of metazoans. However, the use of flies for studying subcellular organization is hampered by a paucity of reliable reagents to label specific organelles. Here, we describe the generation of mouse monoclonal antibodies against a set of markers of the secretory and endocytic pathways, along with goat polyclonal antibodies against two Golgi proteins. We show that the monoclonal antibodies are highly specific and sufficiently sensitive to detect endogenous proteins in crude extracts by immunoblotting with little background staining. By immunofluorescence the major compartments of the membrane traffic system (including the endoplasmic reticulum, the Golgi, and early and late endosomes are labeled by at least one antibody. Moreover, the antibodies can be used to label organelles in fly tissues including salivary glands and wing imaginal discs. We anticipate that these antibodies will provide a useful tool kit to facilitate the investigation of how the endomembrane system functions and varies in the diverse tissue types of metazoans.

Effects of turning and through lane sharing on traffic performance at intersections

Science.gov (United States)

Li, Xiang; Sun, Jian-Qiao

2016-02-01

Turning vehicles strongly influence traffic flows at intersections. Effective regulation of turning vehicles is important to achieve better traffic performance. This paper studies the impact of lane sharing and turning signals on traffic performance at intersections by using cellular automata. Both right-turn and left-turn lane sharing are studied. Interactions between vehicles and pedestrians are considered. The transportation efficiency, road safety and energy economy are the traffic performance metrics. Extensive simulations are carried out to study the traffic performance indices. It is observed that shared turning lanes and permissive left-turn signal improve the transportation efficiency and reduce the fuel consumption in most cases, but the safety is usually sacrificed. It is not always beneficial for the through vehicles when they are allowed to be in the turning lanes.

The late endosomal HOPS complex anchors active G-protein signaling essential for pathogenesis in magnaporthe oryzae.

Directory of Open Access Journals (Sweden)

Ravikrishna Ramanujam

Full Text Available In Magnaporthe oryzae, the causal ascomycete of the devastating rice blast disease, the conidial germ tube tip must sense and respond to a wide array of requisite cues from the host in order to switch from polarized to isotropic growth, ultimately forming the dome-shaped infection cell known as the appressorium. Although the role for G-protein mediated Cyclic AMP signaling in appressorium formation was first identified almost two decades ago, little is known about the spatio-temporal dynamics of the cascade and how the signal is transmitted through the intracellular network during cell growth and morphogenesis. In this study, we demonstrate that the late endosomal compartments, comprising of a PI3P-rich (Phosphatidylinositol 3-phosphate highly dynamic tubulo-vesicular network, scaffold active MagA/GαS, Rgs1 (a GAP for MagA, Adenylate cyclase and Pth11 (a non-canonical GPCR in the likely absence of AKAP-like anchors during early pathogenic development in M. oryzae. Loss of HOPS component Vps39 and consequently the late endosomal function caused a disruption of adenylate cyclase localization, cAMP signaling and appressorium formation. Remarkably, exogenous cAMP rescued the appressorium formation defects associated with VPS39 deletion in M. oryzae. We propose that sequestration of key G-protein signaling components on dynamic late endosomes and/or endolysosomes, provides an effective molecular means to compartmentalize and control the spatio-temporal activation and rapid downregulation (likely via vacuolar degradation of cAMP signaling amidst changing cellular geometry during pathogenic development in M. oryzae.

TRAFFIC SIMULATION FOR MIXED TRAFFIC SYSTEMS

African Journals Online (AJOL)

EGETE

2012-05-04

May 4, 2012 ... Traffic problem is classified into single and mixed, especially in most developing countries, where motorbikes are ..... The traffic light control system presented by its location on ... multi-destination dynamic routing and real-time.

Speed uncertainty and speed regulation in conflict detection and resolution in Air Traffic Control

OpenAIRE

Archambault , Nicolas

2004-01-01

International audience; With the predicted increase of air traffic volume, new air traffic management models are under investigation in order to increase airspace capacity and keep low delays while maintaining transportation safety standards. One of the tasks implied is to solve conflicts, i.e. maintain a sufficient separation between aircraft. Conflict resolution relies on conflict detection ; indeed predicting aircraft trajectories within a time window allows to detect the conflicts and app...

Wind farms' influences for traffic safety. Study of distance requirements for road traffic, railway traffic, sea traffic and air travel; Tuulivoimaloiden vaikutukset liikenneturvallisuuteen. Selvitys etaeisyysvaatimuksista tie-, rautatie-, meri- ja lentoliikenteen osalta

Energy Technology Data Exchange (ETDEWEB)

Hytoenen, K.; Harju, M.; Piispanen, J.; Haulos, S.

2012-11-15

As wind power projects, both design and implementation, have increased rapidly, wind power manufacturers have stated that infrastructure of traffic restricts and complicates implementation of wind power projects. Failure of wind farm and both snow and ice thrown from blades and tower are commonly thought to be dangerous for settlement and for road, railway and sea traffic. Based on the accessible information accidents, which are connected to wind farms, won't cause notable harm for outsiders. Majority of the human injury and fatal accidents concerns the personnel of the wind farm in implementation and operational phases, not outsiders. Purpose of this study has been to clarify how distances requirements in different forms of traffic in Finland differ from other countries that are included in this comparison, and to find grounds for possibly revising these guidelines. In Finland there are still longest distance requirements when placing wind farm nearby the main roads, even after Finnish Traffic Agency revised guidelines in 2012. Regarding roads, differences between all the countries were minor. Also distance requirements regarding railways differ relatively little. Sweden has shortest distance requirement. Distance requirements in Finland and Denmark have been reduced in past few years. Any of these countries have not stated actual defined distance requirements related for example to shipping channels or lanes. International Civil Aviation Organization ICAO has settled internationally rules for obstacle limitation surface nearby airports and marking flight obstacles. Countries have stated national aviation regulations according to these rules and for this reason these regulations are quite consistent. In this study distance requirements are recommended to be calculated for certain basis. In case of roads and railways a general minimum distance requirement is suggested to be 1 x wind farm overall height. Road owner is allowed to require higher distance for

Traffic signal synchronization.

Science.gov (United States)

Huang, Ding-wei; Huang, Wei-neng

2003-05-01

The benefits of traffic signal synchronization are examined within the cellular automata approach. The microsimulations of traffic flow are obtained with different settings of signal period T and time delay delta. Both numerical results and analytical approximations are presented. For undersaturated traffic, the green-light wave solutions can be realized. For saturated traffic, the correlation among the traffic signals has no effect on the throughput. For oversaturated traffic, the benefits of synchronization are manifest only when stochastic noise is suppressed.

Automated mixed traffic vehicle control and scheduling study

Science.gov (United States)

Peng, T. K. C.; Chon, K.

1976-01-01

The operation and the expected performance of a proposed automatic guideway transit system which uses low speed automated mixed traffic vehicles (AMTVs) were analyzed. Vehicle scheduling and headway control policies were evaluated with a transit system simulation model. The effect of mixed traffic interference on the average vehicle speed was examined with a vehicle pedestrian interface model. Control parameters regulating vehicle speed were evaluated for safe stopping and passenger comfort. Some preliminary data on the cost and operation of an experimental AMTV system are included. These data were the result of a separate task conducted at JPL, and were included as background information.

CCC- and WASH-mediated endosomal sorting of LDLR is required for normal clearance of circulating LDL

NARCIS (Netherlands)

Bartuzi, Paulina; Billadeau, Daniel D.; Favier, Robert; Rong, Shunxing; Dekker, Daphne; Fedoseienko, Alina; Fieten, Hille; Wijers, Melinde; Levels, Johannes H.; Huijkman, Nicolette; Kloosterhuis, Niels; Van der Molen, Henk; Brufau, Gemma; Groen, Albert K.; Elliott, Alison M.; Kuivenhoven, Jan Albert; Plecko, Barbara; Grangl, Gernot; McGaughran, Julie; Horton, Jay D.; Burstein, Ezra; Hofker, Marten H.; van de Sluis, Bart

2016-01-01

The low-density lipoprotein receptor (LDLR) plays a pivotal role in clearing atherogenic circulating low-density lipoprotein (LDL) cholesterol. Here we show that the COMMD/CCDC22/CCDC93 (CCC) and the Wiskott-Aldrich syndrome protein and SCAR homologue (WASH) complexes are both crucial for endosomal

CCC- and WASH-mediated endosomal sorting of LDLR is required for normal clearance of circulating LDL

NARCIS (Netherlands)

Bartuzi, Paulina; Billadeau, Daniel D.; Favier, Robert; Rong, Shunxing; Dekker, Daphne; Fedoseienko, Alina; Fieten, Hille; Wijers, Melinde; Levels, Johannes H.; Huijkman, Nicolette; Kloosterhuis, Niels; van der Molen, Henk; Brufau, Gemma; Groen, Albert K.; Elliott, Alison M.; Kuivenhoven, Jan Albert; Plecko, Barbara; Grangl, Gernot; McGaughran, Julie; Horton, Jay D.; Burstein, Ezra; Hofker, Marten H.; van de Sluis, Bart

The low-density lipoprotein receptor (LDLR) plays a pivotal role in clearing atherogenic circulating low-density lipoprotein (LDL) cholesterol. Here we show that the COMMD/CCDC22/CCDC93 (CCC) and the Wiskott-Aldrich syndrome protein and SCAR homologue (WASH) complexes are both crucial for endosomal

Kinesin-3 and dynein cooperate in long-range retrograde endosome motility along a nonuniform microtubule array

NARCIS (Netherlands)

Schuster, M.; Kilaru, S.; Fink, G.; Collemare, J.A.R.; Roger, Y.; Steinberg, G.

2011-01-01

The polarity of microtubules (MTs) determines the motors for intracellular motility, with kinesins moving to plus ends and dynein to minus ends. In elongated cells of Ustilago maydis, dynein is thought to move early endosomes (EEs) toward the septum (retrograde), whereas kinesin-3 transports them to

Imaging endosomes and autophagosomes in whole mammalian cells using correlative cryo-fluorescence and cryo-soft X-ray microscopy (cryo-CLXM)

International Nuclear Information System (INIS)

Duke, Elizabeth M.H.; Razi, Minoo; Weston, Anne; Guttmann, Peter; Werner, Stephan; Henzler, Katja; Schneider, Gerd; Tooze, Sharon A.; Collinson, Lucy M.

2014-01-01

Cryo-soft X-ray tomography (cryo-SXT) is a powerful imaging technique that can extract ultrastructural information from whole, unstained mammalian cells as close to the living state as possible. Subcellular organelles including the nucleus, the Golgi apparatus and mitochondria have been identified by morphology alone, due to the similarity in contrast to transmission electron micrographs. In this study, we used cryo-SXT to image endosomes and autophagosomes, organelles that are particularly susceptible to chemical fixation artefacts during sample preparation for electron microscopy. We used two approaches to identify these compartments. For early and recycling endosomes, which are accessible to externally-loaded markers, we used an anti-transferrin receptor antibody conjugated to 10 nm gold particles. For autophagosomes, which are not accessible to externally-applied markers, we developed a correlative cryo-fluorescence and cryo-SXT workflow (cryo-CLXM) to localise GFP-LC3 and RFP-Atg9. We used a stand-alone cryo-fluorescence stage in the home laboratory to localise the cloned fluorophores, followed by cryo-soft X-ray tomography at the synchrotron to analyse cellular ultrastructure. We mapped the 3D ultrastructure of the endocytic and autophagic structures, and discovered clusters of omegasomes arising from ‘hotspots’ on the ER. Thus, immunogold markers and cryo-CLXM can be used to analyse cellular processes that are inaccessible using other imaging modalities. - Highlights: • We image whole, unstained mammalian cells using cryo-soft X-ray tomography. • Endosomes are identified using a gold marker for the transferrin receptor. • A new workflow for correlative cryo-fluorescence and cryo-SXT is used to locate early autophagosomes. • Interactions between endosomes, endoplasmic reticulum and forming autophagosomes are mapped in 3D. • Multiple omegasomes are shown to form at ‘hotspots’ on the endoplasmic reticulum

The impacts of road traffic management on urban air quality

Energy Technology Data Exchange (ETDEWEB)

Oduyemi, K.O.K. [School of Construction and Environment, University of Abertay Dundee, Dundee (United Kingdom); Davidson, B. [Department of Environmental Health and Consumer Protection, Dundee City Council, Tayside House, Crichton Street, Dundee (United Kingdom)

1998-07-11

The effects of road traffic emissions on urban air quality are investigated, using long-term nitrogen dioxide (NO{sub 2}) data. The effectiveness of the several traffic management measures that have been made in Dundee city centre, UK, within the last 5 years in relation to urban air quality is discussed. The information assessed during this study indicates that the annual mean NO{sub 2} levels at all the study sites are, at present, below the current EC and WHO (long-term) air quality standards for NO{sub 2} concentration in the ambient air. Traffic restrictions appear to be effective in protecting urban air quality. The annual mean NO{sub 2} concentration at two of the study sites is currently close to 40 {mu}g/m{sup 3}, a value published in the Air Quality Regulations 1997 for the air quality objective to be achieved by the year 2005. Proactive traffic management mitigation measures are proposed for these sites and a methodology for the consideration of traffic management alternatives, based upon traffic flow modal split, is described. Some measures proposed are based upon a survey of vehicle occupancy rates, carried out at the busiest of the four study sites. The methodology and assessment procedures presented should be invaluable to assessors of traffic management and local air quality management in a small city, both at the planning and at the auditing stage

Traffic Generator (TrafficGen) Version 1.4.2: Users Guide

Science.gov (United States)

2016-06-01

the network with Transmission Control Protocol and User Datagram Protocol Internet Protocol traffic. Each node generating network traffic in an...TrafficGen Graphical User Interface (GUI) 3 3.1 Anatomy of the User Interface 3 3.2 Scenario Configuration and MGEN Files 4 4. Working with...for public release; distribution is unlimited. vi List of Figures Fig. 1 TrafficGen user interface

33 CFR 162.134 - Connecting waters from Lake Huron to Lake Erie; traffic rules.

Science.gov (United States)

2010-07-01

... 33 Navigation and Navigable Waters 2 2010-07-01 2010-07-01 false Connecting waters from Lake Huron to Lake Erie; traffic rules. 162.134 Section 162.134 Navigation and Navigable Waters COAST GUARD... REGULATIONS § 162.134 Connecting waters from Lake Huron to Lake Erie; traffic rules. (a) Detroit River. The...

Genetic reconstitution of the human Adenovirus type 2 temperature-sensitive 1 mutant defective in endosomal escape

Directory of Open Access Journals (Sweden)

Gastaldelli Michele

2009-10-01

Full Text Available Abstract Human Adenoviruses infect the upper and lower respiratory tracts, the urinary and digestive tracts, lymphoid systems and heart, and give rise to epidemic conjunctivitis. More than 51 human serotypes have been identified to-date, and classified into 6 species A-F. The species C Adenoviruses Ad2 and Ad5 (Ad2/5 cause upper and lower respiratory disease, but how viral structure relates to the selection of particular infectious uptake pathways is not known. An adenovirus mutant, Ad2-ts1 had been isolated upon chemical mutagenesis in the past, and shown to have unprocessed capsid proteins. Ad2-ts1 fails to package the viral protease L3/p23, and Ad2-ts1 virions do not efficiently escape from endosomes. It had been suggested that the C22187T point mutation leading to the substitution of the conserved proline 137 to leucine (P137L in the L3/p23 protease was at least in part responsible for this phenotype. To clarify if the C22187T mutation is necessary and sufficient for the Ad2-ts1 phenotype, we sequenced the genes encoding the structural proteins of Ad2-ts1, and confirmed that the Ad2-ts1 DNA carries the point mutation C22187T. Introduction of C22187T to the wild-type Ad2 genome in a bacterial artificial chromosome (Ad2-BAC gave Ad2-BAC46 virions with the full Ad2-ts1 phenotype. Reversion of Ad2-BAC46 gave wild-type Ad2 particles indicating that P137L is necessary and sufficient for the Ad2-ts1 phenotype. The kinetics of Ad2-ts1 uptake into cells were comparable to Ad2 suggesting similar endocytic uptake mechanisms. Surprisingly, infectious Ad2 or Ad5 but not Ad2-ts1 uptake required CALM (clathrin assembly lymphoid myeloid protein, which controls clathrin-mediated endocytosis and membrane transport between endosomes and the trans-Golgi-network. The data show that no other mutations than P137L in the viral protease are necessary to give rise to particles that are defective in capsid processing and endosomal escape. This provides a basis for

Queueing and traffic

NARCIS (Netherlands)

Baër, Niek

2015-01-01

Traffic jams are everywhere, some are caused by constructions or accidents but a large portion occurs naturally. These "natural" traffic jams are a result of variable driving speeds combined with a high number of vehicles. To prevent these traffic jams, we must understand traffic in general, and to

Accumulation of dsRNA in endosomes contributes to inefficient RNA interference in the fall armyworm, Spodoptera frugiperda.

Science.gov (United States)

Yoon, June-Sun; Gurusamy, Dhandapani; Palli, Subba Reddy

2017-11-01

RNA interference (RNAi) efficiency varies among insects studied. The barriers for successful RNAi include the presence of double-stranded ribonucleases (dsRNase) in the lumen and hemolymph that could potentially digest double-stranded RNA (dsRNA) and the variability in the transport of dsRNA into and within the cells. We recently showed that the dsRNAs are transported into lepidopteran cells, but they are not processed into small interference RNAs (siRNAs) because they are trapped in acidic bodies. In the current study, we focused on the identification of acidic bodies in which dsRNAs accumulate in Sf9 cells. Time-lapse imaging studies showed that dsRNAs enter Sf9 cells and accumulate in acidic bodies within 20 min after their addition to the medium. CypHer-5E-labeled dsRNA also accumulated in the midgut and fat body dissected from Spodoptera frugiperda larvae with similar patterns observed in Sf9 cells. Pharmacological inhibitor assays showed that the dsRNAs use clathrin mediated endocytosis pathway for transport into the cells. We investigated the potential dsRNA accumulation sites employing LysoTracker and double labeling experiments using the constructs to express a fusion of green fluorescence protein with early or late endosomal marker proteins and CypHer-5E-labeled dsRNA. Interestingly, CypHer-5E-labeled dsRNA accumulated predominantly in early and late endosomes. These data suggest that entrapment of internalized dsRNA in endosomes is one of the major factors contributing to inefficient RNAi response in lepidopteran insects. Copyright © 2017 Elsevier Ltd. All rights reserved.

Traffic Games: Modeling Freeway Traffic with Game Theory.

Science.gov (United States)

Cortés-Berrueco, Luis E; Gershenson, Carlos; Stephens, Christopher R

2016-01-01

We apply game theory to a vehicular traffic model to study the effect of driver strategies on traffic flow. The resulting model inherits the realistic dynamics achieved by a two-lane traffic model and aims to incorporate phenomena caused by driver-driver interactions. To achieve this goal, a game-theoretic description of driver interaction was developed. This game-theoretic formalization allows one to model different lane-changing behaviors and to keep track of mobility performance. We simulate the evolution of cooperation, traffic flow, and mobility performance for different modeled behaviors. The analysis of these results indicates a mobility optimization process achieved by drivers' interactions.

Traffic planning for non-homogeneous traffic

Indian Academy of Sciences (India)

Vehicles based on similar traffic operating characteristics are grouped into ... of distances to the video monitor involved using a measuring wheel to mark ... Secondly, the observers reviewed the videotape to sample traffic entity ...... makes a strong case for including service lanes for slow moving vehicles for improving the.

REGULATING THE FLOW OF TRANSPORT WITH ALLOWANCE FOR THE INTERVAL OF MOTION

Directory of Open Access Journals (Sweden)

Аndrew Hul

2017-07-01

Full Text Available Purpose: We show that the regulation of traffic is especially important under conditions of infrastructure. The aim was to show that the objectives of traffic light regulation is to minimize and, where possible, exceptions meetings conflicting streams of vehicles. The authors show that the applied aspects of optimization of traffic lights is to eliminate mash situation and ensure the safety of all road users. Methods: The method used in the comparison is to select the best technical solution, feasibility to test mathematical tools and engineering for the practical implementation of the proposed technical solution. Results: The results of the study is repeated then that shows the organization of the preconditions for the introduction of traffic light regulation based on the calculation cycle of the traffic light object. It is proposed to cycle more than 120 seconds, as if waiting longer allowing drivers can count signal lights faulty and start moving to the blocking signal. The main tasks of traffic light regulation is to minimize and, where possible, exceptions meetings conflicting streams of vehicles and eliminate mash situation and ensure the safety of all participants. An important part of the work on the introduction of traffic light regulation is the calculation cycle of the traffic light object. For reasons of safety, cycle time of more than 120 is considered unacceptable, since the longer waiting signal allowing drivers can count faulty lights and start moving to the prohibitive signal. Discussion: In conclusion, the article revealed that the divergent forms of regulation of the intersection can also be used for general stress on public roads. The methods of testing for the road tion type.

Area-wide traffic calming for preventing traffic related injuries.

Science.gov (United States)

Bunn, F; Collier, T; Frost, C; Ker, K; Roberts, I; Wentz, R

2003-01-01

It is estimated that by 2020 road traffic crashes will have moved from ninth to third in the world disease burden ranking, as measured in disability adjusted life years, and second in developing countries. The identification of effective strategies for the prevention of traffic related injuries is of global health importance. Area-wide traffic calming schemes that discourage through traffic on residential roads is one such strategy. To evaluate the effectiveness of area-wide traffic calming in preventing traffic related crashes, injuries, and deaths. We searched the following electronic databases: Cochrane Injuries Group's Specialised Register, Cochrane Controlled Trials Register, MEDLINE, EMBASE and TRANSPORT (NTIS, TRIS, TRANSDOC). We searched the web sites of road safety organisations, handsearched conference proceedings, checked reference lists of relevant papers and contacted experts in the area. The search was not restricted by language or publication status. Randomised controlled trials, and controlled before-after studies of area-wide traffic calming schemes. Two reviewers independently extracted data on type of study, characteristics of intervention and control areas, and length of data collection periods. Before and after data were collected on the total number of road traffic crashes, all road user deaths and injuries, pedestrian-motor vehicle collisions and road user deaths. The statistical package STATA was used to calculate rate ratios for each study, which were then pooled to give an overall estimate using a random effects model. We found no randomised controlled trials, but 16 controlled before-after trials met our inclusion criteria. Seven studies were done in Germany, six in the UK, two in Australia and one in the Netherlands. There were no studies in low or middle income countries. Eight trials reported the number of road traffic crashes resulting in deaths. The pooled rate ratio was 0.63 (0.14, 2.59 95% CI). Sixteen studies reported the number

The road traffic crashes as a neglected public health concern; an observational study from Iranian population.

Science.gov (United States)

Bakhtiyari, Mahmood; Delpisheh, Ali; Monfared, Ayad Bahadori; Kazemi-Galougahi, Mohammad Hassan; Mehmandar, Mohammad Reza; Riahi, Mohammad; Salehi, Masoud; Mansournia, Mohammad Ali

2015-01-01

Traffic crashes are multifactorial events caused by human factors, technical issues, and environmental conditions. The present study aimed to determine the role of human factors in traffic crashes in Iran using the proportional odds regression model. The database of all traffic crashes in Iran in 2010 (n = 592, 168) registered through the "COM.114" police forms was investigated. Human risk factors leading to traffic crashes were determined and the odds ratio (OR) of each risk factor was estimated using an ordinal regression model and adjusted for potential confounding factors such as age, gender, and lighting status within and outside of cities. The drivers' mean age ± standard deviation was 34.1 ± 14.0 years. The most prevalent risk factors leading to death within cities were disregarding traffic rules and regulations (45%), driver rushing (31%), and alcohol consumption (12.3%). Using the proportional odds regression model, alcohol consumption was the most significant human risk factor in traffic crashes within cities (OR = 6.5, 95% confidence interval [CI], 4.88-8.65) and outside of cities (OR = 1.73, 95% CI, 1.22-3.29). Public health strategies and preventive policies should be focused on more common human risk factors such as disregarding traffic rules and regulations, drivers' rushing, and alcohol consumption due to their greater population attributable fraction and more intuitive impacts on society.

Noxious substances in the air - traffic planning measures - traffic of the future

Energy Technology Data Exchange (ETDEWEB)

Koestenberger, H

1985-01-01

Necessary bundle of measures: Extension of public transport and restriction of individual traffic, extension and activation of large main roads (by-passes) to unload inhabited areas, building garages, creation of residential streets, pedestrian precincts and cycle paths. The best possible traffic system can only be achieved if all means of transport are used efficiently. It is the duty of traffic planners to develop an overall traffic system with the aims of benefiting the whole community. Due to wrong slowing down of traffic, the reduction of emitted quantities of noxious substances from private cars can be counteracted by general slowing down of traffic; frequent braking and restarting. The functional separation of residential areas for living, areas for working, supply, education and leisure pursuits which has been aimed at in recent decades must be slowly changed. This could reduce the traffic and mobility (mixed functions). The aims for traffic of the future are: suitability for the environment, economy, safety and capacity. In an integrated road network, the traffic must take over the correct purpose of traffic. (orig.).

Traffic accidents in Brazil from 1998 to 2010: many changes and few effects.

Science.gov (United States)

Bacchieri, Giancarlo; Barros, Aluísio J D

2011-10-01

The paper describes the situation of road traffic accidents in Brazil since 1998, when a new Brazilian traffic law was approved, up to the year 2010. A review of both academic and non-academic literature was carried out, including journals (both indexed and non-indexed), technical reports, author searches, searches in paper reference lists and direct contact with researchers. The main problems related to road traffic accidents in Brazil identified were the increase in the absolute number of deaths and in the mortality rates, a rapid increase in the number of motorcycles, and drink & driving. Influent authors in the field and centers of expertise were identified. Some potential solutions are presented by the authors, who suggest that the public offices related to traffic regulation and control are not taking suitable measures for control and reduction of road traffic accidents.

Towards Reducing the Dangers Associated with Road Traffic ...

African Journals Online (AJOL)

USER

2018-04-25

Apr 25, 2018 ... ensure drivers' personal safety and to comply with traffic regulations explain high seat belt .... which used to be the public transport fares charged for commuting in Bedford 'mummy trucks' .... females generally are risk-averse and have high-risk perception levels ... places of worship and social gatherings.

49 CFR 236.401 - Automatic block signal system and interlocking standards applicable to traffic control systems.

Science.gov (United States)

2010-10-01

... TRAIN CONTROL SYSTEMS, DEVICES, AND APPLIANCES Traffic Control Systems Standards § 236.401 Automatic... 49 Transportation 4 2010-10-01 2010-10-01 false Automatic block signal system and interlocking standards applicable to traffic control systems. 236.401 Section 236.401 Transportation Other Regulations...

Traffic Infrastructure in the Development of the Croatian Traffic System

Directory of Open Access Journals (Sweden)

Damir Šimulčik

2012-10-01

Full Text Available The absence of a long-term traffic policy and of the policyof financing the constntction and maintenance of traffic infrastructurefacilities, represents a synthesis of numerous unresolvedrelations whose negative effects are felt in the overalleconomic and traffic development and consequently theevaluation of national potentials in the field. Adverse aspectcaused by the lack of a clear and feasible policy of financing thetraffic infrastructure facilities, is also a result of not having definedan adequate traffic policy, programme and strategiccourses of development, nor financing models that would be inaccordance with the market and economy system.This indicates that it is necessary to determine a policy forfinancing the constntction and maintenance of traffic infrastntcture,which has to be based on scientific development,team work, availability of plans and programmes to scientistsand experts, determined methodology based on marketing andeconomic logic in defining the programme and strategic tasksand assignments so as to make them feasible.In the near future, intensive preparations for investments inthe overall traffic sysiem are necessary, especially regarding thetraffic infrastntcture facilities - the pivotal points in the processof evaluating the traffic in our national tenitory. Croatia needsto define clearly its strategy in constructing and maintaining thegeneral traffic infrastructure, appointing at the same time thosewho will carry out the given tasks.

Air Traffic Management Research at NASA

Science.gov (United States)

Farley, Todd

2012-01-01

The U.S. air transportation system is the most productive in the world, moving far more people and goods than any other. It is also the safest system in the world, thanks in part to its venerable air traffic control system. But as demand for air travel continues to grow, the air traffic control systems aging infrastructure and labor-intensive procedures are impinging on its ability to keep pace with demand. And that impinges on the growth of our economy. Part of NASA's current mission in aeronautics research is to invent new technologies and procedures for ATC that will enable our national airspace system to accommodate the increasing demand for air transportation well into the next generation while still maintaining its excellent record for safety. It is a challenging mission, as efforts to modernize have, for decades, been hamstrung by the inability to assure safety to the satisfaction of system operators, system regulators, and/or the traveling public. In this talk, we'll provide a brief history of air traffic control, focusing on the tension between efficiency and safety assurance, and we'll highlight some new NASA technologies coming down the pike.

Emissions of nitrogen oxides from road traffic - regulations, emissions and effects

International Nuclear Information System (INIS)

Sjoedin, Aake; Pihl-Karlsson, Gunilla; Johansson, Manne; Forsberg, Bertil; Erlandsson, Lennart

2004-10-01

The report is a review that aims to improve the basis for additional measures against the road traffic emissions of, in particular, NO x . An important question in the context is whether health effects of NO 2 should serve as a norm for the actions for emission reductions of NO x , or if the environmental effects of NO x -emissions in the form of acidification, eutrophication and ozone should play this role. WHO notes, in its latest review of health effect research, that one cannot demonstrate that NO 2 alone has any direct effects in concentrations at the current whole-year mean norm (40 μg/m 3 ). Such health effects that has been demonstrated in epidemiologic studies at these concentrations are caused by other traffic related emissions (e. g. particles) for which NO 2 constitutes a good indicator. WHO indicates the need for additional sharpening of the norms for ozone and particles. In this context, it is important to note that emissions of NO x on a regional scale contributes to formation of ozone as well as particles. Therefore there exist reasons to decrease the emissions of NO x in order to reach future recommended values for ozone and particles emissions. In the evaluations that will be done during 2004-2005 of the so called Goeteborg protocol, the EU's Ceiling Directives and the CAFE-programme, it is expected that new emissions objective for NO x will be suggested for 2015-2020, to cope with health and environment objectives in Europe. The report shows that that development that currently happens within the vehicle industry, for engines and exhaust emission control system are pursued to meet future exhaust requirement in the USA gives good conditions for the road traffic sector to contribute to that these objectives will reached

77 FR 49859 - Proposed Traffic Records Program Assessment Advisory

Science.gov (United States)

2012-08-17

... decision-making. States need timely, accurate, complete, and uniform traffic records to identify and.... This document provides information on the contents, capabilities, and data quality attributes of an... comments and additional information, see http://www.regulations.gov . Note that all comments received will...

The development of the IAEA transport regulations and its implementation in Europe

International Nuclear Information System (INIS)

Collin, F.W.

1987-01-01

The radiation protection principles are laid down in a binding from in IAEA SS No 9. Regulations of world-wide validity (and, hence, for Europe) are IMO/IMDG Code for sea-going traffic, and ICAO/Technical regulations or IATA/Hazardous goods transport regulations, resp. for air traffic. The regulations by the organizations ADR, RID, ADN, ADNR, and CMEA are valid as transfrontier regulations in Europe. (DG) [de

Localization of the AP-3 adaptor complex defines a novel endosomal exit site for lysosomal membrane proteins

NARCIS (Netherlands)

Peden, A.A.; Oorschot, V.; Hesser, B.A.; Austin, C.D.; Scheller, R.H.; Klumperman, J.

2004-01-01

The adaptor protein (AP) 3 adaptor complex has been implicated in the transport of lysosomal membrane proteins, but its precise site of action has remained controversial. Here, we show by immuno-electron microscopy that AP-3 is associated with budding profiles evolving from a tubular endosomal

Performance of an Automated-Mixed-Traffic-Vehicle /AMTV/ System. [urban people mover

Science.gov (United States)

Peng, T. K. C.; Chon, K.

1978-01-01

This study analyzes the operation and evaluates the expected performance of a proposed automatic guideway transit system which uses low-speed Automated Mixed Traffic Vehicles (AMTV's). Vehicle scheduling and headway control policies are evaluated with a transit system simulation model. The effect of mixed-traffic interference on the average vehicle speed is examined with a vehicle-pedestrian interface model. Control parameters regulating vehicle speed are evaluated for safe stopping and passenger comfort.

The Vici Syndrome Protein EPG5 Is a Rab7 Effector that Determines the Fusion Specificity of Autophagosomes with Late Endosomes/Lysosomes.

Science.gov (United States)

Wang, Zheng; Miao, Guangyan; Xue, Xue; Guo, Xiangyang; Yuan, Chongzhen; Wang, Zhaoyu; Zhang, Gangming; Chen, Yingyu; Feng, Du; Hu, Junjie; Zhang, Hong

2016-09-01

Mutations in the human autophagy gene EPG5 cause the multisystem disorder Vici syndrome. Here we demonstrated that EPG5 is a Rab7 effector that determines the fusion specificity of autophagosomes with late endosomes/lysosomes. EPG5 is recruited to late endosomes/lysosomes by direct interaction with Rab7 and the late endosomal/lysosomal R-SNARE VAMP7/8. EPG5 also binds to LC3/LGG-1 (mammalian and C. elegans Atg8 homolog, respectively) and to assembled STX17-SNAP29 Qabc SNARE complexes on autophagosomes. EPG5 stabilizes and facilitates the assembly of STX17-SNAP29-VAMP7/8 trans-SNARE complexes, and promotes STX17-SNAP29-VAMP7-mediated fusion of reconstituted proteoliposomes. Loss of EPG5 activity causes abnormal fusion of autophagosomes with various endocytic vesicles, in part due to elevated assembly of STX17-SNAP25-VAMP8 complexes. SNAP25 knockdown partially suppresses the autophagy defect caused by EPG5 depletion. Our study reveals that EPG5 is a Rab7 effector involved in autophagosome maturation, providing insight into the molecular mechanism underlying Vici syndrome. Copyright © 2016 Elsevier Inc. All rights reserved.

Arf6-Dependent Intracellular Trafficking of Pasteurella multocida Toxin and pH-Dependent Translocation from Late Endosomes

Directory of Open Access Journals (Sweden)

Tracy P. M. Chong

2011-03-01

Full Text Available The potent mitogenic toxin from Pasteurella multocida (PMT is the major virulence factor associated with a number of epizootic and zoonotic diseases caused by infection with this respiratory pathogen. PMT is a glutamine-specific protein deamidase that acts on its intracellular G-protein targets to increase intracellular calcium, cytoskeletal, and mitogenic signaling. PMT enters cells through receptor-mediated endocytosis and then translocates into the cytosol through a pH-dependent process that is inhibited by NH4Cl or bafilomycin A1. However, the detailed mechanisms that govern cellular entry, trafficking, and translocation of PMT remain unclear. Co-localization studies described herein revealed that while PMT shares an initial entry pathway with transferrin (Tfn and cholera toxin (CT, the trafficking pathways of Tfn, CT, and PMT subsequently diverge, as Tfn is trafficked to recycling endosomes, CT is trafficked retrograde to the ER, and PMT is trafficked to late endosomes. Our studies implicate the small regulatory GTPase Arf6 in the endocytic trafficking of PMT. Translocation of PMT from the endocytic vesicle occurs through a pH-dependent process that is also dependent on both microtubule and actin dynamics, as evidenced by inhibition of PMT activity in our SRE-based reporter assay, with nocodazole and cytochalasin D, respectively, suggesting that membrane translocation and cytotoxicity of PMT is dependent on its transfer to late endosomal compartments. In contrast, disruption of Golgi-ER trafficking with brefeldin A increased PMT activity, suggesting that inhibiting PMT trafficking to non-productive compartments that do not lead to translocation, while promoting formation of an acidic tubulovesicle system more conducive to translocation, enhances PMT translocation and activity.

Involvement of the endosomal-lysosomal system correlates with regional pathology in Creutzfeldt-Jakob disease

DEFF Research Database (Denmark)

Kovács, Gábor G; Gelpi, Ellen; Ströbel, Thomas

2007-01-01

The endosomal-lysosomal system (ELS) has been suggested to play a role in the pathogenesis of prion diseases. The purpose of this study was to examine how experimental observations can be translated to human neuropathology and whether alterations of the ELS relate to neuropathologic changes...... correlate with regional pathology. Overloading of this system might impair the function of lysosomal enzymes and thus may mimic some features of lysosomal storage disorders. Udgivelsesdato: 2007-Jul...

Cellular regulation of the dopamine transporter

DEFF Research Database (Denmark)

Eriksen, Jacob

2010-01-01

-membrane spanning protein Tac, thereby creating an extracellular antibody epitope. Upon expression in HEK293 cells this TacDAT fusion protein displayed functional properties similar to the wild type transporter. In an ELISA based internalization assay, TacDAT intracellular accumulation was increased by inhibitors...... of lysosomal degradation and moreover TacDAT colocalized with the late endosomal marker Rab7. In the DA cell line 1Rb3An27 TacDAT also co-localized with EGFP-Rab7 and not with the recycling endosomal marker EGFP-Rab11. To assess whether sorting to late endosomes/lysosomes was a property also inherent...... to natively expressed transporter, DAT was visualized directly in cultured DA neurons using the fluorescent cocaine analog JHC 1-64. These data showed pronounced colocalization upon constitutive internalization with Lysotracker, a late endosomal/lysosomal marker; however only little cololization was observed...

Causes of road traffic accidents in Juba

Directory of Open Access Journals (Sweden)

Akway M. Cham

2017-05-01

Full Text Available Introduction: Road traffic accidents (RTAs are a major cause of death and disability in South Sudan. The purpose of this study was to investigate whether violation of traffic rules is the main cause of RTAs. Method: A cross sectional study design was used with quantitative data covering January – December 2014. The main objective of the research was to understand the epidemiology of RTAs in order to develop preventive measures. A total of 1725cases from road RTAs data were extracted from the directorate of traffic police Central Equatoria state Juba and Juba teaching hospital. Results: Most (99.5% of the RTA drivers were not under the influence of alcohol. Most accidents were caused by male drivers (99%. The highest number of RTAs took place in August (11%. Drivers of private vehicles caused most accidents (37%. Most drivers (46% were aged 20-30 years. RTAs occurred most often on city roads (89.83%. Conclusion: This leads us to conclude that a comprehensive safety system is needed that are premised on the idea of community-based awareness of traffic rules and safety regulations. Resources are limited so there is a need to harness local resources including the local community. More efforts are needed to improve road safety education among the youth/integrate safety into road design.

Visual impairment and road traffic accidents among drivers in Jimma Town, Southwest Ethiopia.

Science.gov (United States)

Biza, Mohamed; Mossie, Andualem; Woldemichael, Kifle; Gelaw, Yeshigeta

2013-04-01

Vision play a vital role in driving where good and efficient visual functioning of the driver is essential. Any significant loss of visual function will diminish a driver's ability to operate a motor vehicle safely and will thus contribute to road traffic injury. However, there is little evidence indicating that defects of vision alone cause road traffic accidents. To determine the impact of visual impairment and other factors on road traffic accident among vehicle drivers. A cross-sectional descriptive study was conducted on 249 sampled drivers in Southwest Ethiopia. A pretested interviewer led questionnaire was used for interview and vision tests were done using Snellen's acuity chart and Ishihara pseudo-isochromatic plates. Statistical analyses were performed using SPSS version 16.0. The mean age of drivers was 33.6 years (SD +/- 10.3). The relative frequency of self reported road traffic accident was 15.3%. The prevalence of uncorrected binocular visual impairment was 1.6% and there was a significant association between visual impairment and road traffic accident (P road traffic accident. There is need for consistent inspection and screening, strict rules and regulations of licensing and health education for drivers to minimize road traffic accident.

Impacts of Traffic Noise and Traffic Volume on Birds of Roadside Habitats

Directory of Open Access Journals (Sweden)

Kirsten M. Parris

2009-06-01

Full Text Available Roadside habitats are important for a range of taxa including plants, insects, mammals, and birds, particularly in developed countries in which large expanses of native vegetation have been cleared for agriculture or urban development. Although roadside vegetation may provide suitable habitat for many species, resident animals can be exposed to high levels of traffic noise, visual disturbance from passing vehicles, and the risk of collision with cars and trucks. Traffic noise can reduce the distance over which acoustic signals such as song can be detected, an effect known as acoustic interference or masking. Studies from the northern hemisphere show that the singing behavior of birds changes in the presence of traffic noise. We investigated the impact of traffic noise and traffic volume on two species of birds, the Grey Shrike-thrush (Colluricincla harmonica and the Grey Fantail (Rhipidura fuliginosa, at 58 roadside sites on the Mornington Peninsula, southeastern Australia. The lower singing Grey Shrike-thrush sang at a higher frequency in the presence of traffic noise, with a predicted increase in dominant frequency of 5.8 Hz/dB of traffic noise, and a total effect size of 209 Hz. In contrast, the higher singing Grey Fantail did not appear to change its song in traffic noise. The probability of detecting each species on a visit to a site declined substantially with increasing traffic noise and traffic volume, with several lines of evidence supporting a larger effect of traffic noise. Traffic noise could hamper detection of song by conspecifics, making it more difficult for birds to establish and maintain territories, attract mates and maintain pair bonds, and possibly leading to reduced breeding success in noisy roadside habitats. Closing key roads during the breeding season is a potential, but untested, management strategy to protect threatened bird species from traffic noise and collision with vehicles at the time of year when they are most

Actin-Sorting Nexin 27 (SNX27)-Retromer Complex Mediates Rapid Parathyroid Hormone Receptor Recycling*

Science.gov (United States)

McGarvey, Jennifer C.; Xiao, Kunhong; Bowman, Shanna L.; Mamonova, Tatyana; Zhang, Qiangmin; Bisello, Alessandro; Sneddon, W. Bruce; Ardura, Juan A.; Jean-Alphonse, Frederic; Vilardaga, Jean-Pierre; Puthenveedu, Manojkumar A.; Friedman, Peter A.

2016-01-01

The G protein-coupled parathyroid hormone receptor (PTHR) regulates mineral-ion homeostasis and bone remodeling. Upon parathyroid hormone (PTH) stimulation, the PTHR internalizes into early endosomes and subsequently traffics to the retromer complex, a sorting platform on early endosomes that promotes recycling of surface receptors. The C terminus of the PTHR contains a type I PDZ ligand that binds PDZ domain-containing proteins. Mass spectrometry identified sorting nexin 27 (SNX27) in isolated endosomes as a PTHR binding partner. PTH treatment enriched endosomal PTHR. SNX27 contains a PDZ domain and serves as a cargo selector for the retromer complex. VPS26, VPS29, and VPS35 retromer subunits were isolated with PTHR in endosomes from cells stimulated with PTH. Molecular dynamics and protein binding studies establish that PTHR and SNX27 interactions depend on the PDZ recognition motif in PTHR and the PDZ domain of SNX27. Depletion of either SNX27 or VPS35 or actin depolymerization decreased the rate of PTHR recycling following agonist stimulation. Mutating the PDZ ligand of PTHR abolished the interaction with SNX27 but did not affect the overall rate of recycling, suggesting that PTHR may directly engage the retromer complex. Coimmunoprecipitation and overlay experiments show that both intact and mutated PTHR bind retromer through the VPS26 protomer and sequentially assemble a ternary complex with PTHR and SNX27. SNX27-independent recycling may involve N-ethylmaleimide-sensitive factor, which binds both PDZ intact and mutant PTHRs. We conclude that PTHR recycles rapidly through at least two pathways, one involving the ASRT complex of actin, SNX27, and retromer and another possibly involving N-ethylmaleimide-sensitive factor. PMID:27008860

Ethical Predispositions to Violate or Obey Traffic Rules and the Mediating Role of Driving Styles.

Science.gov (United States)

Holman, Andrei C; Popusoi, Simona A

2018-07-04

Traffic law violations are the most important determinants of traffic accidents. This study examined the influence of drivers' ethical perspective on their traffic violations and the mediating role of driving styles in these relationships. A sample of 313 drivers participated in the study. Contrary to expectations, high ethical relativism was found to be associated with fewer aggressive traffic violations. This suggests that the informal normative system that parallels the official traffic regulations is the more salient reference against which drivers usually assess the adequacy of their behaviors, highly relativistic drivers being more willing to also take into consideration the official traffic code in these appraisals. We also found significant interactions between idealism and relativism on both ordinary and aggressive violations. Idealism emerged as a predictor of these violations only in drivers low in relativism. In this group, one's concern about the harm that he/she might cause to other traffic participants induces higher traffic law abidingness. These influences of idealism were mediated by certain driving styles. Drivers' general ethical perspective fosters a specific pattern of driving styles and, consequently, the tendency to commit or to refrain from traffic violations.

Modeling the heterogeneous traffic correlations in urban road systems using traffic-enhanced community detection approach

Science.gov (United States)

Lu, Feng; Liu, Kang; Duan, Yingying; Cheng, Shifen; Du, Fei

2018-07-01

A better characterization of the traffic influence among urban roads is crucial for traffic control and traffic forecasting. The existence of spatial heterogeneity imposes great influence on modeling the extent and degree of road traffic correlation, which is usually neglected by the traditional distance based method. In this paper, we propose a traffic-enhanced community detection approach to spatially reveal the traffic correlation in city road networks. First, the road network is modeled as a traffic-enhanced dual graph with the closeness between two road segments determined not only by their topological connection, but also by the traffic correlation between them. Then a flow-based community detection algorithm called Infomap is utilized to identify the road segment clusters. Evaluated by Moran's I, Calinski-Harabaz Index and the traffic interpolation application, we find that compared to the distance based method and the community based method, our proposed traffic-enhanced community based method behaves better in capturing the extent of traffic relevance as both the topological structure of the road network and the traffic correlations among urban roads are considered. It can be used in more traffic-related applications, such as traffic forecasting, traffic control and guidance.

How and when urban vehicular traffic pollutes the environment

International Nuclear Information System (INIS)

Cera, L.; Di Mascio, P.

1993-01-01

This paper first compares monitored values of urban traffic air and noise pollution with Italian and European Communities pollution limits to assess the efficacy of current and proposed pollution regulations. The paper then focuses on the environmental impacts of the generation and discharge of waste waters during the construction and use of road surfaces. The related problems discussed include: soil erosion due to the increased exposure of bordering terrestrial ecosystems to damaging atmospheric pollutants; and the reception by roadside land areas of pollutants absorbed in rainfall runoff which has come into contact with road surfaces. Some suggestions are made on how to abate these particular types of traffic derived pollutants

WASH and WAVE actin regulators of the Wiskott-Aldrich syndrome protein (WASP) family are controlled by analogous structurally related complexes.

Science.gov (United States)

Jia, Da; Gomez, Timothy S; Metlagel, Zoltan; Umetani, Junko; Otwinowski, Zbyszek; Rosen, Michael K; Billadeau, Daniel D

2010-06-08

We recently showed that the Wiskott-Aldrich syndrome protein (WASP) family member, WASH, localizes to endosomal subdomains and regulates endocytic vesicle scission in an Arp2/3-dependent manner. Mechanisms regulating WASH activity are unknown. Here we show that WASH functions in cells within a 500 kDa core complex containing Strumpellin, FAM21, KIAA1033 (SWIP), and CCDC53. Although recombinant WASH is constitutively active toward the Arp2/3 complex, the reconstituted core assembly is inhibited, suggesting that it functions in cells to regulate actin dynamics through WASH. FAM21 interacts directly with CAPZ and inhibits its actin-capping activity. Four of the five core components show distant (approximately 15% amino acid sequence identify) but significant structural homology to components of a complex that negatively regulates the WASP family member, WAVE. Moreover, biochemical and electron microscopic analyses show that the WASH and WAVE complexes are structurally similar. Thus, these two distantly related WASP family members are controlled by analogous structurally related mechanisms. Strumpellin is mutated in the human disease hereditary spastic paraplegia, and its link to WASH suggests that misregulation of actin dynamics on endosomes may play a role in this disorder.

Comparison of Microscopic Drivers' Probabilistic Lane-changing Models With Real Traffic Microscopic Data

Directory of Open Access Journals (Sweden)

Seyyed Mohammad Sadat Hoseini

2011-07-01

Full Text Available The difficulties of microscopic-level simulation models to accurately reproduce real traffic phenomena stem not only from the complexity of calibration and validation operations, but also from the structural inadequacies of the sub-models themselves. Both of these drawbacks originate from the scant information available on real phenomena because of the difficulty in gathering accurate field data. This paper studies the traffic behaviour of individual drivers utilizing vehicle trajectory data extracted from digital images collected from freeways in Iran. These data are used to evaluate the four proposed microscopic traffic models. One of the models is based on the traffic regulations in Iran and the three others are probabilistic models that use a decision factor for calculating the probability of choosing a position on the freeway by a driver. The decision factors for three probabilistic models are increasing speed, decreasing risk of collision, and increasing speed combined with decreasing risk of collision. The models are simulated by a cellular automata simulator and compared with the real data. It is shown that the model based on driving regulations is not valid, but that other models appear useful for predicting the driver’s behaviour on freeway segments in Iran during noncongested conditions.

Adaptive traffic signal control with actor-critic methods in a real-world traffic network with different traffic disruption events

NARCIS (Netherlands)

Aslani, Mohammad; Mesgari, Mohammad Saadi; Wiering, Marco

2017-01-01

The transportation demand is rapidly growing in metropolises, resulting in chronic traffic con-gestions in dense downtown areas. Adaptive traffic signal control as the principle part of in-telligent transportation systems has a primary role to effectively reduce traffic congestion by making a

A Study on the Model of Traffic Flow and Vehicle Exhaust Emission

Directory of Open Access Journals (Sweden)

Han Xue

2013-01-01

Full Text Available The increase of traffic flow in cities causes traffic congestion and accidents as well as air pollution. Traffic problems have attracted the interest of many researchers from the perspective of theory and engineering. In order to provide a simple and practical method for measuring the exhaust emission and assessing the effect of pollution control, a model is based on the relationship between traffic flow and vehicle exhaust emission under a certain level of road capacity constraints. In the proposed model, the hydrocarbons (HC, carbon monoxide (CO, and nitrogen oxides (NOx are considered as the indexes of total exhaust emission, and the speed is used as an intermediate variable. To verify the rationality and practicality of the model, a case study for Beijing, China, is provided in which the effects of taxi fare regulation and the specific vehicle emission reduction policy are analyzed.

From Goods to Traffic:First Steps Toward an Auction-based Traffic Signal Controller

OpenAIRE

Raphael, Jeffery; Maskell, Simon; Sklar, Elizabeth Ida

2015-01-01

Traffic congestion is a major issue that plagues many urban road networks large and small. Traffic engineers are now leaning towards Intelligent Traffic Systems as many physical changes to road networks are costly or infeasible. Multi-Agent Systems (MAS) have become a popular paradigm for intelligent solutions to traffic management problems. There are many MAS approaches to traffic management that utilise market mechanisms. In market-based approaches, drivers “pay” to use the roadways. Howeve...

A Two-Stage Fuzzy Logic Control Method of Traffic Signal Based on Traffic Urgency Degree

OpenAIRE

Yan Ge

2014-01-01

City intersection traffic signal control is an important method to improve the efficiency of road network and alleviate traffic congestion. This paper researches traffic signal fuzzy control method on a single intersection. A two-stage traffic signal control method based on traffic urgency degree is proposed according to two-stage fuzzy inference on single intersection. At the first stage, calculate traffic urgency degree for all red phases using traffic urgency evaluation module and select t...

Safety in traffic for vulnerable military road users

Directory of Open Access Journals (Sweden)

Aleksandar J. Bulajić

2012-04-01

Full Text Available The Army of Serbia, as a relatively closed system, regulates the field of traffic safety; however, during peacetime, general rules apply to all participants in traffic circulation. The Republic of Serbia is in the group of countries with a high number of road fatalities. The level of traffic safety in the Serbian Army has been on constant increase since 2000, although the relevant transport authorities in the military are not yet satisfied with the achieved level (even one lost life is too much. The increase can be deceptive, since if we take into account the substantial reduction in vehicle use due to various factors in the last few years (poor financial situation in the country as well as in the military, under-investment in the purchase of new vehicles, purchase of transportation services, fewer drivers drafted and more vehicles driven by trained officers, etc., it is not surprising that there are fewer accidents and fewer road deaths and injuries among military personnel. This paper aims at approaching the problem of pedestrian safety as a segment of road safety and at making an educational impact on all members of the military, because they all participate in traffic daily, if not as drivers or passengers, then certainly in large numbers as pedestrians. The basis of this paper is aimed at shedding light on the causes of pedestrian road fatalities due to their mistakes, i. e. 'negligence' of the participants in car accidents with the participation of pedestrians, as well as at proposing measures to reduce and prevent traffic accidents with pedestrians.

Properties of Traffic Risk Coefficient

Science.gov (United States)

Tang, Tie-Qiao; Huang, Hai-Jun; Shang, Hua-Yan; Xue, Yu

2009-10-01

We use the model with the consideration of the traffic interruption probability (Physica A 387(2008)6845) to study the relationship between the traffic risk coefficient and the traffic interruption probability. The analytical and numerical results show that the traffic interruption probability will reduce the traffic risk coefficient and that the reduction is related to the density, which shows that this model can improve traffic security.

Analysis of vehicular traffic flow in the major areas of Kuala Lumpur utilizing open-traffic

Science.gov (United States)

Manogaran, Saargunawathy; Ali, Muhammad; Yusof, Kamaludin Mohamad; Suhaili, Ramdhan

2017-09-01

Vehicular traffic congestion occurs when a large number of drivers are overcrowded on the road and the traffic flow does not run smoothly. Traffic congestion causes chaos on the road and interruption to daily activities of users. Time consumed on road give lots of negative effects on productivity, social behavior, environmental and cost to economy. Congestion is worsens and leads to havoc during the emergency such as flood, accidents, road maintenance and etc., where behavior of traffic flow is always unpredictable and uncontrollable. Real-time and historical traffic data are critical inputs for most traffic flow analysis applications. Researcher attempt to predict traffic using simulations as there is no exact model of traffic flow exists due to its high complexity. Open Traffic is an open source platform available for traffic data analysis linked to Open Street Map (OSM). This research is aimed to study and understand the Open Traffic platform. The real-time traffic flow pattern in Kuala Lumpur area was successfully been extracted and analyzed using Open Traffic. It was observed that the congestion occurs on every major road in Kuala Lumpur and most of it owes to the offices and the economic and commercial centers during rush hours. At some roads the congestion occurs at night due to the tourism activities.

Regulation of B cell differentiation by intracellular membrane associated proteins and microRNAs: role in the antibody response

Directory of Open Access Journals (Sweden)

Zheng eLou

2015-10-01

Full Text Available B cells are central to adaptive immunity and their functions in antibody responses are exquisitely regulated. As suggested by recent findings, B cell differentiation is mediated by intracellular membrane structures (including endosomes, lysosomes and autophagosomes and protein factors specifically associated with these membranes, including Rab7, Atg5 and Atg7. These factors participate in vesicle formation/trafficking, signal transduction and induction of gene expression to promote antigen presentation, CSR/SHM, and generation/maintenance of plasma cells and memory B cells. Their expression is induced in B cells activated to differentiate and further fine-tuned by immune-modulating microRNAs, which coordinates CSR/SHM, plasma cell differentiation and memory B cell differentiation. These short non-coding RNAs would individually target multiple factors associated with the same intracellular membrane compartments and collaboratively target a single factor in addition to regulate AID and Blimp-1. These, together with regulation of microRNA biogenesis and activities by endosomes and autophagosomes, show that intracellular membranes and microRNAs, two broadly relevant cell constituents, play important roles in balancing gene expression to specify B cell differentiation processes for optimal antibody responses.

Road Traffic Congestion Management Based on a Search-Allocation Approach

Directory of Open Access Journals (Sweden)

Raiyn Jamal

2017-03-01

Full Text Available This paper introduces a new scheme for road traffic management in smart cities, aimed at reducing road traffic congestion. The scheme is based on a combination of searching, updating, and allocation techniques (SUA. An SUA approach is proposed to reduce the processing time for forecasting the conditions of all road sections in real-time, which is typically considerable and complex. It searches for the shortest route based on historical observations, then computes travel time forecasts based on vehicular location in real-time. Using updated information, which includes travel time forecasts and accident forecasts, the vehicle is allocated the appropriate section. The novelty of the SUA scheme lies in its updating of vehicles in every time to reduce traffic congestion. Furthermore, the SUA approach supports autonomy and management by self-regulation, which recommends its use in smart cities that support internet of things (IoT technologies.

Finnish air traffic until 2025. Four scenarios; Suomen lentoliikenne vuoteen 2025 - neljae skenaariota

Energy Technology Data Exchange (ETDEWEB)

Aalto, E.; Pollanen, M.; Mantynen, J.; Makela, T.; Rauhamaki, H. [Tampere Univ. of Technology (Finland). Transport Research Centre Verne

2012-07-01

Air traffic is a very dynamic field of business. Its operating environment features multiple factors of change, which affect air traffic and its operators both in the short and in the long run. This study charts the significant factors in Finnish air traffic and creates four distinct scenarios for the future. These scenarios were produced on the basis of the currently available facts and are attempts to provide several alternatives for the future. The study's goal has been to produce a visionary depiction that helps one to visualise and understand the whole picture. The perspective chosen for consideration of the future is the year 2025. The study describes four scenarios differing clearly from each other. In the first, entitled Globaalista maailmasta blokkeihin (From a global world to blocs), states form clusters and regional co-operation takes precedence over the current global direction of development. The second, Eurooppa omalla reitillaeaen (Europe on its own way), depicts Europe ahead of the rest of the world in terms of environmental regulation for air traffic. Uudistuksilla talouskukoistukseen (Economic boom through reforms) is based on an optimistic economic outlook. In contrast, Oeljykriisistae taloustaantumaan (From oil crisis to recession), describes a world where economic growth has been forestalled by rising oil prices due to increasing scarcity. The study investigates passenger numbers, air traffic within Finland, the international route network, and the changes these display in the various scenarios. The estimates are designed to highlight the development trends affecting air traffic in different operating environments. Since Finland is a small country and set apart from the rest of Europe in a fashion similar to island states, air traffic is of major importance and cannot be efficiently replaced by other forms of traffic. In particular, the individual scenarios stress the impact of international interaction and economic development with regard

Automobile control technology and traffic control

Energy Technology Data Exchange (ETDEWEB)

Takaba, Sadao [Univ. of Tokyo (Japan)

1988-09-05

In the field of automobile control technology, electronic was first adopted for the electronic fuel control as an answer to the exhaust gas regulations. The operations of the driving system, frame system or the automobile itself which is the combination of the two were optimized by adding sensors, computers, actuators, etc. to alleviate the burden of the driver, offering easier drivability and confortableness. For local driving control, measurement of distance up to obstacle has been practiced using the ultrasonic radar sensor. Research and development of microwave radar sensor have been carried out for years. Automatic driving has been a dream technology, and the study for the technology was started since early times. Remarkable progress was made recently in the navigation system for traffic control in wide area. New automobile traffic information communication and other systems are being developed. Historical description is made on the control and information systems for road transportation, dividing the period into the 1st, 2nd and 3rd generations. 10 references.

Traffic safety strategies

Directory of Open Access Journals (Sweden)

V. Sadauskas

2003-04-01

Full Text Available Fast development of the number of vehicles is closely related not only to large benefit for the public but also to certain undesirable social and economic consequences. Firstly - large numbers of injured and killed people are involved into the accidents. The target to improve traffic safety situation in Lithuania can be reached only after the detailed evaluation of transport system, environment, traffic participants, road and vehicle. Taking into consideration the accident situation in Lithuania and its causes the followings priority trends are suggested: The improvement of the coordination of road traffic safety system, the training and education of road users, the explanation of the importance of traffic safety and its propagation, the improvement of traffic conditions. Recommendations and proposals for differentiated criterion of maximum speed limit selection taking into account different factors are provided in the work.

Travelers ability to observe changes in traffic intensities and traffic light settings

NARCIS (Netherlands)

Vreeswijk, Jacob Dirk; Do, Michael; Middag, Wilco; Martens, Marieke Hendrikje; van Berkum, Eric C.; van Arem, Bart; ITSC,

2011-01-01

Travel choice behavior is an important determinant in traffic and subject to human imperfection and bounded rationality. In decision-making processes travelers seldom act perfectly rational. Traffic models and traffic network management measure could become more realistic and effective, if

Traffic breakdown at a signal: classical theory versus the three-phase theory of city traffic

International Nuclear Information System (INIS)

Kerner, Boris S; Schreckenberg, Michael; Klenov, Sergey L

2014-01-01

Physical reasons for a crucial difference between the results of a three-phase theory developed recently (Kerner 2011 Phys. Rev. E 84 045102(R); 2013 Europhys. Lett. 102 28010; 2014 Physica A 397 76) and the classical theory are explained. Microscopic characteristics of traffic passing a traffic signal during the green signal phase and their dependence on the duration of the green phase have been found. It turns out that a moving synchronized flow pattern (MSP), which occurs in under-saturated traffic at the signal, causes ‘compression’ of traffic flow: the rate of MSP discharge can be considerably larger than the saturation flow rate of the classical traffic theory of city traffic. This leads to a considerably larger rate of traffic passing the signal in comparison with the saturation flow rate. This effect together with traffic behavior at the upstream queue front explains the metastability of under-saturated traffic with respect to a random time-delayed traffic breakdown. (paper)

Real time traffic models, decision support for traffic management

NARCIS (Netherlands)

Wismans, Luc Johannes Josephus; de Romph, E.; Friso, K.; Zantema, K.

2014-01-01

Reliable and accurate short-term traffic state prediction can improve the performance of real-time traffic management systems significantly. Using this short-time prediction based on current measurements delivered by advanced surveillance systems will support decision-making processes on various

Real Time Traffic Models, Decision Support for Traffic Management

NARCIS (Netherlands)

Wismans, L.; De Romph, E.; Friso, K.; Zantema, K.

2014-01-01

Reliable and accurate short-term traffic state prediction can improve the performance of real-time traffic management systems significantly. Using this short-time prediction based on current measurements delivered by advanced surveillance systems will support decision-making processes on various

Remark on the Causes of Traffic Accidents and Traffic Awareness: Examples of Usak Province

Directory of Open Access Journals (Sweden)

Ercan ÖZEN

2014-01-01

Full Text Available Traffic accidents cause important losses on the social and economics structure of countries. While economic losses are able to be compensated in a short time, it isn’t possible to compensate social and communal losses. This study tackles with not the economic aspect of traffic accidents but the social dimension, evaluates the causes of the traffic accidents with respect to views of individuals on traffic and aims to measure the levels of awareness of individuals about traffic. To achieve these goals, a group of 506 people were surveyed in Uşak province and the acquired data were analyzed via one-way ANOVA test in SPSS 16 program. According to the analysis results, drivers and pedestrians are considered to have significant flaws in traffic accidents. But alongside this thought, the idea that traffic accidents are caused by the road having technical and physical problems has come to the fore. This results present a valid motive for the established approach “Vision Zero” , being practised in Sweden, to be adopted in Turkey. When the level of awareness about traffic is examined, it is seen that individuals generally have got inadequate information related traffic rules and traffic authorities. Besides, the fact that individuals are inadequate in terms of courtesy and respect in traffic and that more serious efforts on traffic education is needed to be spent come out as the important findings of this study. The findings of this study can be thought as a directive tool to the policy makers.

Evaluation of diversion strategies in the context of advanced traffic management systems (ATMS) for an urban traffic corridor with heterogeneous traffic

Energy Technology Data Exchange (ETDEWEB)

Korlapati, D.R.

2007-07-01

Due to urbanization and accelerated growth in vehicular traffic, most big cities in India face problems related to traffic management resulting in severe congestion, pollution, and a high rate of accidents during peak hours. Lane blocking incidents on arterials or urban traffic corridors cause major disruption to traffic flow. Peak hour congestion with low average speeds and high accident rates are commonly associated with traffic in major cities in India. The situation is deteriorating further as creation of new facilities are almost impossible, with resource and space constraints. In such scenarios, application of advanced technologies seems to offer hope. One such application area is Advanced Traffic Management Systems (ATMS), a component of intelligent transportation system (ITS). Due to the unique traffic characteristics prevailing in India, the application of such systems needs to first be evaluated before implementation. This paper proposed a research methodology for the evaluation of diversion strategies in the context of ATMS for an urban corridor in India. The evaluation framework combined several relevant modules related to various aspects of traffic control, surveillance and advisory. As part of this study, a simulation model and a simulation optimization model were developed. The simulation model was microscopic in nature and captured the driver behaviour and traffic characteristics realistically by modeling the complex interactions among vehicles traversing a corridor. It was concluded that the results and observations were useful indicators to gauge the potential success of diversion plans. 10 refs., 1 tab., 2 figs.

Alcohol abuse, road traffic and safety on the site

CERN Multimedia

DSU Department

2008-01-01

In the light of the serious road accident that occurred on 3 February 2008, you are reminded that: the legal blood alcohol limit in force under the Host States’ traffic regulations (0.5 g per 1000 ml of blood) also applies on the CERN site; the consumption of alcohol is forbidden during working hours and is only tolerated in the restaurants at certain times, unless an exception is granted for special events; failure to observe these rules may result in disciplinary action by the Organization, independently of any sanctions that may be applicable pursuant to the road traffic regulations of the Host State concerned. Furthermore, the Reception and Access Control Service, the site guards and the Fire Brigade have been instructed to stop any driver in an obvious state of intoxication and to ask him to abandon his vehicle on the spot. In case of disagreement, they may also suggest that he submits to a voluntary blood alcohol level test at the CERN Medical Service or Fire Brigade...

Examining perimeter gating control of urban traffic networkswith locally adaptive traffic signals

NARCIS (Netherlands)

Keyvan Ekbatani, M.; Gao, X.; Gayah, V.V.; Knoop, V.L.

2015-01-01

Traditionally, urban traffic is controlled by traffic lights. Recent findings of the Macroscopic or Network Fundamental Diagram (MFD or NFD) have led to the development of novel traffic control strategies that can be applied at a networkwide level. One pertinent example is perimeter flow control

Modernizing the Professional Capabilities of Driving Instructors and Traffic Safety Teachers

Directory of Open Access Journals (Sweden)

Kolarov Ivan

2009-06-01

Full Text Available The effective traffic safety training is a long process; it starts from very young age and continues throughout life. This is not only a process of acquiring knowledge and skills, but also a matter of social importance. Pedagogical forms of education depend on the position of a man in society, respectively, on abilities to percept the environment. The recent investigations of traffic accidents show, that the young drivers generally have poorer than average self-reflection skills. It is therefore important to foster self-reflection skills and attitudes from an early age, especially in matters that relate to road and driving safety. All of staff that have career in traffic safety training (mainly instructors for driving and traffic safety teachers in Europe have their responsibility for decreasing the traffic accidents and incidents by polishing their skills. They work under variety conditions, specific National Lows and Regulations, but the features of their target groups are one and the same. In order to meet the new challenges put by EU Commission they need to have common vision about traffic safety training in Europe, to know what traffic safety means for different age groups, to know what is the best practice of their colleagues, including curricula, methods, training materials, and to be aware their work is very important for traffic safety. Aim of this paper is to present an approach for non-formal instructors for driving and traffic safety teachers training for modernizing their professional capabilities with students form 0 to 30 years old. The investigations and main results are based on the theoretical investigation for hierarchical level of behavior, made by Hatakka for instructors for driving training. The students are divided in four age groups: pre-school children (0-6 years, schoolchildren (7-12 years, teenagers (13-17 years and young adults (18-30 years. For each age group a curriculum is developed according to common EU rules and

Traffic management simulation development : summary.

Science.gov (United States)

2011-01-01

Increasingly, Florida traffic is monitored electronically by components of the Intelligent Traffic System (ITS), which send data to regional traffic management centers and assist management of traffic flows and incident response using software called...

A novel chimeric cell-penetrating peptide with membrane-disruptive properties for efficient endosomal escape.

Science.gov (United States)

Salomone, Fabrizio; Cardarelli, Francesco; Di Luca, Mariagrazia; Boccardi, Claudia; Nifosì, Riccardo; Bardi, Giuseppe; Di Bari, Lorenzo; Serresi, Michela; Beltram, Fabio

2012-11-10

Efficient endocytosis into a wide range of target cells and low toxicity make the arginine-rich Tat peptide (Tat(11): YGRKKRRQRRR, residues 47-57 of HIV-1 Tat protein) an excellent transporter for delivery purposes. Unfortunately, molecules taken up by endocytosis undergo endosomal entrapment and possible metabolic degradation. Escape from the endosome is therefore actively researched. In this context, antimicrobial peptides (AMPs) provide viable templates for the design of new membrane-disruptive motifs. In particular the Cecropin-A and Melittin hybrids (CMs) are among the smallest and most effective peptides with membrane-perturbing abilities. Here we present a novel chimeric peptide in which the Tat(11) motif is fused to the CM(18) hybrid (KWKLFKKIGAVLKVLTTG, residues 1-7 of Cecropin-A and 2-12 of Melittin). When administered to cells, CM(18)-Tat(11) combines the two desired functionalities: efficient uptake and destabilization of endocytotic-vesicle membranes. We show that this chimeric peptide effectively increases cargo-molecule cytoplasm availability and allows the subsequent intracellular localization of diverse membrane-impermeable molecules (i.e. Tat(11)-EGFP fusion protein, calcein, dextrans, and plasmidic DNA) with no detectable cytotoxicity. The present results open the way to the rational engineering of "modular" cell-penetrating peptides (CPPs) that combine (i) efficient translocation from the extracellular milieu into vesicles and (ii) efficient release of molecules from vesicles into the cytoplasm. Copyright © 2012 Elsevier B.V. All rights reserved.

Bcl-xL regulates CD1d-mediated antigen presentation to NKT cells by altering CD1d trafficking through the endocytic pathway.

Science.gov (United States)

Subrahmanyam, Priyanka B; Carey, Gregory B; Webb, Tonya J

2014-09-01

NKT cells are a unique subset of T cells that recognize glycolipid Ags presented in the context of CD1d molecules. NKT cells mount strong antitumor responses and are a major focus in developing effective cancer immunotherapy. It is known that CD1d molecules are constantly internalized from the cell surface, recycled through the endocytic compartments, and re-expressed on the cell surface. However, little is known about the regulation of CD1d-mediated Ag processing and presentation in B cell lymphoma. Prosurvival factors of the Bcl-2 family, such as Bcl-xL, are often upregulated in B cell lymphomas and are intimately linked to sphingolipid metabolism, as well as the endocytic compartments. We hypothesized that Bcl-xL can regulate CD1d-mediated Ag presentation to NKT cells. We found that overexpression or induction of Bcl-xL led to increased Ag presentation to NKT cells. Conversely, the inhibition or knockdown of Bcl-xL led to decreased NKT cell activation. Furthermore, knockdown of Bcl-xL resulted in the loss of CD1d trafficking to lysosome-associated membrane protein 1(+) compartments. Rab7, a late endosomal protein, was upregulated and CD1d molecules accumulated in the Rab7(+) late endosomal compartment. These results demonstrate that Bcl-xL regulates CD1d-mediated Ag processing and presentation to NKT cells by altering the late endosomal compartment and changing the intracellular localization of CD1d. Copyright © 2014 by The American Association of Immunologists, Inc.

High-level traffic-violation detection for embedded traffic analysis

NARCIS (Netherlands)

Vijverberg, J.A.; de Koning, A.H.M.; Han, Jungong; de With, P.H.N.; Cornelissen, D.

2007-01-01

This paper presents the design of a robust and real-time traffic-violation detection system for cameras on intersections. We use background segmentation and a novel road-model to obtain the candidate traffic participants. A region-based tracking system, equipped with static occlusion-reasoning,

Trajectory Based Traffic Analysis

DEFF Research Database (Denmark)

Krogh, Benjamin Bjerre; Andersen, Ove; Lewis-Kelham, Edwin

2013-01-01

We present the INTRA system for interactive path-based traffic analysis. The analyses are developed in collaboration with traffic researchers and provide novel insights into conditions such as congestion, travel-time, choice of route, and traffic-flow. INTRA supports interactive point-and-click a......We present the INTRA system for interactive path-based traffic analysis. The analyses are developed in collaboration with traffic researchers and provide novel insights into conditions such as congestion, travel-time, choice of route, and traffic-flow. INTRA supports interactive point......-and-click analysis, due to a novel and efficient indexing structure. With the web-site daisy.aau.dk/its/spqdemo/we will demonstrate several analyses, using a very large real-world data set consisting of 1.9 billion GPS records (1.5 million trajectories) recorded from more than 13000 vehicles, and touching most...

African Swine Fever Virus Undergoes Outer Envelope Disruption, Capsid Disassembly and Inner Envelope Fusion before Core Release from Multivesicular Endosomes.

Directory of Open Access Journals (Sweden)

Bruno Hernáez

2016-04-01

Full Text Available African swine fever virus (ASFV is a nucleocytoplasmic large DNA virus (NCLDV that causes a highly lethal disease in domestic pigs. As other NCLDVs, the extracellular form of ASFV possesses a multilayered structure consisting of a genome-containing nucleoid successively wrapped by a thick protein core shell, an inner lipid membrane, an icosahedral protein capsid and an outer lipid envelope. This structural complexity suggests an intricate mechanism of internalization in order to deliver the virus genome into the cytoplasm. By using flow cytometry in combination with pharmacological entry inhibitors, as well as fluorescence and electron microscopy approaches, we have dissected the entry and uncoating pathway used by ASFV to infect the macrophage, its natural host cell. We found that purified extracellular ASFV is internalized by both constitutive macropinocytosis and clathrin-mediated endocytosis. Once inside the cell, ASFV particles move from early endosomes or macropinosomes to late, multivesicular endosomes where they become uncoated. Virus uncoating requires acidic pH and involves the disruption of the outer membrane as well as of the protein capsid. As a consequence, the inner viral membrane becomes exposed and fuses with the limiting endosomal membrane to release the viral core into the cytosol. Interestingly, virus fusion is dependent on virus protein pE248R, a transmembrane polypeptide of the inner envelope that shares sequence similarity with some members of the poxviral entry/fusion complex. Collective evidence supports an entry model for ASFV that might also explain the uncoating of other multienveloped icosahedral NCLDVs.

78 FR 53107 - Drawbridge Operation Regulation; Passaic River, Kearny and Newark, NJ

Science.gov (United States)

2013-08-28

... reducing vehicular traffic congestion during the morning and afternoon vehicular rush hour periods due to... alleviate traffic congestion resulting from area roadway closures. It is expected that this change to the regulations would provide relief to vehicular traffic while continuing to meet the reasonable needs of...

Right-­turn traffic volume adjustment in traffic signal warrant analysis : final report.

Science.gov (United States)

2016-05-06

This report was based on the research project, Right-Turn Traffic Volume Adjustment in Traffic Signal Warrants, sponsored by the Nevada Department of Transportation (NDOT) and SOLARIS. Right-turn traffic does not affect intersection performance in th...

Reorganization of the Endosomal System in Salmonella-Infected Cells: The Ultrastructure of Salmonella-Induced Tubular Compartments

Science.gov (United States)

Krieger, Viktoria; Liebl, David; Zhang, Yuying; Rajashekar, Roopa; Chlanda, Petr; Giesker, Katrin; Chikkaballi, Deepak; Hensel, Michael

2014-01-01

During the intracellular life of Salmonella enterica, a unique membrane-bound compartment termed Salmonella-containing vacuole, or SCV, is formed. By means of translocated effector proteins, intracellular Salmonella also induce the formation of extensive, highly dynamic membrane tubules termed Salmonella-induced filaments or SIF. Here we report the first detailed ultrastructural analyses of the SCV and SIF by electron microscopy (EM), EM tomography and live cell correlative light and electron microscopy (CLEM). We found that a subset of SIF is composed of double membranes that enclose portions of host cell cytosol and cytoskeletal filaments within its inner lumen. Despite some morphological similarities, we found that the formation of SIF double membranes is independent from autophagy and requires the function of the effector proteins SseF and SseG. The lumen of SIF network is accessible to various types of endocytosed material and our CLEM analysis of double membrane SIF demonstrated that fluid phase markers accumulate only between the inner and outer membrane of these structures, a space continual with endosomal lumen. Our work reveals how manipulation of the endosomal membrane system by an intracellular pathogen results in a unique tubular membrane compartmentalization of the host cell, generating a shielded niche permissive for intracellular proliferation of Salmonella. PMID:25254663

SOME EMPIRICAL RELATIONS BETWEEN TRAVEL SPEED, TRAFFIC VOLUME AND TRAFFIC COMPOSITION IN URBAN ARTERIALS

Directory of Open Access Journals (Sweden)

Eleni I. VLAHOGIANNI, Ph.D.

2007-01-01

Full Text Available The effects of traffic mix (the percentage of cars, trucks, buses and so on are of particular interest in the speed-volume relationship in urban signalized arterials under various geometric and control characteristics. The paper presents some empirical observations on the relation between travel speed, traffic volume and traffic composition in urban signalized arterials. A methodology based on emerging self-organizing structures of neural networks to identify regions in the speed-volume relationship with respect to traffic composition and Bayesian networks to evaluate the effect of different types of motorized vehicles on prevailing traffic conditions is proposed. Results based on data from a large urban network indicate that the variability in traffic conditions can be described by eight regions in speed-volume relationship with respect to traffic composition. Further evaluation of the effect of motorized vehicles in each region separately indicates that the effect of traffic composition decreases with the onset of congestion. Moreover, taxis and motorcycles are the primary affecting parameter of the form of the speed-volume relationship in urban arterials.

Endogenous Vascular Endothelial Growth Factor-A (VEGF-A) Maintains Endothelial Cell Homeostasis by Regulating VEGF Receptor-2 Transcription*

Science.gov (United States)

E, Guangqi; Cao, Ying; Bhattacharya, Santanu; Dutta, Shamit; Wang, Enfeng; Mukhopadhyay, Debabrata

2012-01-01

Vascular endothelial growth factor A (VEGF-A) is one of the most important factors controlling angiogenesis. Although the functions of exogenous VEGF-A have been widely studied, the roles of endogenous VEGF-A remain unclear. Here we focused on the mechanistic functions of endogenous VEGF-A in endothelial cells. We found that it is complexed with VEGF receptor 2 (VEGFR-2) and maintains a basal expression level for VEGFR-2 and its downstream signaling activation. Endogenous VEGF-A also controls expression of key endothelial specific genes including VEGFR-2, Tie-2, and vascular endothelial cadherin. Of importance, endogenous VEGF-A differs from exogenous VEGF-A by regulating VEGFR-2 transcription through mediation of FoxC2 binding to the FOX:ETS motif, and the complex formed by endogenous VEGF-A with VEGFR-2 is localized within the EEA1 (early endosome antigen 1) endosomal compartment. Taken together, our results emphasize the importance of endogenous VEGF-A in endothelial cells by regulating key vascular proteins and maintaining the endothelial homeostasis. PMID:22167188

The impact of traffic sign deficit on road traffic accidents in Nigeria.

Science.gov (United States)

Ezeibe, Christian; Ilo, Chukwudi; Oguonu, Chika; Ali, Alphonsus; Abada, Ifeanyi; Ezeibe, Ezinwanne; Oguonu, Chukwunonso; Abada, Felicia; Izueke, Edwin; Agbo, Humphrey

2018-04-04

This study assesses the impact of traffic sign deficit on road traffic accidents in Nigeria. The participants were 720 commercial vehicle drivers. While simple random sampling was used to select 6 out of 137 federal highways, stratified random sampling was used to select six categories of commercial vehicle drivers. The study used qual-dominant mixed methods approach comprising key informant interviews; group interviews; field observation; policy appraisal and secondary literature on traffic signs. Result shows that the failure of government to provide and maintain traffic signs in order to guide road users through the numerous accident black spots on the highways is the major cause of road accidents in Nigeria. The study argues that provision and maintenance of traffic signs present opportunity to promoting safety on the highways and achieving the sustainable development goals.

Using Traffic Conflict Method in Evaluating Traffic Safety at the Reconstructed Intersection

Directory of Open Access Journals (Sweden)

Zdravko Bukljaš

2002-05-01

Full Text Available As part of organised social system, traffic is subjected togeneral social tendency towards adequate safety and sustainabilityof relations in such a system, probabilistically marked bythe risk of danger. Inte1polation of subjective factor facilitatesthe occwTence of negative phenomena. Road traffic system ischaracterised by extremely massive participation in traffic, contributingthus to a greater possibility of negative features characteristicfor imperfect human mechanism. This is precisely thereason why this paper deals with the problem of traffic safety onthe concrete example of the intersection between the SavskaStreet and the Slavonska and LjubljanskaAvenues over the periodof time prior to constmcting the new unde1pass solution,and the period of time immediately after the construction workswere completed. The used data have been provided by the TrafficPolice Department - Lucko.

Strategies for the Activation and Release of the Membranolytic Peptide Melittin from Liposomes Using Endosomal pH as a Trigger

NARCIS (Netherlands)

Oude Blenke, E.; Sleszynska, M.; Evers, M. J W; Storm, G.; Martin, N. I.; Mastrobattista, E.

2017-01-01

Endosomolytic peptides are often coupled to drug delivery systems to enhance endosomal escape, which is crucial for the delivery of macromolecular drugs that are vulnerable to degradation in the endolysosomal pathway. Melittin is a 26 amino acid peptide derived from bee venom that has a very high

Web traffic and firm performance

DEFF Research Database (Denmark)

Farooq, Omar; Aguenaou, Samir

2013-01-01

Does the traffic generated by websites of firms signal anything to stock market participants? Does higher web-traffic translate into availability of more information and therefore lower agency problems? And if answers to above questions are in affirmative, does higher web-traffic traffic translate...... into better firm performance? This paper aims to answer these questions by documenting a positive relationship between the extent of web-traffic and firm performance in the MENA region during the 2010. We argue that higher web-traffic lowers the agency problems in firms by disseminating more information...... to stock market participants. Consequently, lower agency problems translate into better performance. Furthermore, we also show that agency reducing role of web-traffic is more pronounced in regimes where information environment is already bad. For example, our results show stronger impact of web...

Research on the Method of Traffic Organization and Optimization Based on Dynamic Traffic Flow Model

Directory of Open Access Journals (Sweden)

Shu-bin Li

2017-01-01

Full Text Available The modern transportation system is becoming sluggish by traffic jams, so much so that it can harm the economic and society in our country. One of the reasons is the surging vehicles day by day. Another reason is the shortage of the traffic supply seriously. But the most important reason is that the traffic organization and optimization hardly met the conditions of modern transport development. In this paper, the practical method of the traffic organization and optimization used in regional area is explored by the dynamic traffic network analysis method. Firstly, the operational states of the regional traffic network are obtained by simulation method based on the self-developed traffic simulation software DynaCHINA, in which the improved traffic flow simulation model was proposed in order to be more suitable for actual domestic urban transport situation. Then the appropriated optimization model and algorithm were proposed according to different optimized content and organization goals, and the traffic simulation processes more suitable to regional optimization were designed exactly. Finally, a regional network in Tai’an city was selected as an example. The simulation results show that the proposed method is effective and feasible. It can provide strong scientific and technological support for the traffic management department.

78 FR 53104 - Drawbridge Operation Regulation; Hackensack River, Kearny and Jersey City, NJ

Science.gov (United States)

2013-08-28

... relief from vehicular traffic congestion during the morning and afternoon vehicular rush hour periods due... periods to alleviate traffic congestion resulting from area roadway closures. It is expected that this change to the regulations would provide relief to vehicular traffic while continuing to meet the...

Orchestrating cytoskeleton and intracellular vesicle traffic to build functional immunological synapses.

Science.gov (United States)

Soares, Helena; Lasserre, Rémi; Alcover, Andrés

2013-11-01

Immunological synapses are specialized cell-cell contacts formed between T lymphocytes and antigen-presenting cells. They are induced upon antigen recognition and are crucial for T-cell activation and effector functions. The generation and function of immunological synapses depend on an active T-cell polarization process, which results from a finely orchestrated crosstalk between the antigen receptor signal transduction machinery, the actin and microtubule cytoskeletons, and controlled vesicle traffic. Although we understand how some of these particular events are regulated, we still lack knowledge on how these multiple cellular elements are harmonized to ensure appropriate T-cell responses. We discuss here our view on how T-cell receptor signal transduction initially commands cytoskeletal and vesicle traffic polarization, which in turn sets the immunological synapse molecular design that regulates T-cell activation. We also discuss how the human immunodeficiency virus (HIV-1) hijacks some of these processes impairing immunological synapse generation and function. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Civil Cases Proof Peculiarities of Road Traffic Accidents Damages

Directory of Open Access Journals (Sweden)

Polyakov D. N.

2012-11-01

Full Text Available The author reveals proof peculiarities in civil cases of reparation of damages harmed by road traffic accident, in relation to the determination of a respondent (debtor. In the article are analyzed the appropriate norms of the RF Civil code regulating the rules and conditions of civil liability for damage caused by using a transport facility as a source of danger

Evolution of traffic emissions in European urban areas: Milan case study

International Nuclear Information System (INIS)

Giugliano, M.; Cemin, A.; Cernuschi, S.

1992-01-01

The worldwide increasing role of mobile sources on the air quality of urban areas requires a careful evaluation of all the potential intervention policies for the reduction of traffic related emissions. The accurate description of the evolution of the source, following the renewal of the fleet with vehicles regulated by stricter emission standards, represents the basic premise for this evaluation. This paper outlines a proposed methodology for the evaluation of the trend of mobile source emissions, implemented on the calculation program EMISMOB, and reports the results obtained with its application to predict the evolution of traffic emissions in the urban area of Milan (Italy) until the year 2010

[Traffic deaths and traffic injuries: also an economic problem for society].

Science.gov (United States)

Martinoli, S

1993-02-09

Based on an epidemiological observation on traffic accidents in the Swiss canton of Ticino, 1985, the following statement is possible: In Switzerland, 900 people die every year as a result of traffic casualties. Many victims of these accidents will be disabled for life. Direct and indirect costs of traffic injuries reach 3 billion Swiss francs a year. Only a small amount (6%) is devoted to medical treatment. The biggest part is due as compensation for lost income, disability allocations and loss of productivity. Among the avoidable fatalities those caused by traffic accidents have a very undesirable effect, because they involve mainly the young. More efforts should be undertaken to lower the mortality on the road, because it changes the Swiss population pyramid in a significant manner.

Visualization of Traffic Accidents

Science.gov (United States)

Wang, Jie; Shen, Yuzhong; Khattak, Asad

2010-01-01

Traffic accidents have tremendous impact on society. Annually approximately 6.4 million vehicle accidents are reported by police in the US and nearly half of them result in catastrophic injuries. Visualizations of traffic accidents using geographic information systems (GIS) greatly facilitate handling and analysis of traffic accidents in many aspects. Environmental Systems Research Institute (ESRI), Inc. is the world leader in GIS research and development. ArcGIS, a software package developed by ESRI, has the capabilities to display events associated with a road network, such as accident locations, and pavement quality. But when event locations related to a road network are processed, the existing algorithm used by ArcGIS does not utilize all the information related to the routes of the road network and produces erroneous visualization results of event locations. This software bug causes serious problems for applications in which accurate location information is critical for emergency responses, such as traffic accidents. This paper aims to address this problem and proposes an improved method that utilizes all relevant information of traffic accidents, namely, route number, direction, and mile post, and extracts correct event locations for accurate traffic accident visualization and analysis. The proposed method generates a new shape file for traffic accidents and displays them on top of the existing road network in ArcGIS. Visualization of traffic accidents along Hampton Roads Bridge Tunnel is included to demonstrate the effectiveness of the proposed method.

Model for Predicting Traffic Signs Functional Service Life – The Republic of Croatia Case Study

Directory of Open Access Journals (Sweden)

Dario Babić

2017-06-01

Full Text Available Traffic signs are the basic elements of communication between the relevant road authorities and road users. They manage, regulate, inform and warn road users to ensure their safe movement throughout transport networks. Traffic signs must be timely visible to all traffic participants in all weather and traffic conditions in order to fulfil their function, which means they must have satisfactory retroreflection properties. This paper presents a research of the deterioration of traffic signs retroreflection. The aim of this article is to develop models that will effectively enable predicting the retroreflection of traffic signs and thus optimize the maintenance activities and replacement of road signs to increase road safety. The research included measurements of retroreflection of retroreflective material Classes I and II (white, red and blue colour and Class III (red and yellow colour. Based on the collected data from the City of Zagreb (Republic of Croatia, the authors developed the models to estimate the functional service life of certain colours and materials used to make traffic signs. Considering that the average coefficient of determination for all the models is between 0.55-0.60, they present an effective tool in the traffic sign maintenance system.

The Stability of Multi-modal Traffic Network

International Nuclear Information System (INIS)

Han Linghui; Sun Huijun; Zhu Chengjuan; Jia Bin; Wu Jianjun

2013-01-01

There is an explicit and implicit assumption in multimodal traffic equilibrium models, that is, if the equilibrium exists, then it will also occur. The assumption is very idealized; in fact, it may be shown that the quite contrary could happen, because in multimodal traffic network, especially in mixed traffic conditions the interaction among traffic modes is asymmetric and the asymmetric interaction may result in the instability of traffic system. In this paper, to study the stability of multimodal traffic system, we respectively present the travel cost function in mixed traffic conditions and in traffic network with dedicated bus lanes. Based on a day-to-day dynamical model, we study the evolution of daily route choice of travelers in multimodal traffic network using 10000 random initial values for different cases. From the results of simulation, it can be concluded that the asymmetric interaction between the cars and buses in mixed traffic conditions can lead the traffic system to instability when traffic demand is larger. We also study the effect of travelers' perception error on the stability of multimodal traffic network. Although the larger perception error can alleviate the effect of interaction between cars and buses and improve the stability of traffic system in mixed traffic conditions, the traffic system also become instable when the traffic demand is larger than a number. For all cases simulated in this study, with the same parameters, traffic system with dedicated bus lane has better stability for traffic demand than that in mixed traffic conditions. We also find that the network with dedicated bus lane has higher portion of travelers by bus than it of mixed traffic network. So it can be concluded that building dedicated bus lane can improve the stability of traffic system and attract more travelers to choose bus reducing the traffic congestion. (general)

Enrichment of Phosphatidylethanolamine in Viral Replication Compartments via Co-opting the Endosomal Rab5 Small GTPase by a Positive-Strand RNA Virus.

Directory of Open Access Journals (Sweden)

Kai Xu

2016-10-01

Full Text Available Positive-strand RNA viruses build extensive membranous replication compartments to support replication and protect the virus from antiviral responses by the host. These viruses require host factors and various lipids to form viral replication complexes (VRCs. The VRCs built by Tomato bushy stunt virus (TBSV are enriched with phosphatidylethanolamine (PE through a previously unknown pathway. To unravel the mechanism of PE enrichment within the TBSV replication compartment, in this paper, the authors demonstrate that TBSV co-opts the guanosine triphosphate (GTP-bound active form of the endosomal Rab5 small GTPase via direct interaction with the viral replication protein. Deletion of Rab5 orthologs in a yeast model host or expression of dominant negative mutants of plant Rab5 greatly decreases TBSV replication and prevents the redistribution of PE to the sites of viral replication. We also show that enrichment of PE in the viral replication compartment is assisted by actin filaments. Interestingly, the closely related Carnation Italian ringspot virus, which replicates on the boundary membrane of mitochondria, uses a similar strategy to the peroxisomal TBSV to hijack the Rab5-positive endosomes into the viral replication compartments. Altogether, usurping the GTP-Rab5-positive endosomes allows TBSV to build a PE-enriched viral replication compartment, which is needed to support peak-level replication. Thus, the Rab family of small GTPases includes critical host factors assisting VRC assembly and genesis of the viral replication compartment.

Reminder: Alcohol abuse, road traffic and safety on the site

CERN Multimedia

HSE Unit

2011-01-01

You are reminded that: - the legal blood alcohol limit in force under the Host States’ traffic regulations (0.5 g per 1000 ml of blood) also applies on the CERN site; - the consumption of alcohol is forbidden during working hours and is only tolerated in the restaurants at certain times, unless an exception is granted for special events (Operational Circular No. 8); - failure to observe these rules may result in disciplinary action by the Organization, independently of any sanctions that may be applicable pursuant to the road traffic regulations of the Host State concerned. Furthermore, the Reception and Access Control Service, the site guards and the Fire Brigade have been instructed to stop any driver in an obvious state of intoxication and to ask him/her to abandon the vehicle on the spot. In case of disagreement, they may also suggest that he/she submits to a voluntary blood alcohol level test at the CERN Medical Service or Fire Brigade. They will also stop and question any obviously intoxicate...

Right-\\0xADturn traffic volume adjustment in traffic signal warrant analysis : final report.

Science.gov (United States)

2016-05-06

This report was based on the research project, Right-Turn Traffic Volume Adjustment in : Traffic Signal Warrants, sponsored by the Nevada Department of Transportation (NDOT) : and SOLARIS. Right-turn traffic does not affect intersection performance i...

RNAi Screen in Drosophila melanogastor Identifies Regulators of Steroidogenesis and Developmental Maturation

DEFF Research Database (Denmark)

Danielsen, Erik Thomas

and duration required for juvenile-adult transition. This PhD project demonstrates the power of Drosophila genetics by taking an in vivo genome-wide RNAi screening approach to uncover genes required for the function of steroid producing tissue and developmental maturation. In total, 1909 genes were found...... to be required for the prothoracic gland function and affected the developmental timing for the juvenile-adult transition. Among the screen hits, we focused on an uncharacterized gene, sit (CG5278), which is highly expressed in the gland and is required for ecdysone production. Sit is a homolog of mammalian very...... flux of cholesterol uptake in the gland cells and affected the endosomal trafficking. Therefore this gene was suggested to be named stuck in traffic (sit). Sit’s role in cholesterol uptake was also supported by the observation that the developmental delayed phenotype from loss of sit expression...

Traffic Flow Visualization and Control

National Research Council Canada - National Science Library

Larson, Robert

1999-01-01

.... Air Force Research Laboratory. It is a video-camera-based, wide-area, traffic surveillance and detection system that provides real-time traffic information to traffic management center operators...

Intelligent Traffic Information System a Real-Time Traffic Information System on the Shiraz Bypass

Directory of Open Access Journals (Sweden)

Sodagaran Amir

2016-01-01

Full Text Available Real-time traffic information system is an Intelligent Transportation System (ITS that allows commuters to make their traveling plan better. In this regard, an intelligent and real-time traffic information system was developed based on the video detection and an image processing algorithm was applied to measure traffic-flow according to the average speed of vehicles. Then, traffic status of each pass way is broadcasted to the electronic boards installed on all decision making entrance / exit. Different levels of congestion related to the routes ahead are shown on the boards with different colors in order to assist commuters. This system was implemented on the Shiraz Dry River’s bypasses which account as vital routes to moderate traffic of city center. Experimental results are promising due to the proximity of determined traffic status by the system compared to the detection done by traffic experts. Average speed improvement is another result of using this system. This intelligent system developed and implemented in Shiraz city for the first time in Iran.s.

Stormwater Management: Calculation of Traffic Area Runoff Loads and Traffic Related Emissions

Directory of Open Access Journals (Sweden)

Maximilian Huber

2016-07-01

Full Text Available Metals such as antimony, cadmium, chromium, copper, lead, nickel, and zinc can be highly relevant pollutants in stormwater runoff from traffic areas because of their occurrence, toxicity, and non-degradability. Long-term measurements of their concentrations, the corresponding water volumes, the catchment areas, and the traffic volumes can be used to calculate specific emission loads and annual runoff loads that are necessary for mass balances. In the literature, the annual runoff loads are often specified by a distinct catchment area (e.g., g/ha. These loads were summarized and discussed in this paper for all seven metals and three types of traffic areas (highways, parking lots, and roads; 45 sites. For example, the calculated median annual runoff loads of all sites are 355 g/ha for copper, 110 g/ha for lead (only data of the 21st century, and 1960 g/ha for zinc. In addition, historical trends, annual variations, and site-specific factors were evaluated for the runoff loads. For Germany, mass balances of traffic related emissions and annual heavy metal runoff loads from highways and total traffic areas were calculated. The influences on the mass fluxes of the heavy metal emissions and the runoff pollution were discussed. However, a statistical analysis of the annual traffic related metal fluxes, in particular for different traffic area categories and land uses, is currently not possible because of a lack of monitoring data.

Synchronized flow in oversaturated city traffic.

Science.gov (United States)

Kerner, Boris S; Klenov, Sergey L; Hermanns, Gerhard; Hemmerle, Peter; Rehborn, Hubert; Schreckenberg, Michael

2013-11-01

Based on numerical simulations with a stochastic three-phase traffic flow model, we reveal that moving queues (moving jams) in oversaturated city traffic dissolve at some distance upstream of the traffic signal while transforming into synchronized flow. It is found that, as in highway traffic [Kerner, Phys. Rev. E 85, 036110 (2012)], such a jam-absorption effect in city traffic is explained by a strong driver's speed adaptation: Time headways (space gaps) between vehicles increase upstream of a moving queue (moving jam), resulting in moving queue dissolution. It turns out that at given traffic signal parameters, the stronger the speed adaptation effect, the shorter the mean distance between the signal location and the road location at which moving queues dissolve fully and oversaturated traffic consists of synchronized flow only. A comparison of the synchronized flow in city traffic found in this Brief Report with synchronized flow in highway traffic is made.

Is it traffic type, volume, or distance? Wheezing in infants living near truck and bus traffic.

Science.gov (United States)

Ryan, Patrick H; LeMasters, Grace; Biagini, Jocelyn; Bernstein, David; Grinshpun, Sergey A; Shukla, Rakesh; Wilson, Kimberly; Villareal, Manuel; Burkle, Jeff; Lockey, James

2005-08-01

Previous studies of air pollution have not examined the association between exposure to varying types, distance, and amounts of traffic and wheezing in very young infants. We sought to determine the relationship between types of traffic, traffic volume, and distance and wheezing among infants less than 1 year of age. A geographic information system and a classification scheme were developed to categorize infants enrolled in the study as living near moving truck and bus traffic (highway >50 miles per hour, >1000 trucks daily, bus traffic (bus and truck traffic had a significantly increased prevalence of wheezing (adjusted odds ratio, 2.50; 95% CI, 1.15-5.42) when compared with unexposed infants. The prevalence of wheezing among nonwhite infants was at least twice that of white infants, regardless of exposure. Infants living less than 400 m from a high volume of moving traffic, however, did not have an increased prevalence of wheezing. These results suggest that the distance from and type of traffic exposures are more significant risk factors than traffic volume for wheezing in early infancy.

A rapid appraisal of traffic policemen about auditory effects of traffic noise pollution from Ambala city

Directory of Open Access Journals (Sweden)

Abhishek Singh

2015-01-01

Full Text Available Context: Traffic policemen are at an increased risk of suffering from hazards of noise pollution because they are engaged in controlling traffic noise, particularly at heavy traffic junctions. The effect is more in this subgroup because they are continuously exposed to it. Aim: The present study was aimed at assessing the knowledge, attitude and practices of traffic policemen regarding auditory effects of traffic noise pollution in Ambala city. Settings and Design: Cross-sectional workplace survey. Materials and Methods: The present descriptive study was carried out in different traffic zones of Ambala city during April-June 2013. The study population consisted of 100 traffic policemen working in different traffic intersections of Ambala city. Statistical Analysis Used: Structured interview schedule was used to collect the data. SPSS version 17.0 was used for analysis. Interpretation of data was performed using percentages and proportions. Results: Majority (75% of the study subjects were exposed to traffic noise pollution for more than 5 years. Of the total subjects, 5% of respondents reported below-average hearing on self-assessment of hearing ability. Seventeen percent of the study population accepted that while hearing over phone they do miss some conversation. Most (98% of the traffic police did not use any personal protective equipment (PPEs like earplugs/earmuffs, and the non-availability of these PPEs (90% is the common reason for the hearing loss. Conclusions: The study concludes that traffic policemen are not much aware regarding impending auditory effects of traffic noise pollution. Duty rotation, duty scheduling and other forms of preventive modalities for exposure limitation are suggested.

Road Traffic Injuries

Institute of Scientific and Technical Information of China (English)

WANG Zheng-guo

2005-01-01

@@ As everybody knows that automobiles have been greatly changing our life. However, everything has two sides, motor vehicles have also caused a huge number of people's deaths, injuries and property damage. Traffic crashes are perhaps the number one public health problem in developed countries [1]. In the United States, pre-retirement years of life lost in traffic crashes are more than that of the two combined leading diseases: cancer and heart disease [1]. Today road traffic crash (RTC) ranks 11th in leading cause of death and accounts for 2.1% of all deaths globally.

Multi-layered nanoparticles for penetrating the endosome and nuclear membrane via a step-wise membrane fusion process.

Science.gov (United States)

Akita, Hidetaka; Kudo, Asako; Minoura, Arisa; Yamaguti, Masaya; Khalil, Ikramy A; Moriguchi, Rumiko; Masuda, Tomoya; Danev, Radostin; Nagayama, Kuniaki; Kogure, Kentaro; Harashima, Hideyoshi

2009-05-01

Efficient targeting of DNA to the nucleus is a prerequisite for effective gene therapy. The gene-delivery vehicle must penetrate through the plasma membrane, and the DNA-impermeable double-membraned nuclear envelope, and deposit its DNA cargo in a form ready for transcription. Here we introduce a concept for overcoming intracellular membrane barriers that involves step-wise membrane fusion. To achieve this, a nanotechnology was developed that creates a multi-layered nanoparticle, which we refer to as a Tetra-lamellar Multi-functional Envelope-type Nano Device (T-MEND). The critical structural elements of the T-MEND are a DNA-polycation condensed core coated with two nuclear membrane-fusogenic inner envelopes and two endosome-fusogenic outer envelopes, which are shed in stepwise fashion. A double-lamellar membrane structure is required for nuclear delivery via the stepwise fusion of double layered nuclear membrane structure. Intracellular membrane fusions to endosomes and nuclear membranes were verified by spectral imaging of fluorescence resonance energy transfer (FRET) between donor and acceptor fluorophores that had been dually labeled on the liposome surface. Coating the core with the minimum number of nucleus-fusogenic lipid envelopes (i.e., 2) is essential to facilitate transcription. As a result, the T-MEND achieves dramatic levels of transgene expression in non-dividing cells.

Packet traffic features of IPv6 and IPv4 protocol traffic

OpenAIRE

ÇİFLİKLİ, Cebrail; GEZER, Ali; ÖZŞAHİN, Abdullah Tuncay

2012-01-01

Nowadays, the IPv6 protocol is in a transition phase in operational networks. The ratio of its traffic volume is increasing day by day. The many provided facilities for IPv6 connection increasethe total IPv6 traffic load. IPv6-over-IPv4 tunnels, pilot programsto provide IPv6 connections, IPv6/IPv4 dual stack operating systems,and free IPv6 tunnel brokers cause the IPv6 protocol to expand quickly. For efficient resource utilization, the characteristics of network traffic should be determ...

Drivers' phone use at red traffic lights: a roadside observation study comparing calls and visual-manual interactions.

Science.gov (United States)

Huth, Véronique; Sanchez, Yann; Brusque, Corinne

2015-01-01

Phone use while driving has become one of the priority issues in road safety, given that it may lead to decreased situation awareness and deteriorated driving performance. It has been suggested that drivers can regulate their exposure to secondary tasks and seek for compatibility of phone use and driving. Phone use strategies include the choice of driving situations with low demands and interruptions of the interaction when the context changes. Traffic light situations at urban intersections imply both a temptation to use the phone while waiting at the red traffic light and a potential threat due to the incompatibility of phone use and driving when the traffic light turns green. These two situations were targeted in a roadside observation study, with the aim to investigate the existence of a phone use strategy at the red traffic light and to test its effectiveness. N=124 phone users and a corresponding control group of non-users were observed. Strategic phone use behaviour was detected for visual-manual interactions, which are more likely to be initiated at the red traffic light and tend to be stopped before the vehicle moves off, while calls are less likely to be limited to the red traffic light situation. As an indicator of impaired situation awareness, delayed start was associated to phone use and in particular to visual-manual interactions, whether phone use was interrupted before moving off or not. Traffic light situations do not seem to allow effective application of phone use strategies, although drivers attempt to do so for the most demanding phone use mode. The underlying factors of phone use need to be studied so as to reduce the temptation of phone use and facilitate exposure regulation strategies. Copyright © 2014. Published by Elsevier Ltd.

Modulation of PML protein expression regulates JCV infection

International Nuclear Information System (INIS)

Gasparovic, Megan L.; Maginnis, Melissa S.; O'Hara, Bethany A.; Dugan, Aisling S.; Atwood, Walter J.

2009-01-01

JC virus (JCV) is a human polyomavirus that infects the majority of the human population worldwide. It is responsible for the fatal demyelinating disease Progressive Multifocal Leukoencephalopathy. JCV binds to cells using the serotonin receptor 5-HT 2A R and α(2-6)- or α(2-3)-linked sialic acid. It enters cells using clathrin-dependent endocytosis and traffics to the early endosome and possibly to the endoplasmic reticulum. Viral DNA is then delivered to the nucleus where transcription, replication, and assembly of progeny occur. We found that the early regulatory protein large T antigen accumulates in microdomains in the nucleus adjacent to ND-10 or PML domains. This observation prompted us to explore the role of these domains in JCV infection. We found that a reduction of nuclear PML enhanced virus infection and that an increase in nuclear PML reduced infection. Infection with JCV did not directly modulate nuclear levels of PML but our data indicate that a host response involving interferon beta is likely to restrict virus infection by increasing nuclear PML.

Dynamic traffic management and air quality; Dynamisch verkeersmanagement en luchtkwaliteit

Energy Technology Data Exchange (ETDEWEB)

NONE

2010-06-15

Traffic flow can be improved by means of dynamic routing and regulation as a result of which traffic might choose a different route. Therefore, in this report also attention is paid to the effects on other road sections, such as improvement of the air quality. Calculations were carried out for the cases Southeast Amsterdam, Alkmaar and De Bilt. The detailed impacts on traffic and on air quality are available in the annex. All the reported air quality impacts are calculated using CAR II, version 8.0 for the year 2010. [Dutch] Doorstroming van het verkeer kan worden bevorderd door middel van dynamisch routeren en doseren. Het verkeer zal door deze maatregel een andere route gaan kiezen. Daarom besteedt de publicatie ook aandacht aan de effecten op de overige wegvakken, zoals verbetering van de luchtkwaliteit. Voor deze publikatie zijn berekeningen uitgevoerd voor de cases Amsterdam-zuidoost, Alkmaar en De Bilt. De gedetailleerde verkeerseffecten en effecten op luchtkwaliteit staan in de gratis beschikbare bijlage. Alle in deze rapportage opgenomen effecten op luchtkwaliteit zijn berekend met CAR II, versie 8.0 voor het jaar 2010.

Urban Road Traffic Simulation Techniques

Directory of Open Access Journals (Sweden)

Ana Maria Nicoleta Mocofan

2011-09-01

Full Text Available For achieving a reliable traffic control system it is necessary to first establish a network parameter evaluation system and also a simulation system for the traffic lights plan. In 40 years of history, the computer aided traffic simulation has developed from a small research group to a large scale technology for traffic systems planning and development. In the following thesis, a presentation of the main modeling and simulation road traffic applications will be provided, along with their utility, as well as the practical application of one of the models in a case study.

Traffic Flow Management Wrap-Up

Science.gov (United States)

Grabbe, Shon

2011-01-01

Traffic Flow Management involves the scheduling and routing of air traffic subject to airport and airspace capacity constraints, and the efficient use of available airspace. Significant challenges in this area include: (1) weather integration and forecasting, (2) accounting for user preferences in the Traffic Flow Management decision making process, and (3) understanding and mitigating the environmental impacts of air traffic on the environment. To address these challenges, researchers in the Traffic Flow Management area are developing modeling, simulation and optimization techniques to route and schedule air traffic flights and flows while accommodating user preferences, accounting for system uncertainties and considering the environmental impacts of aviation. This presentation will highlight some of the major challenges facing researchers in this domain, while also showcasing recent innovations designed to address these challenges.

Monitoring road traffic congestion using a macroscopic traffic model and a statistical monitoring scheme

KAUST Repository

Zeroual, Abdelhafid; Harrou, Fouzi; Sun, Ying; Messai, Nadhir

2017-01-01

Monitoring vehicle traffic flow plays a central role in enhancing traffic management, transportation safety and cost savings. In this paper, we propose an innovative approach for detection of traffic congestion. Specifically, we combine the flexibility and simplicity of a piecewise switched linear (PWSL) macroscopic traffic model and the greater capacity of the exponentially-weighted moving average (EWMA) monitoring chart. Macroscopic models, which have few, easily calibrated parameters, are employed to describe a free traffic flow at the macroscopic level. Then, we apply the EWMA monitoring chart to the uncorrelated residuals obtained from the constructed PWSL model to detect congested situations. In this strategy, wavelet-based multiscale filtering of data has been used before the application of the EWMA scheme to improve further the robustness of this method to measurement noise and reduce the false alarms due to modeling errors. The performance of the PWSL-EWMA approach is successfully tested on traffic data from the three lane highway portion of the Interstate 210 (I-210) highway of the west of California and the four lane highway portion of the State Route 60 (SR60) highway from the east of California, provided by the Caltrans Performance Measurement System (PeMS). Results show the ability of the PWSL-EWMA approach to monitor vehicle traffic, confirming the promising application of this statistical tool to the supervision of traffic flow congestion.

Monitoring road traffic congestion using a macroscopic traffic model and a statistical monitoring scheme

KAUST Repository

Zeroual, Abdelhafid

2017-08-19

Monitoring vehicle traffic flow plays a central role in enhancing traffic management, transportation safety and cost savings. In this paper, we propose an innovative approach for detection of traffic congestion. Specifically, we combine the flexibility and simplicity of a piecewise switched linear (PWSL) macroscopic traffic model and the greater capacity of the exponentially-weighted moving average (EWMA) monitoring chart. Macroscopic models, which have few, easily calibrated parameters, are employed to describe a free traffic flow at the macroscopic level. Then, we apply the EWMA monitoring chart to the uncorrelated residuals obtained from the constructed PWSL model to detect congested situations. In this strategy, wavelet-based multiscale filtering of data has been used before the application of the EWMA scheme to improve further the robustness of this method to measurement noise and reduce the false alarms due to modeling errors. The performance of the PWSL-EWMA approach is successfully tested on traffic data from the three lane highway portion of the Interstate 210 (I-210) highway of the west of California and the four lane highway portion of the State Route 60 (SR60) highway from the east of California, provided by the Caltrans Performance Measurement System (PeMS). Results show the ability of the PWSL-EWMA approach to monitor vehicle traffic, confirming the promising application of this statistical tool to the supervision of traffic flow congestion.

A Public Health Perspective of Road Traffic Accidents

Science.gov (United States)

Gopalakrishnan, S.

2012-01-01

Road traffic accidents (RTAs) have emerged as an important public health issue which needs to be tackled by a multi-disciplinary approach. The trend in RTA injuries and death is becoming alarming in countries like India. The number of fatal and disabling road accident happening is increasing day by day and is a real public health challenge for all the concerned agencies to prevent it. The approach to implement the rules and regulations available to prevent road accidents is often ineffective and half-hearted. Awareness creation, strict implementation of traffic rules, and scientific engineering measures are the need of the hour to prevent this public health catastrophe. This article is intended to create awareness among the health professionals about the various modalities available to prevent road accidents and also to inculcate a sense of responsibility toward spreading the message of road safety as a good citizen of our country. PMID:24479025

Comparisons of Traffic Collisions between Expressways and Rural Roads in Truck Drivers

Directory of Open Access Journals (Sweden)

Sangbok Lee

2016-03-01

Conclusion: This study can be used as a guideline and a base line to develop a plan of action to prevent traffic accidents. It can also help to prepare formal regulations about a truck driver's vehicle maintenance and driving attitude for a precaution on road accidents.

Pedestrian Friendly Traffic Signal Control.

Science.gov (United States)

2016-01-01

This project continues research aimed at real-time detection and use of pedestrian : traffic flow information to enhance adaptive traffic signal control in urban areas : where pedestrian traffic is substantial and must be given appropriate attention ...

Vps1 in the late endosome-to-vacuole traffic

Indian Academy of Sciences (India)

2013-01-13

Jan 13, 2013 ... the plasma membrane lipids and follows the endocytic inter- nalization pathways ... plasm and found also as punctuate structures (figure 1A). Our explanation ..... 1992 Morphological classification of the yeast vacuolar protein.

Costs of traffic injuries

DEFF Research Database (Denmark)

Kruse, Marie

2015-01-01

assessed using Danish national healthcare registers. Productivity costs were computed using duration analysis (Cox regression models). In a subanalysis, cost per severe traffic injury was computed for the 12 995 individuals that experienced a severe injury. RESULTS: The socioeconomic cost of a traffic...... injury was €1406 (2009 price level) in the first year, and €8950 over a 10-year period. Per 100 000 population, the 10-year cost was €6 565 668. A severe traffic injury costs €4969 per person in the first year, and €4 006 685 per 100 000 population over a 10-year period. Victims of traffic injuries...

Traffic simulation for mixed traffic systems | Mbam | Global Journal of ...

African Journals Online (AJOL)

Traffic problem is classified into single and mixed, especially in most developing countries, where motorbikes are used as the most popular transportation system. The aim of this paper is to introduce the motorbike symbol into the traffic light control system to separate cars/lorries indicator from that of motorbike. This is likely ...

Traffic information computing platform for big data

Energy Technology Data Exchange (ETDEWEB)

Duan, Zongtao, E-mail: ztduan@chd.edu.cn; Li, Ying, E-mail: ztduan@chd.edu.cn; Zheng, Xibin, E-mail: ztduan@chd.edu.cn; Liu, Yan, E-mail: ztduan@chd.edu.cn; Dai, Jiting, E-mail: ztduan@chd.edu.cn; Kang, Jun, E-mail: ztduan@chd.edu.cn [Chang' an University School of Information Engineering, Xi' an, China and Shaanxi Engineering and Technical Research Center for Road and Traffic Detection, Xi' an (China)

2014-10-06

Big data environment create data conditions for improving the quality of traffic information service. The target of this article is to construct a traffic information computing platform for big data environment. Through in-depth analysis the connotation and technology characteristics of big data and traffic information service, a distributed traffic atomic information computing platform architecture is proposed. Under the big data environment, this type of traffic atomic information computing architecture helps to guarantee the traffic safety and efficient operation, more intelligent and personalized traffic information service can be used for the traffic information users.

Traffic information computing platform for big data

International Nuclear Information System (INIS)

Duan, Zongtao; Li, Ying; Zheng, Xibin; Liu, Yan; Dai, Jiting; Kang, Jun

2014-01-01

Big data environment create data conditions for improving the quality of traffic information service. The target of this article is to construct a traffic information computing platform for big data environment. Through in-depth analysis the connotation and technology characteristics of big data and traffic information service, a distributed traffic atomic information computing platform architecture is proposed. Under the big data environment, this type of traffic atomic information computing architecture helps to guarantee the traffic safety and efficient operation, more intelligent and personalized traffic information service can be used for the traffic information users

Traffic forecasts ignoring induced demand

DEFF Research Database (Denmark)

Næss, Petter; Nicolaisen, Morten Skou; Strand, Arvid

2012-01-01

the model calculations included only a part of the induced traffic, the difference in cost-benefit results compared to the model excluding all induced traffic was substantial. The results show lower travel time savings, more adverse environmental impacts and a considerably lower benefitcost ratio when...... induced traffic is partly accounted for than when it is ignored. By exaggerating the economic benefits of road capacity increase and underestimating its negative effects, omission of induced traffic can result in over-allocation of public money on road construction and correspondingly less focus on other...... performance of a proposed road project in Copenhagen with and without short-term induced traffic included in the transport model. The available transport model was not able to include long-term induced traffic resulting from changes in land use and in the level of service of public transport. Even though...

Traffic intensity monitoring using multiple object detection with traffic surveillance cameras

Science.gov (United States)

Hamdan, H. G. Muhammad; Khalifah, O. O.

2017-11-01

Object detection and tracking is a field of research that has many applications in the current generation with increasing number of cameras on the streets and lower cost for Internet of Things(IoT). In this paper, a traffic intensity monitoring system is implemented based on the Macroscopic Urban Traffic model is proposed using computer vision as its source. The input of this program is extracted from a traffic surveillance camera which has another program running a neural network classification which can identify and differentiate the vehicle type is implanted. The neural network toolbox is trained with positive and negative input to increase accuracy. The accuracy of the program is compared to other related works done and the trends of the traffic intensity from a road is also calculated. relevant articles in literature searches, great care should be taken in constructing both. Lastly the limitation and the future work is concluded.

Intelligent Traffic Light Based on PLC Control

Science.gov (United States)

Mei, Lin; Zhang, Lijian; Wang, Lingling

2017-11-01

The traditional traffic light system with a fixed control mode and single control function is contradicted with the current traffic section. The traditional one has been unable to meet the functional requirements of the existing flexible traffic control system. This paper research and develop an intelligent traffic light called PLC control system. It uses PLC as control core, using a sensor module for receiving real-time information of vehicles, traffic control mode for information to select the traffic lights. Of which control mode is flexible and changeable, and it also set the countdown reminder to improve the effectiveness of traffic lights, which can realize the goal of intelligent traffic diversion, intelligent traffic diversion.

ITSA: Internet Traffic Statistics Archive

NARCIS (Netherlands)

Hoogesteger, Martijn; de Oliveira Schmidt, R.; Pras, Aiko

Motivated by the fact that comprehensive and long term Internet traffic measurements can be hard to obtain, we have proposed and developed the Internet Traffic Statistics Archive (ITSA). Since 2013, ITSA concentrates reports on high-level statistics of Internet traffic worldwide. Examples of

Fuzzy Multiobjective Traffic Light Signal Optimization

Directory of Open Access Journals (Sweden)

N. Shahsavari Pour

2013-01-01

Full Text Available Traffic congestion is a major concern for many cities throughout the world. In a general traffic light controller, the traffic lights change at a constant cycle time. Hence it does not provide an optimal solution. Many traffic light controllers in current use are based on the “time-of-the-day” scheme, which use a limited number of predetermined traffic light patterns and implement these patterns depending upon the time of the day. These automated systems do not provide an optimal control for fluctuating traffic volumes. In this paper, the fuzzy traffic light controller is used to optimize the control of fluctuating traffic volumes such as oversaturated or unusual load conditions. The problem is solved by genetic algorithm, and a new defuzzification method is introduced. The performance of the new defuzzification method (NDM is compared with the centroid point defuzzification method (CPDM by using ANOVA. Finally, an illustrative example is presented to show the competency of proposed algorithm.

Traffic Management as a Service: The Traffic Flow Pattern Classification Problem

Directory of Open Access Journals (Sweden)

Carlos T. Calafate

2015-01-01

Full Text Available Intelligent Transportation System (ITS technologies can be implemented to reduce both fuel consumption and the associated emission of greenhouse gases. However, such systems require intelligent and effective route planning solutions to reduce travel time and promote stable traveling speeds. To achieve such goal these systems should account for both estimated and real-time traffic congestion states, but obtaining reliable traffic congestion estimations for all the streets/avenues in a city for the different times of the day, for every day in a year, is a complex task. Modeling such a tremendous amount of data can be time-consuming and, additionally, centralized computation of optimal routes based on such time-dependencies has very high data processing requirements. In this paper we approach this problem through a heuristic to considerably reduce the modeling effort while maintaining the benefits of time-dependent traffic congestion modeling. In particular, we propose grouping streets by taking into account real traces describing the daily traffic pattern. The effectiveness of this heuristic is assessed for the city of Valencia, Spain, and the results obtained show that it is possible to reduce the required number of daily traffic flow patterns by a factor of 4210 while maintaining the essence of time-dependent modeling requirements.

Lagrangian Multi-Class Traffic State Estimation

NARCIS (Netherlands)

Yuan, Y.

2013-01-01

Road traffic is important to everybody in the world. People travel and commute everyday. For those who travel by cars (or other types of road vehicles), traffic congestion is a daily experience. One essential goal of traffic researchers is to reduce traffic congestion and to improve the whole

Traffic law enforcement by non-police bodies. The "Escape" Project, Deliverable 4. Project funded by the European Commission under the Transport RTD Programme of the 4th Framework Programme.

NARCIS (Netherlands)

Heidstra, J. Goldenbeld, C. Gelau, C. Mäkinen, T. Jayet, M.-C. & Evers, C.

2000-01-01

To ensure some minimal standard for traffic system operation and safety, a system of traffic laws and regulations is necessary. Enforcement of these traffic laws is believed to influence driving behaviour through a mechanism of deterrence: the threat of legal punishment should convince road users to

Traffic analysis toolbox volume XI : weather and traffic analysis, modeling and simulation.

Science.gov (United States)

2010-12-01

This document presents a weather module for the traffic analysis tools program. It provides traffic engineers, transportation modelers and decisions makers with a guide that can incorporate weather impacts into transportation system analysis and mode...

State Traffic Safety Information

Data.gov (United States)

Department of Transportation — The State Traffic Safety Information (STSI) portal is part of the larger Fatality Analysis Reporting System (FARS) Encyclopedia. STSI provides state-by-state traffic...

Dynamic control of traffic lights

NARCIS (Netherlands)

Haijema, Rene; Hendrix, Eligius M.T.; Wal, van der Jan

2017-01-01

Traffic lights are put in place to dynamically change priority between traffic participants. Commonly, the duration of green intervals and the grouping, and ordering in which traffic flows are served are pre-fixed. In this chapter, the problem of minimizing vehicle delay at isolated intersections is

A Public Health Perspective of Road Traffic Accidents

Directory of Open Access Journals (Sweden)

S Gopalakrishnan

2012-01-01

Full Text Available Road traffic accidents (RTAs have emerged as an important public health issue which needs to be tackled by a multi-disciplinary approach. The trend in RTA injuries and death is becoming alarming in countries like India. The number of fatal and disabling road accident happening is increasing day by day and is a real public health challenge for all the concerned agencies to prevent it. The approach to implement the rules and regulations available to prevent road accidents is often ineffective and half-hearted. Awareness creation, strict implementation of traffic rules, and scientific engineering measures are the need of the hour to prevent this public health catastrophe. This article is intended to create awareness among the health professionals about the various modalities available to prevent road accidents and also to inculcate a sense of responsibility toward spreading the message of road safety as a good citizen of our country.

Consensus-Based Cooperative Control Based on Pollution Sensing and Traffic Information for Urban Traffic Networks.

Science.gov (United States)

Artuñedo, Antonio; Del Toro, Raúl M; Haber, Rodolfo E

2017-04-26

Nowadays many studies are being conducted to develop solutions for improving the performance of urban traffic networks. One of the main challenges is the necessary cooperation among different entities such as vehicles or infrastructure systems and how to exploit the information available through networks of sensors deployed as infrastructures for smart cities. In this work an algorithm for cooperative control of urban subsystems is proposed to provide a solution for mobility problems in cities. The interconnected traffic lights controller ( TLC ) network adapts traffic lights cycles, based on traffic and air pollution sensory information, in order to improve the performance of urban traffic networks. The presence of air pollution in cities is not only caused by road traffic but there are other pollution sources that contribute to increase or decrease the pollution level. Due to the distributed and heterogeneous nature of the different components involved, a system of systems engineering approach is applied to design a consensus-based control algorithm. The designed control strategy contains a consensus-based component that uses the information shared in the network for reaching a consensus in the state of TLC network components. Discrete event systems specification is applied for modelling and simulation. The proposed solution is assessed by simulation studies with very promising results to deal with simultaneous responses to both pollution levels and traffic flows in urban traffic networks.

Consensus-Based Cooperative Control Based on Pollution Sensing and Traffic Information for Urban Traffic Networks

Directory of Open Access Journals (Sweden)

Antonio Artuñedo

2017-04-01

Full Text Available Nowadays many studies are being conducted to develop solutions for improving the performance of urban traffic networks. One of the main challenges is the necessary cooperation among different entities such as vehicles or infrastructure systems and how to exploit the information available through networks of sensors deployed as infrastructures for smart cities. In this work an algorithm for cooperative control of urban subsystems is proposed to provide a solution for mobility problems in cities. The interconnected traffic lights controller (TLC network adapts traffic lights cycles, based on traffic and air pollution sensory information, in order to improve the performance of urban traffic networks. The presence of air pollution in cities is not only caused by road traffic but there are other pollution sources that contribute to increase or decrease the pollution level. Due to the distributed and heterogeneous nature of the different components involved, a system of systems engineering approach is applied to design a consensus-based control algorithm. The designed control strategy contains a consensus-based component that uses the information shared in the network for reaching a consensus in the state of TLC network components. Discrete event systems specification is applied for modelling and simulation. The proposed solution is assessed by simulation studies with very promising results to deal with simultaneous responses to both pollution levels and traffic flows in urban traffic networks.

Dynamic Traffic Congestion Simulation and Dissipation Control Based on Traffic Flow Theory Model and Neural Network Data Calibration Algorithm

Directory of Open Access Journals (Sweden)

Li Wang

2017-01-01

Full Text Available Traffic congestion is a common problem in many countries, especially in big cities. At present, China’s urban road traffic accidents occur frequently, the occurrence frequency is high, the accident causes traffic congestion, and accidents cause traffic congestion and vice versa. The occurrence of traffic accidents usually leads to the reduction of road traffic capacity and the formation of traffic bottlenecks, causing the traffic congestion. In this paper, the formation and propagation of traffic congestion are simulated by using the improved medium traffic model, and the control strategy of congestion dissipation is studied. From the point of view of quantitative traffic congestion, the paper provides the fact that the simulation platform of urban traffic integration is constructed, and a feasible data analysis, learning, and parameter calibration method based on RBF neural network is proposed, which is used to determine the corresponding decision support system. The simulation results prove that the control strategy proposed in this paper is effective and feasible. According to the temporal and spatial evolution of the paper, we can see that the network has been improved on the whole.

Framework for Traffic Congestion Prediction

NARCIS (Netherlands)

Zaki, J.F.W.; Ali-Eldin, A.M.T.; Hussein, S.E.; Saraya, S.F.; Areed, F.F.

2016-01-01

Traffic Congestion is a complex dilemma facing most major cities. It has undergone a lot of research since the early 80s in an attempt to predict traffic in the short-term. Recently, Intelligent Transportation Systems (ITS) became an integral part of traffic research which helped in modeling and

Traffic sign detection and analysis

DEFF Research Database (Denmark)

Møgelmose, Andreas; Trivedi, Mohan M.; Moeslund, Thomas B.

2012-01-01

Traffic sign recognition (TSR) is a research field that has seen much activity in the recent decade. This paper introduces the problem and presents 4 recent papers on traffic sign detection and 4 recent papers on traffic sign classification. It attempts to extract recent trends in the field...

Forecasting Multivariate Road Traffic Flows Using Bayesian Dynamic Graphical Models, Splines and Other Traffic Variables

NARCIS (Netherlands)

Anacleto, Osvaldo; Queen, Catriona; Albers, Casper J.

Traffic flow data are routinely collected for many networks worldwide. These invariably large data sets can be used as part of a traffic management system, for which good traffic flow forecasting models are crucial. The linear multiregression dynamic model (LMDM) has been shown to be promising for

Air traffic control activity increases attention capacity in air traffic controllers.

Science.gov (United States)

Ribas, Valdenilson Ribeiro; Martins, Hugo André de Lima; Amorim, Gutemberg Guerra; Ribas, Renata de Melo Guerra; de Almeida, Cláudia Ângela Vilela; Ribas, Valéria Ribeiro; de Vasconcelos, Carlos Augusto Carvalho; Lima, Murilo Duarte Costa; Sougey, Everton Botelho; de Castro, Raul Manhães

2010-01-01

Air traffic controllers simultaneously develop complex and multiple tasks in the course of their activities. In this context, concern is raised over the high level of attention needed by these professionals which can ultimately be affected by stress and fatigue. The objective of this study was to assess attention level in air traffic controllers (ATCo). 45 flight protection professionals were evaluated, comprising 30 ATCo, subdivided into ATCo with ten or more years in the profession (ATCo≥10, n=15) and ATCo with less than ten years in the profession (ATCo air traffic control activity after ten years may be associated with a high level of attention.

Traffic and Granular Flow ’07

CERN Document Server

Chevoir, François; Gondret, Philippe; Lassarre, Sylvain; Lebacque, Jean-Patrick; Schreckenberg, Michael

2009-01-01

This book covers several research fields, all of which deal with transport. Three main topics are treated: road traffic, granular matter, and biological transport. Different points of view, i.e. modelling, simulations, experiments, and phenomenological observations, are considered. Sub-topics include: highway or urban vehicular traffic (dynamics of traffic, macro/micro modelling, measurements, data analysis, security issues, psychological issues), pedestrian traffic, animal traffic (e.g. social insects), collective motion in biological systems (molecular motors...), granular flow (dense flows, intermittent flows, solid/liquid transition, jamming, force networks, fluid and solid friction), networks (biological networks, urban traffic, the internet, vulnerability of networks, optimal transport networks) and cellular automata applied to the various aforementioned fields.

32 CFR 634.32 - Traffic violation reports.

Science.gov (United States)

2010-07-01

... 32 National Defense 4 2010-07-01 2010-07-01 true Traffic violation reports. 634.32 Section 634.32 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY (CONTINUED) LAW ENFORCEMENT AND CRIMINAL INVESTIGATIONS MOTOR VEHICLE TRAFFIC SUPERVISION Traffic Supervision § 634.32 Traffic violation reports. (a) Most traffic violations...

Probabilistic description of traffic flow

International Nuclear Information System (INIS)

Mahnke, R.; Kaupuzs, J.; Lubashevsky, I.

2005-01-01

A stochastic description of traffic flow, called probabilistic traffic flow theory, is developed. The general master equation is applied to relatively simple models to describe the formation and dissolution of traffic congestions. Our approach is mainly based on spatially homogeneous systems like periodically closed circular rings without on- and off-ramps. We consider a stochastic one-step process of growth or shrinkage of a car cluster (jam). As generalization we discuss the coexistence of several car clusters of different sizes. The basic problem is to find a physically motivated ansatz for the transition rates of the attachment and detachment of individual cars to a car cluster consistent with the empirical observations in real traffic. The emphasis is put on the analogy with first-order phase transitions and nucleation phenomena in physical systems like supersaturated vapour. The results are summarized in the flux-density relation, the so-called fundamental diagram of traffic flow, and compared with empirical data. Different regimes of traffic flow are discussed: free flow, congested mode as stop-and-go regime, and heavy viscous traffic. The traffic breakdown is studied based on the master equation as well as the Fokker-Planck approximation to calculate mean first passage times or escape rates. Generalizations are developed to allow for on-ramp effects. The calculated flux-density relation and characteristic breakdown times coincide with empirical data measured on highways. Finally, a brief summary of the stochastic cellular automata approach is given

Traffic Flow Prediction Using MI Algorithm and Considering Noisy and Data Loss Conditions: An Application to Minnesota Traffic Flow Prediction

Directory of Open Access Journals (Sweden)

Seyed Hadi Hosseini

2014-10-01

Full Text Available Traffic flow forecasting is useful for controlling traffic flow, traffic lights, and travel times. This study uses a multi-layer perceptron neural network and the mutual information (MI technique to forecast traffic flow and compares the prediction results with conventional traffic flow forecasting methods. The MI method is used to calculate the interdependency of historical traffic data and future traffic flow. In numerical case studies, the proposed traffic flow forecasting method was tested against data loss, changes in weather conditions, traffic congestion, and accidents. The outcomes were highly acceptable for all cases and showed the robustness of the proposed flow forecasting method.

Mechanisms of isoform-specific Na/K pump regulation by short- and long-term adrenergic activation in rat ventricular myocytes.

Science.gov (United States)

Yin, Jian; Guo, Hui-Cai; Yu, Ding; Wang, Hui-Ci; Li, Jun-Xia; Wang, Yong-Li

2014-01-01

Many stressful conditions, including cardiovascular diseases, induce long-term elevations in circulating catecholamines, thereby leading to changes of the Na/K pump and thus affecting myocardial functions. However, only short-term adrenergic regulation of the Na/K pump has been reported. The present study is the first investigation of long-term adrenergic regulation of the Na/K pump and the potential mechanism. After acutely isolated Sprague-Dawley rat myocytes were incubated with noradrenaline or isoprenaline for 24 h, Na/K pump high- (IPH) and low-affinity current (IPL), α-isoform mRNA, and α-isoform protein were examined using patch-clamp, RT-PCR, and Western blotting techniques, respectively. After the short-term incubation, isoprenaline reduced the IPL through a PKA-dependent pathway that involves α1-isoform translocation from the membrane to early endosomes, and noradrenaline increased the IPH through a PKC-dependent pathway that involves α2-isoform translocation from late endosomes to the membrane. After long-term incubation, isoprenaline increased the IPL, α1-isoform mRNA, and α1-isoform protein, and noradrenaline reduced the IPH, α2-isoform mRNA, and α1-isoform protein through a PKA-or PKC-dependent pathway, respectively. These results suggest that long-term adrenergic Na/K pump regulation is isoform-specific and negatively feeds back on the short-term response. Furthermore, long-term regulation involves transcription and translation of the respective α-isoform, whereas short-term regulation involves the translocation of the available α-isoform to the plasma membrane. © 2014 S. Karger AG, Basel.

Mechanisms of Isoform-Specific Na/K Pump Regulation by Short- and Long-Term Adrenergic Activation in Rat Ventricular Myocytes

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Jian Yin

2014-05-01

Full Text Available Background: Many stressful conditions, including cardiovascular diseases, induce long-term elevations in circulating catecholamines, thereby leading to changes of the Na/K pump and thus affecting myocardial functions. However, only short-term adrenergic regulation of the Na/K pump has been reported. The present study is the first investigation of long-term adrenergic regulation of the Na/K pump and the potential mechanism. Methods: After acutely isolated Sprague-Dawley rat myocytes were incubated with noradrenaline or isoprenaline for 24 h, Na/K pump high- (IPH and low-affinity current (IPL, α-isoform mRNA, and α-isoform protein were examined using patch-clamp, RT-PCR, and Western blotting techniques, respectively. Results: After the short-term incubation, isoprenaline reduced the IPL through a PKA-dependent pathway that involves α1-isoform translocation from the membrane to early endosomes, and noradrenaline increased the IPH through a PKC-dependent pathway that involves α2-isoform translocation from late endosomes to the membrane. After long-term incubation, isoprenaline increased the IPL, α1-isoform mRNA, and α1-isoform protein, and noradrenaline reduced the IPH, α2-isoform mRNA, and α1-isoform protein through a PKA-or PKC-dependent pathway, respectively. Conclusions: These results suggest that long-term adrenergic Na/K pump regulation is isoform-specific and negatively feeds back on the short-term response. Furthermore, long-term regulation involves transcription and translation of the respective α-isoform, whereas short-term regulation involves the translocation of the available α-isoform to the plasma membrane.

Framework for Traffic Congestion Management

Directory of Open Access Journals (Sweden)

Mahmud Hassan TALUKDAR

2013-06-01

Full Text Available Traffic Congestion is one of many serious global problems in all great cities resulted from rapid urbanization which always exert negative externalities upon society. The solution of traffic congestion is highly geocentric and due to its heterogeneous nature, curbing congestion is one of the hard tasks for transport planners. It is not possible to suggest unique traffic congestion management framework which could be absolutely applied for every great cities. Conversely, it is quite feasible to develop a framework which could be used with or without minor adjustment to deal with congestion problem. So, the main aim of this paper is to prepare a traffic congestion mitigation framework which will be useful for urban planners, transport planners, civil engineers, transport policy makers, congestion management researchers who are directly or indirectly involved or willing to involve in the task of traffic congestion management. Literature review is the main source of information of this study. In this paper, firstly, traffic congestion is defined on the theoretical point of view and then the causes of traffic congestion are briefly described. After describing the causes, common management measures, using world- wide, are described and framework for supply side and demand side congestion management measures are prepared.

The subapical compartment : a traffic center in membrane polarity development

NARCIS (Netherlands)

Hoekstra, D; Tyteca, D; van IJzendoorn, SCD

2004-01-01

Spatially separated apical and basolateral plasma membrane domains that have distinct functions and molecular compositions are a characteristic feature of epithelial cell polarity. The subapical compartment (SAC), also known as the common endosome (CE), where endocytic pathways from both surfaces

Emergent traffic jams

International Nuclear Information System (INIS)

Nagel, K.; Paczuski, M.

1995-01-01

We study a single-lane traffic model that is based on human driving behavior. The outflow from a traffic jam self-organizes to a critical state of maximum throughput. Small perturbations of the outflow far downstream create emergent traffic jams with a power law distribution P(t)∼t -3/2 of lifetimes t. On varying the vehicle density in a closed system, this critical state separates lamellar and jammed regimes and exhibits 1/f noise in the power spectrum. Using random walk arguments, in conjunction with a cascade equation, we develop a phenomenological theory that predicts the critical exponents for this transition and explains the self-organizing behavior. These predictions are consistent with all of our numerical results

Emergent traffic jams

Science.gov (United States)

Nagel, Kai; Paczuski, Maya

1995-04-01

We study a single-lane traffic model that is based on human driving behavior. The outflow from a traffic jam self-organizes to a critical state of maximum throughput. Small perturbations of the outflow far downstream create emergent traffic jams with a power law distribution P(t)~t-3/2 of lifetimes t. On varying the vehicle density in a closed system, this critical state separates lamellar and jammed regimes and exhibits 1/f noise in the power spectrum. Using random walk arguments, in conjunction with a cascade equation, we develop a phenomenological theory that predicts the critical exponents for this transition and explains the self-organizing behavior. These predictions are consistent with all of our numerical results.

Simulation of traffic control signal systems

Science.gov (United States)

Connolly, P. J.; Concannon, P. A.; Ricci, R. C.

1974-01-01

In recent years there has been considerable interest in the development and testing of control strategies for networks of urban traffic signal systems by simulation. Simulation is an inexpensive and timely method for evaluating the effect of these traffic control strategies since traffic phenomena are too complex to be defined by analytical models and since a controlled experiment may be hazardous, expensive, and slow in producing meaningful results. This paper describes the application of an urban traffic corridor program, to evaluate the effectiveness of different traffic control strategies for the Massachusetts Avenue TOPICS Project.

[Comics for traffic education: evaluation of a traffic safety campaign].

Science.gov (United States)

Bonfadelli, H

1989-01-01

Traffic safety campaigns often are ineffective to change driving behavior because they don't reach the target group or are recognized only by people who are already interested or concerned. The evaluation of a traffic safety campaign called "Leo Lässig", addressed to young new drivers, shows that recognition and acceptance by the target group were stimulated by the age-conform means of comic-strips.

SIGNAL CONTROLLED JUNCTIONS CALCULATIONS IN TRAFFIC-CAPACITY ASSESSMENT - AIMSUN, OMNITRANS, WEBSTER AND TP 10/2010 RESULTS COMPARISON

Directory of Open Access Journals (Sweden)

Ľubomír ČERNICKÝ

2016-03-01

Full Text Available Every increase in traffic volume on road network in towns can lead to overcrowding of road network. This results in undesirable external costs such as traffic congestions, which cause high loses in time during transportation, increased fuel consumption and thus higher production of greenhouse gases and noise. This all ultimately reduces the attractiveness of the area. The increase of traffic volume and therefrom derived traffic problems are needed to be solved during traffic-capacity assessment of every larger investment. The software can help to assess increased traffic in solved area and thus help authorities to make a right decision during approving of the investment plan. This article is focused on comparison of two software – Aimsun and OmniTrans, and calculations according to Webster and technical regulations for assessing junction capacity in the Slovak Republic. The packages outputs are also compared to the measured data at the assessed junction in this article. The analysis showed that outputs of various tools differ, generally all packages showed higher delays compared to measured data at the main road and lower delays compared to measured data at the side roads.

Impact Analysis of Land Use on Traffic Congestion Using Real-Time Traffic and POI

Directory of Open Access Journals (Sweden)

Tianqi Zhang

2017-01-01

Full Text Available This paper proposed a new method to describe, compare, and classify the traffic congestion points in Beijing, China, by using the online map data and further revealed the relationship between traffic congestion and land use. The data of the point of interest (POI and the real-time traffic was extracted from an electronic map of the area in the fourth ring road of Beijing. The POIs were quantified based on the architectural area of the land use; the congestion points were identified based on real-time traffic. Then, the cluster analysis using the attributes of congestion time was conducted to identify the main traffic congestion areas. The result of a linear regression analysis between the congestion time and the land use showed that the influence of the high proportion of commercial land use on the traffic congestion was significant. Also, we considered five types of land use through performing a linear regression analysis between the congestion time and the ratio of four types of land use. The results showed that the reasonable ratio of land use types could efficiently reduce congestion time. This study makes contributions to the policy-making of urban land use.

Spatial patterns monitoring of road traffic injuries in Karachi metropolis.

Science.gov (United States)

Lateef, Muhammad U

2011-06-01

This article aims to assess the pattern of road traffic injuries (RTIs) and fatalities in Karachi metropolis. Assessing the pattern of RTIs in Karachi at this juncture is important for many reasons. The rapid motorisation in the recent years due to the availability of credit has significantly increased the traffic volume of the city. Since then, the roads of Karachi have continuously developed at a rapid pace. This development has come with a high human loss, because the construction of multilevel flyovers, signal-free corridors and the resulting high-speed traffic ultimately increase the severity of injuries. The reasons for this high proportion are inadequate infrastructure, poor enforcement of safety regulations, high crash severity index and greater population of vulnerable road user groups (riders and pedestrians). This research is the first of its kind in the country to have a geocoded database of fatalities and injuries in a geographical information system for the entire city of Karachi. In fact, road crashes are both predictable and preventable. Developing countries should learn from the experience of highly motorised nations to avoid the high burden of RTIs by adopting road safety and prevention measures.

The Crossroads of Synaptic Growth Signaling, Membrane Traffic and Neurological Disease: Insights from Drosophila.

Science.gov (United States)

Deshpande, Mugdha; Rodal, Avital A

2016-02-01

Neurons require target-derived autocrine and paracrine growth factors to maintain proper identity, innervation, homeostasis and survival. Neuronal growth factor signaling is highly dependent on membrane traffic, both for the packaging and release of the growth factors themselves, and for regulation of intracellular signaling by their transmembrane receptors. Here, we review recent findings from the Drosophila larval neuromuscular junction (NMJ) that illustrate how specific steps of intracellular traffic and inter-organelle interactions impinge on signaling, particularly in the bone morphogenic protein, Wingless and c-Jun-activated kinase pathways, regulating elaboration and stability of NMJ arbors, construction of synapses and synaptic transmission and homeostasis. These membrane trafficking and signaling pathways have been implicated in human motor neuron diseases including amyotrophic lateral sclerosis and hereditary spastic paraplegia, highlighting their importance for neuronal health and survival. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Recruitment of Cbl-b to B cell antigen receptor couples antigen recognition to Toll-like receptor 9 activation in late endosomes.

Directory of Open Access Journals (Sweden)

Margaret Veselits

Full Text Available Casitas B-lineage lymphoma-b (Cbl-b is a ubiquitin ligase (E3 that modulates signaling by tagging molecules for degradation. It is a complex protein with multiple domains and binding partners that are not involved in ubiquitinating substrates. Herein, we demonstrate that Cbl-b, but not c-Cbl, is recruited to the clustered B cell antigen receptor (BCR and that Cbl-b is required for entry of endocytosed BCRs into late endosomes. The E3 activity of Cbl-b is not necessary for BCR endocytic trafficking. Rather, the ubiquitin associated (UBA domain is required. Furthermore, the Cbl-b UBA domain is sufficient to confer the receptor trafficking functions of Cbl-b on c-Cbl. Cbl-b is also required for entry of the Toll-like receptor 9 (TLR9 into late endosomes and for the in vitro activation of TLR9 by BCR-captured ligands. These data indicate that Cbl-b acts as a scaffolding molecule to coordinate the delivery of the BCR and TLR9 into subcellular compartments required for productively delivering BCR-captured ligands to TLR9.

Remotely Accessed Vehicle Traffic Management System

Science.gov (United States)

Al-Alawi, Raida

2010-06-01

The ever increasing number of vehicles in most metropolitan cities around the world and the limitation in altering the transportation infrastructure, led to serious traffic congestion and an increase in the travelling time. In this work we exploit the emergence of novel technologies such as the internet, to design an intelligent Traffic Management System (TMS) that can remotely monitor and control a network of traffic light controllers located at different sites. The system is based on utilizing Embedded Web Servers (EWS) technology to design a web-based TMS. The EWS located at each intersection uses IP technology for communicating remotely with a Central Traffic Management Unit (CTMU) located at the traffic department authority. Friendly GUI software installed at the CTMU will be able to monitor the sequence of operation of the traffic lights and the presence of traffic at each intersection as well as remotely controlling the operation of the signals. The system has been validated by constructing a prototype that resembles the real application.

Traffic management simulation development.

Science.gov (United States)

2011-01-03

Microscopic simulation can provide significant support to traffic management center (TMC) operations. However, traffic simulation applications require data that are expensive and time-consuming to collect. Data collected by TMCs can be used as a prim...

Fair and efficient prices in traffic. Propositions on price policy for the sustainable development of traffic in Switzerland

International Nuclear Information System (INIS)

Maibach, M.; Ott, W.; Schreyer, Ch.

2000-01-01

Fair and efficient prices are a central topic in traffic policy, both at the EU level and in Switzerland. They should allow the efficient use of the infrastructure, set up fair terms of competition between road and rail traffic and reduce traffic loading. At the same time, they should also make it possible to finance traffic in a sustainable manner. One of the main directions of attack is letting those responsible pay the external costs which result from traffic jams, accidents and environmental pollution. With the planned introduction of the usage-dependent heavy traffic levy, Switzerland has made an important step in the direction of true cost-allocation for goods vehicles. As for the rest of road traffic, however, considerable external costs are still not covered. On the other hand, rail traffic is not able to cover its infrastructure costs itself in spite of compensation it receives for the provision of public services

Fair and efficient prices in traffic. Propositions on price policy for the sustainable development of traffic in Switzerland

International Nuclear Information System (INIS)

Maibach, M.; Ott, W.; Schreyer, Ch.

1999-01-01

Fair and efficient prices are a central topic in traffic policy, both at the European Union level and in Switzerland. They should allow the efficient use of the infrastructure, set up fair terms of competition between road and rail traffic and reduce traffic loading. At the same time, they should also make it possible to finance traffic in a sustainable manner. One of the main directions of attack is letting those responsible pay the external costs which result from traffic jams, accidents and environmental pollution. With the planned introduction of the usage-dependent heavy traffic levy, Switzerland has made an important step in the direction of true cost-allocation for goods vehicles. As for the rest of road traffic, however, considerable external costs are still not covered. On the other hand, rail traffic is not able to cover its infrastructure costs itself in spite of compensation it receives for the provision of public services [de

Simulation of load traffic and steeped speed control of conveyor

Science.gov (United States)

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.

A computerized traffic control algorithm to determine optimal traffic signal settings. Ph.D. Thesis - Toledo Univ.

Science.gov (United States)

Seldner, K.

1977-01-01

An algorithm was developed to optimally control the traffic signals at each intersection using a discrete time traffic model applicable to heavy or peak traffic. Off line optimization procedures were applied to compute the cycle splits required to minimize the lengths of the vehicle queues and delay at each intersection. The method was applied to an extensive traffic network in Toledo, Ohio. Results obtained with the derived optimal settings are compared with the control settings presently in use.

Dissociative Tendencies and Traffic Incidents

Directory of Open Access Journals (Sweden)

Valle, Virginia

2012-01-01

Full Text Available This paper analyses the relationship between dissociative experiences and road traffic incidents (crashes and traffic tickets in drivers (n=295 from Mar del Plata (Argentina city. A self-report questionnaire was applied to assess traffic crash involvement and sociodemographic variables. Dissociative tendencies were assessed by a modified version of the DES scale. To examine differences in DES scores tests of the difference of means were applied. Drivers who reported to be previously involved in traffic incidents obtained higher puntuations in the dissociative experiences scale than drivers who did not report such events. This result is observed for the total scale and for the three sub-scales (absorption, amnesia and depersonalization. However, differences appeared mainly for minor damage collisions. Further studies are needed to evaluate the role of dissociative tendencies as a risk factor in road traffic safety.

TBC-8, a putative RAB-2 GAP, regulates dense core vesicle maturation in Caenorhabditis elegans.

Science.gov (United States)

Hannemann, Mandy; Sasidharan, Nikhil; Hegermann, Jan; Kutscher, Lena M; Koenig, Sabine; Eimer, Stefan

2012-01-01

Dense core vesicles (DCVs) are thought to be generated at the late Golgi apparatus as immature DCVs, which subsequently undergo a maturation process through clathrin-mediated membrane remodeling events. This maturation process is required for efficient processing of neuropeptides within DCVs and for removal of factors that would otherwise interfere with DCV release. Previously, we have shown that the GTPase, RAB-2, and its effector, RIC-19, are involved in DCV maturation in Caenorhabditis elegans motoneurons. In rab-2 mutants, specific cargo is lost from maturing DCVs and missorted into the endosomal/lysosomal degradation route. Cargo loss could be prevented by blocking endosomal delivery. This suggests that RAB-2 is involved in retention of DCV components during the sorting process at the Golgi-endosomal interface. To understand how RAB-2 activity is regulated at the Golgi, we screened for RAB-2-specific GTPase activating proteins (GAPs). We identified a potential RAB-2 GAP, TBC-8, which is exclusively expressed in neurons and which, when depleted, shows similar DCV maturation defects as rab-2 mutants. We could demonstrate that RAB-2 binds to its putative GAP, TBC-8. Interestingly, TBC-8 also binds to the RAB-2 effector, RIC-19. This interaction appears to be conserved as TBC-8 also interacted with the human ortholog of RIC-19, ICA69. Therefore, we propose that a dynamic ON/OFF cycling of RAB-2 at the Golgi induced by the GAP/effector complex is required for proper DCV maturation.

Tourism in protected areas: Disentangling road and traffic effects on intra-guild scavenging processes.

Science.gov (United States)

Donázar, José Antonio; Ceballos, Olga; Cortés-Avizanda, Ainara

2018-07-15

The expansion of road networks and the increase in traffic have emerged in recent years as key threats to the conservation of biodiversity. This is particularly concerning in many protected areas because the increase of recreational activities requiring the use of vehicles. Effects of roads and traffic within guild scenarios and ecological processes remain however poorly known. Here we examined how road proximity and traffic intensity influence patterns of resource use in an Old-World avian scavenger guild living in a protected natural park in northern Spain. We experimentally placed 130 carcasses at different distances from a scenic road in the centre of the park. Vehicles were recorded by means of traffic counters which revealed that maximum numbers were reached during weekends and holidays and during the middle hours of the day. Avian scavenger attendance at carcasses was recorded by means of camera-traps. Obligated scavengers, Eurasian griffon (Gyps fulvus) and Egyptian vultures (Neophron percnopterus) were frequently observed (59.4% and 37.7% of the consumed carcasses) together with five other facultative scavenger species. We found that the richness (number of species) and the probability of consumption of the resource were reduced the smaller the distance to the road and in days with higher traffic intensity. The same factors affected the probability of presence of all the scavenger species. Moreover, some of them, notably griffon vultures, showed hourly patterns of carcass attendance suggesting avoidance of maximum traffic levels. Our results highlight that roads and traffic would trigger consequences on the structure and functioning of scavenger food webs, which may be particularly concerning in protected areas with remarkable levels of biodiversity. Future regulations at protected areas should couple both traffic and tourist affluence with wildlife conservation. In this way important ecological processes would be preserved while maintaining a good

Wireless traffic steering for green cellular networks

CERN Document Server

Zhang, Shan; Zhou, Sheng; Niu, Zhisheng; Shen, Xuemin (Sherman)

2016-01-01

This book introduces wireless traffic steering as a paradigm to realize green communication in multi-tier heterogeneous cellular networks. By matching network resources and dynamic mobile traffic demand, traffic steering helps to reduce on-grid power consumption with on-demand services provided. This book reviews existing solutions from the perspectives of energy consumption reduction and renewable energy harvesting. Specifically, it explains how traffic steering can improve energy efficiency through intelligent traffic-resource matching. Several promising traffic steering approaches for dynamic network planning and renewable energy demand-supply balancing are discussed. This book presents an energy-aware traffic steering method for networks with energy harvesting, which optimizes the traffic allocated to each cell based on the renewable energy status. Renewable energy demand-supply balancing is a key factor in energy dynamics, aimed at enhancing renewable energy sustainability to reduce on-grid energy consum...

Using the Tools in TRADE I: A Decision Support System for Traffic Light maintenance

NARCIS (Netherlands)

van Vlijmen, S.F.M.; Wieringa, Roelf J.

1997-01-01

In this report we develop a specification of requirements and conceptual design of a decision support system for maintenance of the lamps in traffic regulation systems. Requirements and design are both specified using semi-formal techniques in TRADE (Toolkit for Requirements and Design Engineering).

Controlled Traffic Farming

OpenAIRE

Controlled Traffic Farming Europe

2011-01-01

Metadata only record Controlled Traffic Farming (CTF) is a farming method used to reduce soil compaction, decrease inputs, and improve soil structure when coupled with reduced-till or no-till practices. This practices utilizes permanent traffic/wheel zones to limit soil compaction to a specific area. This website provides practical information on CTF, case studies, workshops, and links to additional resources.

Traffic flow wide-area surveillance system definition

Energy Technology Data Exchange (ETDEWEB)

Allgood, G.O.; Ferrell, R.K.; Kercel, S.W.; Abston, R.A.; Carnal, C.L. [Oak Ridge National Lab., TN (United States); Moynihan, P.I. [Jet Propulsion Lab., Pasadena, CA (United States)

1994-11-01

Traffic Flow Wide-Area Surveillance (TFWAS) is a system for assessing the state of traffic flow over a wide area for enhanced traffic control and improved traffic management and planning. The primary purpose of a TFWAS system is to provide a detailed traffic flow description and context description to sophisticated traffic management and control systems being developed or envisioned for the future. A successful TFWAS system must possess the attributes of safety, reconfigurability, reliability, and expandability. The primary safety premise of TFWAS is to ensure that no action or failure of the TFWAS system or its components can result in risk of injury to humans. A wide variety of communication techniques is available for use with TFWAS systems. These communication techniques can be broken down into two categories, landlines and wireless. Currently used and possible future traffic sensing technologies have been examined. Important criteria for selecting TFWAS sensors include sensor capabilities, costs, operational constraints, sensor compatibility with the infrastructure, and extent. TFWAS is a concept that can take advantage of the strengths of different traffic sensing technologies, can readily adapt to newly developed technologies, and can grow with the development of new traffic control strategies. By developing innovative algorithms that will take information from a variety of sensor types and develop descriptions of traffic flows over a wide area, a more comprehensive understanding of the traffic state can be provided to the control system to perform the most reasonable control actions over the entire wide area. The capability of characterizing the state of traffic over an entire region should revolutionize developments in traffic control strategies.

Qualitative and Quantitative Analysis of Congested Marine Traffic Environment – An Application Using Marine Traffic Simulation System

Directory of Open Access Journals (Sweden)

Kazuhiko Hasegawa

2013-06-01

Full Text Available Difficulty of sailing is quite subjective matter. It depends on various factors. Using Marine Traffic Simulation System (MTSS developed by Osaka University this challenging subject is discussed. In this system realistic traffic flow including collision avoidance manoeuvres can be reproduced in a given area. Simulation is done for southward of Tokyo Bay, Strait of Singapore and off-Shanghai area changing traffic volume from 5 or 50 to 150 or 200% of the present volume. As a result, strong proportional relation between near-miss ratio and traffic density per hour per sailed area is found, independent on traffic volume, area size and configuration. The quantitative evaluation index of the difficulty of sailing, here called risk rate of the area is defined using thus defined traffic density and near-miss ratio.

32 CFR 634.28 - Traffic accident investigation.

Science.gov (United States)

2010-07-01

... 32 National Defense 4 2010-07-01 2010-07-01 true Traffic accident investigation. 634.28 Section... ENFORCEMENT AND CRIMINAL INVESTIGATIONS MOTOR VEHICLE TRAFFIC SUPERVISION Traffic Supervision § 634.28 Traffic accident investigation. Installation law enforcement personnel must make detailed investigations of...

Flow level performance approximations for elastic traffic integrated with prioritized stream traffic

NARCIS (Netherlands)

Malhotra, R.; Berg, J.L. van den

2007-01-01

Almost all traffic in todays networks can be classified as being either stream or elastic. The support of these two traffic types is possible either with a Differentiated (DiffServ) or an Integrated Services (IntServ) architecture. However, both DiffServ and IntServ rely on efficient scheduling

Criterion for traffic phases in single vehicle data and empirical test of a microscopic three-phase traffic theory

International Nuclear Information System (INIS)

Kerner, Boris S; Klenov, Sergey L; Hiller, Andreas

2006-01-01

Based on empirical and numerical microscopic analyses, the physical nature of a qualitatively different behaviour of the wide moving jam phase in comparison with the synchronized flow phase-microscopic traffic flow interruption within the wide moving jam phase-is found. A microscopic criterion for distinguishing the synchronized flow and wide moving jam phases in single vehicle data measured at a single freeway location is presented. Based on this criterion, empirical microscopic classification of different local congested traffic states is performed. Simulations made show that the microscopic criterion and macroscopic spatiotemporal objective criteria lead to the same identification of the synchronized flow and wide moving jam phases in congested traffic. Microscopic models in the context of three-phase traffic theory have been tested based on the microscopic criterion for the phases in congested traffic. It is found that microscopic three-phase traffic models can explain both microscopic and macroscopic empirical congested pattern features. It is obtained that microscopic frequency distributions for vehicle speed difference as well as fundamental diagrams and speed correlation functions can depend on the spatial co-ordinate considerably. It turns out that microscopic optimal velocity (OV) functions and time headway distributions are not necessarily qualitatively different, even if local congested traffic states are qualitatively different. The reason for this is that important spatiotemporal features of congested traffic patterns are lost in these as well as in many other macroscopic and microscopic traffic characteristics, which are widely used as the empirical basis for a test of traffic flow models, specifically, cellular automata traffic flow models

Traffic calming for the prevention of road traffic injuries: systematic review and meta-analysis.

Science.gov (United States)

Bunn, F; Collier, T; Frost, C; Ker, K; Roberts, I; Wentz, R

2003-09-01

To assess whether area-wide traffic calming schemes can reduce road crash related deaths and injuries. Systematic review and meta-analysis. Cochrane Injuries Group Specialised Register, Cochrane Central Register of Controlled Trials, Medline, EMBASE, Sociological Abstracts Science (and social science) citation index, National Technical Information service, Psychlit, Transport Research Information Service, International Road Research Documentation, and Transdoc, and web sites of road safety organisation were searched; experts were contacted, conference proceedings were handsearched, and relevant reference lists were checked. Randomised controlled trials, and controlled before/after studies of area-wide traffic calming schemes designed to discourage and slow down through traffic on residential roads. Data were collected on road user deaths, injuries, and traffic crashes. For each study rate ratios were calculated, the ratio of event rates before and after intervention in the traffic calmed area divided by the corresponding ratio of event rates in the control area, which were pooled to give an overall estimate using a random effects model. Sixteen controlled before/after studies met our inclusion criteria. Eight studies reported the number of road user deaths: pooled rate ratio 0.63 (95% confidence interval (CI) 0.14 to 2.59). Sixteen studies reported the number of injuries (fatal and non-fatal): pooled rate ratio 0.89 (95% CI 0.80 to 1.00). All studies were in high income countries. Area-wide traffic calming in towns and cities has the potential to reduce road traffic injuries. However, further rigorous evaluations of this intervention are needed, especially in low and middle income countries.

Model for traffic emissions estimation

Science.gov (United States)

Alexopoulos, A.; Assimacopoulos, D.; Mitsoulis, E.

A model is developed for the spatial and temporal evaluation of traffic emissions in metropolitan areas based on sparse measurements. All traffic data available are fully employed and the pollutant emissions are determined with the highest precision possible. The main roads are regarded as line sources of constant traffic parameters in the time interval considered. The method is flexible and allows for the estimation of distributed small traffic sources (non-line/area sources). The emissions from the latter are assumed to be proportional to the local population density as well as to the traffic density leading to local main arteries. The contribution of moving vehicles to air pollution in the Greater Athens Area for the period 1986-1988 is analyzed using the proposed model. Emissions and other related parameters are evaluated. Emissions from area sources were found to have a noticeable share of the overall air pollution.

Study of physio-psychological effects on traffic wardens due to traffic noise pollution; exposure-effect relation.

Science.gov (United States)

Tabraiz, Shamas; Ahmad, Saeed; Shehzadi, Iffat; Asif, Muhammad Bilal

2015-01-01

Noise pollution has increased to alarming extent in most of the urban areas in Pakistan. It is assumed even more perilous than air and water pollution due to its direct acute and chronic physio-psychological effects. The objective of this study is to analyze and evaluate the psychological and physiological effects caused by traffic noise on traffic wardens and to find relation type between exposure time and effect. Three wardens check posts near roads were selected for survey in Taxila and Islamabad cities of Pakistan. Survey conducted included noise measurements at aforementioned check posts for one month and Performa based interviews of traffic wardens. Analysis of results showed that noise levels varied between 85-106 dB hence violating OSHA regulations. Major psychological effects found in wardens were aggravated depression 58%, stress 65%, public conflict 71%, irritation and annoyance 54%, behavioral affects 59% and speech interference 56%. Physiological effects found were hypertension 87%, muscle tension 64%, exhaustion 48%, low performance levels 55%, concentration loss 93%, hearing impairment 69%, headache 74% and cardiovascular issue 71%. Relation between exposure time and effects were evaluated by using simple regression test in excel. Percentage of psychological and physiological effects in wardens varied with the exposure time; aggravated depression (R(2) = 0.946, P = 0.133), stress suffering (R(2) = 0.014, P = 0.173), public conflict (R(2) = 0.946, P = 0.133), irritation and annoyance (R(2) = 0.371, P = 0.137), behavioral affects (R(2) = 0.596, P = 0.0616) and speech interference (R(2) = 0.355, P = 0.445), hypertension (R(2) = 0.96, P = 0.00095) and cardiovascular issue (R(2) = 0.775, P = 0.044).

OPTIMASI BANDWITH MENGGUNAKAN TRAFFIC SHAPPING

Directory of Open Access Journals (Sweden)

Imam Riadi

2010-01-01

Full Text Available Perkembangan layanan komunikasi telah berkembang sangat pesat. Salah satunya adalah pemanfaatan penggunaan bandwith untuk mengakases jaringan Internet. Traffic shapping bandwidth dapat memberikan efesiensi dalam hal pemanfaatan bandwidth pada instansi yang melakukan manajemen dalam lalulintas jaringannya. Metode yang digunakan dalam penelitian ini adalah metode literatur yaitu metode pengumpulan data yang dilakukan dengan mengumpulkan sumber- sumber data yang terkait dan metode eksperimen yaitu melakukan penelitian dengan mengkonfigurasi Mikrotik RouterOS untuk melakukan traffic shapping bandwidth. Hasil penelitian ini adalah router yang dapat melakukan traffic shapping sehingga kualitas koneksi menjadi lebih baik untuk mengakses dan mengambil data dari sebuah website. Kata kunci : Optimasi, Bandwith, Traffic Shapping, MikroTik. 

Bicycle traffic in urban areas

Directory of Open Access Journals (Sweden)

Anđelković Zorica

2015-01-01

Full Text Available Cycling is a term describing the use of bicycles, but also any mean of transport driven solely by human power. Development of bicycle traffic in urban areas involves construction of cycling infrastructure, adapting streets and other traffic infrastructure to a form suitable for cycling and other means of transport (individual motorized traffic, public transport, walking, ensuring the adequate budget and systematic planning and development of sustainable transport in cities. The paper presents basic settings and conditions as input elements to plan bicycle traffic in urban areas, as well as program- design conditions which lead the activities of planners and designers of urban roads in connection with cyclists.

Traffic light control by multiagent reinforcement learning systems

NARCIS (Netherlands)

Bakker, B.; Whiteson, S.; Kester, L.; Groen, F.C.A.; Babuška, R.; Groen, F.C.A.

2010-01-01

Traffic light control is one of the main means of controlling road traffic. Improving traffic control is important because it can lead to higher traffic throughput and reduced traffic congestion. This chapter describes multiagent reinforcement learning techniques for automatic optimization of

Traffic Light Control by Multiagent Reinforcement Learning Systems

NARCIS (Netherlands)

Bakker, B.; Whiteson, S.; Kester, L.J.H.M.; Groen, F.C.A.

2010-01-01

Traffic light control is one of the main means of controlling road traffic. Improving traffic control is important because it can lead to higher traffic throughput and reduced traffic congestion. This chapter describes multiagent reinforcement learning techniques for automatic optimization of

The importance of spatial orientation and knowledge of traffic signs for children's traffic safety.

Science.gov (United States)

Trifunović, Aleksandar; Pešić, Dalibor; Čičević, Svetlana; Antić, Boris

2017-05-01

Pre-school children, as well as children from lower grades in primary school, who although rarely, completely independently participate in traffic, represent a vulnerable population from the standpoint of traffic safety. The greatest number of children were injured or killed in road traffic crashes on their way from home to kindergarten or school. Mostly due to lack of experience, children's behavior is confusing and often reckless and hasty. Safe behavior in the traffic environment demands certain cognitive skills. Unlike adults, children have less than fully developed peripheral vision. Also, changes occur in color perception, i.e. discrimination. All this leads to the conclusion that the stage of physical and mental development of the child is very important for safe participation in traffic. So, to estimate if they are sufficiently equipped to participate safely in traffic, a sensitive test for young children that may be suitable for their level of cognitive development is required. Accordingly, road safety education should be arranged in such a way that considers the child's level of development, as has been shown to be more effective when started at younger ages. Play is the most natural and easiest way of learning because it is the lens through which children experience their world, and the world of others. Having this in mind, if we want to measure the abilities of a child, and their preparedness for safety participation in traffic, unavoidable is to use non-verbal tests. The purpose of this study is to explore primary schooler's spatial, and abilities of color perception and memorization, as well as their performances in interpreting the meaning of traffic signs. In addition, neighborhood environmental correlates (rural-urban) and possible individual differences influences on the relationship among these abilities was examined. Knowledge about these factors affecting children's safety can be applied to improve relevant intervention measures for promoting

Endosomal recognition of Lactococcus lactis G121 and its RNA by dendritic cells is key to its allergy-protective effects.

Science.gov (United States)

Stein, Karina; Brand, Stephanie; Jenckel, André; Sigmund, Anna; Chen, Zhijian James; Kirschning, Carsten J; Kauth, Marion; Heine, Holger

2017-02-01

Bacterial cowshed isolates are allergy protective in mice; however, the underlying mechanisms are largely unknown. We examined the ability of Lactococcus lactis G121 to prevent allergic inflammatory reactions. We sought to identify the ligands and pattern recognition receptors through which L lactis G121 confers allergy protection. L lactis G121-induced cytokine release and surface expression of costimulatory molecules by untreated or inhibitor-treated (bafilomycin and cytochalasin D) human monocyte-derived dendritic cells (moDCs), bone marrow-derived mouse dendritic cells (BMDCs), and moDC/naive CD4 + T-cell cocultures were analyzed by using ELISA and flow cytometry. The pathology of ovalbumin-induced acute allergic airway inflammation after adoptive transfer of BMDCs was examined by means of microscopy. L lactis G121-treated murine BMDCs and human moDCs released T H 1-polarizing cytokines and induced T H 1 T cells. Inhibiting phagocytosis and endosomal acidification in BMDCs or moDCs impaired the release of T H 1-polarizing cytokines, costimulatory molecule expression, and T-cell activation on L lactis G121 challenge. In vivo allergy protection mediated by L lactis G121 was dependent on endosomal acidification in dendritic cells (DCs). Toll-like receptor (Tlr) 13 -/- BMDCs showed a weak response to L lactis G121 and were unresponsive to its RNA. The T H 1-polarizing activity of L lactis G121-treated human DCs was blocked by TLR8-specific inhibitors, mediated by L lactis G121 RNA, and synergistically enhanced by activation of nucleotide-binding oligomerization domain-containing protein (NOD) 2. Bacterial RNA is the main driver of L lactis G121-mediated protection against experimentally induced allergy and requires both bacterial uptake by DCs and endosomal acidification. In mice L lactis G121 RNA signals through TLR13; however, the most likely intracellular receptor in human subjects is TLR8. Copyright © 2016 American Academy of Allergy, Asthma & Immunology

Traffic Information Unit, Traffic Information System, Vehicle Management System, Vehicle, and Method of Controlling a Vehicle

NARCIS (Netherlands)

Papp, Z.; Doodeman, G.J.N.; Nelisse, M.W.; Sijs, J.; Theeuwes, J.A.C.; Driessen, B.J.F.

2010-01-01

A traffic information unit (MD1, MD2, MD3) according to the invention comprises a facility (MI) for tracking vehicle state information of individual vehicles present at a traffic infrastructure and a facility (T) for transmitting said vehicle state information to a vehicle (70B, 70E). A traffic

Towards an agent based traffic regulation and recommendation system for the on-road air quality control.

Science.gov (United States)

Sadiq, Abderrahmane; El Fazziki, Abdelaziz; Ouarzazi, Jamal; Sadgal, Mohamed

2016-01-01

This paper presents an integrated and adaptive problem-solving approach to control the on-road air quality by modeling the road infrastructure, managing traffic based on pollution level and generating recommendations for road users. The aim is to reduce vehicle emissions in the most polluted road segments and optimizing the pollution levels. For this we propose the use of historical and real time pollution records and contextual data to calculate the air quality index on road networks and generate recommendations for reassigning traffic flow in order to improve the on-road air quality. The resulting air quality indexes are used in the system's traffic network generation, which the cartography is represented by a weighted graph. The weights evolve according to the pollution indexes and path properties and the graph is therefore dynamic. Furthermore, the systems use the available pollution data and meteorological records in order to predict the on-road pollutant levels by using an artificial neural network based prediction model. The proposed approach combines the benefits of multi-agent systems, Big data technology, machine learning tools and the available data sources. For the shortest path searching in the road network, we use the Dijkstra algorithm over Hadoop MapReduce framework. The use Hadoop framework in the data retrieve and analysis process has significantly improved the performance of the proposed system. Also, the agent technology allowed proposing a suitable solution in terms of robustness and agility.

PENGEMBANGAN SISTEM PENGENDALIAN TRAFFIC DAN WEB FILTERING PADA JARINGAN INTERNET BERBASIS HOTSPOT

Directory of Open Access Journals (Sweden)

Riska Robianto

2016-12-01

Full Text Available Utilization of Internet services to the hotspot network raises particular problems in traffic control and web filtering. Linux as an operating system that is open (open source, offers a variety of internet firewall systems to assist in the management of traffic, bandwidth, web filtering and one using Smoothwall Express to Linux which can convert a personal computer (PC to the Internet firewall reliable and stable. Smoothwall allows to integrated with the add-ons extends like AdvProxy and URL Filter to improve the performance of Smoohtwall and simplify configuration of the traffic control and web filtering. The research was conducted by collecting data from various sources related, then experiment with implementing Smoohtwall by adding add-ons extends to help administrators manage and regulate the bandwidth allocation of each client, block websites (URLs that are not needed. Based on these results bandwidth management can be divided equally and can block websites (URLs that are not needed like porn sites, warez, bad word, and also the quality of the network connection updatesites more stable.

VBR video traffic models

CERN Document Server

Tanwir, Savera

2014-01-01

There has been a phenomenal growth in video applications over the past few years. An accurate traffic model of Variable Bit Rate (VBR) video is necessary for performance evaluation of a network design and for generating synthetic traffic that can be used for benchmarking a network. A large number of models for VBR video traffic have been proposed in the literature for different types of video in the past 20 years. Here, the authors have classified and surveyed these models and have also evaluated the models for H.264 AVC and MVC encoded video and discussed their findings.

Photonics approach to traffic signs

Science.gov (United States)

Litwin, Dariusz; Galas, Jacek; CzyŻewski, Adam; Rymsza, Barbara; Kornalewski, Leszek; Kryszczyński, Tadeusz; Mikucki, Jerzy; Wikliński, Piotr; Daszkiewicz, Marek; Malasek, Jacek

2016-12-01

The automotive industry has been always a driving force for all economies. Despite of its beneficial meaning to every society it brings also many issues including wide area of road safety. The latter has been enforced by the increasing number of cars and the dynamic development of the traffic as a whole. Road signs and traffic lights are crucial in context of good traffic arrangement and its fluency. Traffic designers are used to treat horizontal road signs independently of vertical signs. However, modern light sources and growing flexibility in shaping optical systems create opportunity to design more advanced and smart solutions. In this paper we present an innovative, multidisciplinary approach that consists in tight interdependence of different traffic signals. We describe new optical systems together with their influence on the perception of the road user. The analysis includes maintenance and visibility in different weather conditions. A special attention has been focused on intersections of complex geometry.

Traffic accidents: an econometric investigation

OpenAIRE

Tito Moreira; Adolfo Sachsida; Loureiro Paulo

2004-01-01

Based on a sample of drivers in Brasilia's streets, this article investigates whether distraction explains traffic accidents. A probit model is estimated to determine the predictive power of several variables on traffic accidents. The main conclusion drawn from this study is that the proxies used to measure distraction, such as the use of cell phones and cigarette smoking in a moving vehicle, are significant factors in determining traffic accidents.

ReFlow: Reports on Internet Traffic

NARCIS (Netherlands)

Hoogesteger, Martijn; de Oliveira Schmidt, R.; Sperotto, Anna; Pras, Aiko

Internet traffic statistics can provide valuable information to network analysts and researchers about the traffic, technologies and main characteristics of today’s networks. For many years Internet2 maintained a public website with statistics about the traffic in the Abilene network. This site was

32 CFR 634.25 - Installation traffic codes.

Science.gov (United States)

2010-07-01

... 32 National Defense 4 2010-07-01 2010-07-01 true Installation traffic codes. 634.25 Section 634.25 National Defense Department of Defense (Continued) DEPARTMENT OF THE ARMY (CONTINUED) LAW ENFORCEMENT AND CRIMINAL INVESTIGATIONS MOTOR VEHICLE TRAFFIC SUPERVISION Traffic Supervision § 634.25 Installation traffic codes. (a) Installation or activit...

Road traffic noise: self-reported noise annoyance versus GIS modelled road traffic noise exposure.

Science.gov (United States)

Birk, Matthias; Ivina, Olga; von Klot, Stephanie; Babisch, Wolfgang; Heinrich, Joachim

2011-11-01

self-reported road traffic noise annoyance is commonly used in epidemiological studies for assessment of potential health effects. Alternatively, some studies have used geographic information system (GIS) modelled exposure to road traffic noise as an objective parameter. The aim of this study was to analyse the association between noise exposure due to neighbouring road traffic and the noise annoyance of adults, taking other determinants into consideration. parents of 951 Munich children from the two German birth cohorts GINIplus and LISAplus reported their annoyance due to road traffic noise at home. GIS modelled road traffic noise exposure (L(den), maximum within a 50 m buffer) from the noise map of the city of Munich was available for all families. GIS-based calculated distance to the closest major road (≥10,000 vehicles per day) and questionnaire based-information about family income, parental education and the type of the street of residence were explored for their potential influence. An ordered logit regression model was applied. The noise levels (L(den)) and the reported noise annoyance were compared with an established exposure-response function. the correlation between noise annoyance and noise exposure (L(den)) was fair (Spearman correlation r(s) = 0.37). The distance to a major road and the type of street were strong predictors for the noise annoyance. The annoyance modelled by the established exposure-response function and that estimated by the ordered logit model were moderately associated (Pearson's correlation r(p) = 0.50). road traffic noise annoyance was associated with GIS modelled neighbouring road traffic noise exposure (L(den)). The distance to a major road and the type of street were additional explanatory factors of the noise annoyance appraisal.

Predicting Information Flows in Network Traffic.

Science.gov (United States)

Hinich, Melvin J.; Molyneux, Robert E.

2003-01-01

Discusses information flow in networks and predicting network traffic and describes a study that uses time series analysis on a day's worth of Internet log data. Examines nonlinearity and traffic invariants, and suggests that prediction of network traffic may not be possible with current techniques. (Author/LRW)

Spectral Analysis of Traffic Functions in Urban Areas

Directory of Open Access Journals (Sweden)

Florin Nemtanu

2015-12-01

Full Text Available The paper is focused on the Fourier transform application in urban traffic analysis and the use of said transform in traffic decomposition. The traffic function is defined as traffic flow generated by different categories of traffic participants. A Fourier analysis was elaborated in terms of identifying the main traffic function components, called traffic sub-functions. This paper presents the results of the method being applied in a real case situation, that is, an intersection in the city of Bucharest where the effect of a bus line was analysed. The analysis was done using different time scales, while three different traffic functions were defined to demonstrate the theoretical effect of the proposed method of analysis. An extension of the method is proposed to be applied in urban areas, especially in the areas covered by predictive traffic control.

Competitive Traffic Assignment in Road Networks

Directory of Open Access Journals (Sweden)

Krylatov Alexander Y.

2016-09-01

Full Text Available Recently in-vehicle route guidance and information systems are rapidly developing. Such systems are expected to reduce congestion in an urban traffic area. This social benefit is believed to be reached by imposing the route choices on the network users that lead to the system optimum traffic assignment. However, guidance service could be offered by different competitive business companies. Then route choices of different mutually independent groups of users may reject traffic assignment from the system optimum state. In this paper, a game theoretic approach is shown to be very efficient to formalize competitive traffic assignment problem with various groups of users in the form of non-cooperative network game with the Nash equilibrium search. The relationships between the Wardrop’s system optimum associated with the traffic assignment problem and the Nash equilibrium associated with the competitive traffic assignment problem are investigated. Moreover, some related aspects of the Nash equilibrium and the Wardrop’s user equilibrium assignments are also discussed.

Myopic (HD-PTP, PTPN23) selectively regulates synaptic neuropeptide release.

Science.gov (United States)

Bulgari, Dinara; Jha, Anupma; Deitcher, David L; Levitan, Edwin S

2018-02-13

Neurotransmission is mediated by synaptic exocytosis of neuropeptide-containing dense-core vesicles (DCVs) and small-molecule transmitter-containing small synaptic vesicles (SSVs). Exocytosis of both vesicle types depends on Ca 2+ and shared secretory proteins. Here, we show that increasing or decreasing expression of Myopic (mop, HD-PTP, PTPN23), a Bro1 domain-containing pseudophosphatase implicated in neuronal development and neuropeptide gene expression, increases synaptic neuropeptide stores at the Drosophila neuromuscular junction (NMJ). This occurs without altering DCV content or transport, but synaptic DCV number and age are increased. The effect on synaptic neuropeptide stores is accounted for by inhibition of activity-induced Ca 2+ -dependent neuropeptide release. cAMP-evoked Ca 2+ -independent synaptic neuropeptide release also requires optimal Myopic expression, showing that Myopic affects the DCV secretory machinery shared by cAMP and Ca 2+ pathways. Presynaptic Myopic is abundant at early endosomes, but interaction with the endosomal sorting complex required for transport III (ESCRT III) protein (CHMP4/Shrub) that mediates Myopic's effect on neuron pruning is not required for control of neuropeptide release. Remarkably, in contrast to the effect on DCVs, Myopic does not affect release from SSVs. Therefore, Myopic selectively regulates synaptic DCV exocytosis that mediates peptidergic transmission at the NMJ.

Is vehicle automation enough to prevent crashes? Role of traffic operations in automated driving environments for traffic safety.

Science.gov (United States)

Jeong, Eunbi; Oh, Cheol; Lee, Seolyoung

2017-07-01

Automated driving systems (ADSs) are expected to prevent traffic accidents caused by driver carelessness on freeways. There is no doubt regarding this safety benefit if all vehicles in the transportation system were equipped with ADSs; however, it is implausible to expect that ADSs will reach 100% market penetration rate (MPR) in the near future. Therefore, the following question arises: 'Can ADSs, which consider only situations in the vicinity of an equipped vehicle, really contribute to a significant reduction in traffic accidents?' To address this issue, the interactions between equipped and unequipped vehicles must be investigated, which is the purpose of this study. This study evaluated traffic safety at different MPRs based on a proposed index to represent the overall rear-end crash risk of the traffic stream. Two approaches were evaluated for adjusting longitudinal vehicle maneuvers: vehicle safety-based maneuvering (VSM), which considers the crash risk of an equipped vehicle and its neighboring vehicles, and traffic safety-based maneuvering (TSM), which considers the overall crash risk in the traffic stream. TSM assumes that traffic operational agencies are able to monitor all the vehicles and to intervene in vehicle maneuvering. An optimization process, which attempts to obtain vehicle maneuvering control parameters to minimize the overall crash risk, is integrated into the proposed evaluation framework. The main purpose of employing the optimization process for vehicle maneuvering in this study is to identify opportunities to improve traffic safety through effective traffic management rather than developing a vehicle control algorithm that can be implemented in practice. The microscopic traffic simulator VISSIM was used to simulate the freeway traffic stream and to conduct systematic evaluations based on the proposed methodology. Both TSM and VSM achieved significant reductions in the potential for rear-end crashes. However, TSM obtained much greater

Prediction of vehicle crashes by drivers' characteristics and past traffic violations in Korea using a zero-inflated negative binomial model.

Science.gov (United States)

Kim, Dae-Hwan; Ramjan, Lucie M; Mak, Kwok-Kei

2016-01-01

Traffic safety is a significant public health challenge, and vehicle crashes account for the majority of injuries. This study aims to identify whether drivers' characteristics and past traffic violations may predict vehicle crashes in Korea. A total of 500,000 drivers were randomly selected from the 11.6 million driver records of the Ministry of Land, Transport and Maritime Affairs in Korea. Records of traffic crashes were obtained from the archives of the Korea Insurance Development Institute. After matching the past violation history for the period 2004-2005 with the number of crashes in year 2006, a total of 488,139 observations were used for the analysis. Zero-inflated negative binomial model was used to determine the incident risk ratio (IRR) of vehicle crashes by past violations of individual drivers. The included covariates were driver's age, gender, district of residence, vehicle choice, and driving experience. Drivers violating (1) a hit-and-run or drunk driving regulation at least once and (2) a signal, central line, or speed regulation more than once had a higher risk of a vehicle crash with respective IRRs of 1.06 and 1.15. Furthermore, female gender, a younger age, fewer years of driving experience, and middle-sized vehicles were all significantly associated with a higher likelihood of vehicle crashes. Drivers' demographic characteristics and past traffic violations could predict vehicle crashes in Korea. Greater resources should be assigned to the provision of traffic safety education programs for the high-risk driver groups.

Analysis of Malicious Traffic in Modbus/TCP Communications

Science.gov (United States)

Kobayashi, Tiago H.; Batista, Aguinaldo B.; Medeiros, João Paulo S.; Filho, José Macedo F.; Brito, Agostinho M.; Pires, Paulo S. Motta

This paper presents the results of our analysis about the influence of Information Technology (IT) malicious traffic on an IP-based automation environment. We utilized a traffic generator, called MACE (Malicious trAffic Composition Environment), to inject malicious traffic in a Modbus/TCP communication system and a sniffer to capture and analyze network traffic. The realized tests show that malicious traffic represents a serious risk to critical information infrastructures. We show that this kind of traffic can increase latency of Modbus/TCP communication and that, in some cases, can put Modbus/TCP devices out of communication.

5 CFR 842.405 - Air traffic controllers, firefighters, law enforcement officers, and nuclear materials couriers.

Science.gov (United States)

2010-01-01

... 5 Administrative Personnel 2 2010-01-01 2010-01-01 false Air traffic controllers, firefighters, law enforcement officers, and nuclear materials couriers. 842.405 Section 842.405 Administrative Personnel OFFICE OF PERSONNEL MANAGEMENT (CONTINUED) CIVIL SERVICE REGULATIONS (CONTINUED) FEDERAL EMPLOYEES RETIREMENT SYSTEM-BASIC ANNUITY Computations ...

30 CFR 56.9100 - Traffic control.

Science.gov (United States)

2010-07-01

... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Traffic control. 56.9100 Section 56.9100 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND NONMETAL MINE... Dumping Traffic Safety § 56.9100 Traffic control. To provide for the safe movement of self-propelled...

Life Times of Simulated Traffic Jams

Science.gov (United States)

Nagel, Kai

We study a model for freeway traffic which includes strong noise taking into account the fluctuations of individual driving behavior. The model shows emergent traffic jams with a self-similar appearance near the throughput maximum of the traffic. The lifetime distribution of these jams shows a short scaling regime, which gets considerably longer if one reduces the fluctuations when driving at maximum speed but leaves the fluctuations for slowing down or accelerating unchanged. The outflow from a traffic jam self-organizes into this state of maximum throughput.

2016 Traffic Safety Culture Index

Science.gov (United States)

... Newsroom SEARCH Driver Behavior & Performance 2016 Traffic Safety Culture Index This report presents the results of our annual Traffic Safety Culture Index survey, providing data on the attitudes and ...

Procedure for Marine Traffic Simulation with AIS Data

Directory of Open Access Journals (Sweden)

Rina Miyake

2015-03-01

Full Text Available It is essential to evaluate safety of marine traffic for the improvement of efficiency and safety of marine traffic. Spread of AIS makes observation of actual marine traffic more easily and faster than before. Besides, description of collision avoidance behaviours of ships are indispensable to simulate a realistic marine traffic. It is important to develop and implement an algorithm of collision avoidance corresponding to a target traffic or target area into the marine traffic simulation because actual actions for collision avoidance depend on circumstances where ships are sailing. The authors developed an automated marine traffic simulation system with AIS data. And in this paper, we proposed a series of systematic procedures for marine traffic simulation including analysing for collision avoidance behaviours using AIS data.

Impact Analysis of Land Use on Traffic Congestion Using Real-Time Traffic and POI

OpenAIRE

Zhang, Tianqi; Sun, Lishan; Yao, Liya; Rong, Jian

2017-01-01

This paper proposed a new method to describe, compare, and classify the traffic congestion points in Beijing, China, by using the online map data and further revealed the relationship between traffic congestion and land use. The data of the point of interest (POI) and the real-time traffic was extracted from an electronic map of the area in the fourth ring road of Beijing. The POIs were quantified based on the architectural area of the land use; the congestion points were identified based on ...

Systemic Approach to Traffic Evaluation of Mostar Airport

Directory of Open Access Journals (Sweden)

Eldo Raguž

2005-11-01

Full Text Available The attempt of this work is to systematically find solutionsfor Mostar Airport development through technical and technologicalharmonization of traffic processes undertaken in twoseparate organizations - airport and air traffic control and coordinationbetween other traffic branches. The work uses theindicators of traffic flows and tourist trends in the region, andtogether with the mentioned simulations it attempts to evaluatethe traffic potentials in the region by affecting the change in thecurrent negative traffic flows at Mostar Airport.

Traffic noise in Hyderabad city, part-II. vehicular contribution to road traffic noise

International Nuclear Information System (INIS)

Sheikh, G.H.

2001-01-01

The results of a road traffic noise survey carried out in Hyderabad city showed that the levels of traffic noise in the City are alarmingly high and much beyond the comfortable limits. There, in order to investigate the level of the noise emitted by different types of vehicles plying on the city roads and to assess their individual contribution to high level traffic noise, studies have been carried out on the measurement of noise emitted by motorcycles, buses, auto-rickshaws, and motor vehicle horns as they normally move on the city roads. The data collected has been analyzed for L/sub v99/, L/sub v90/, L/sub v50/, L/sub v10/ and L/sub v1/ and results are discussed with reference to the existing motor vehicle rules in Pakistan and motor vehicle noise emission limits set by the EEC and other developed countries. Some suggestion have also been made to limit high level traffic noise. (author)

Military Regulated Airspace: Atlantic / Gulf of Mexico

Data.gov (United States)

National Oceanic and Atmospheric Administration, Department of Commerce — Military regulated airspace areas depict the Air Traffic Control Assigned Airspace (ATCAA) and Airspace Corridor areas. The MarineCadastre.gov team worked with the...

Light signals for road traffic control.

NARCIS (Netherlands)

Schreuder, D.A.

1981-01-01

Signals for road traffic control are a major constituent of the modern traffic scene, particularly in built-up areas. A vast amount of research has been executed in the last two decennia, resulting in a fairly generally accepted view on what the requirements for effective traffic lights are. For the

Road Traffic in China

NARCIS (Netherlands)

Jie, L.; Van Zuylen, H.J.

2014-01-01

Traffic is tightly related to the social and economic development in a country. In China the development of the economy has been very fast in the past 30 years and this is still continuing. The transport infrastructure shows a similar pattern, while traffic is also rapidly growing. In urban areas

On traffic modelling in GPRS networks

DEFF Research Database (Denmark)

Madsen, Tatiana Kozlova; Schwefel, Hans-Peter; Prasad, Ramjee

2005-01-01

Optimal design and dimensioning of wireless data networks, such as GPRS, requires the knowledge of traffic characteristics of different data services. This paper presents an in-detail analysis of an IP-level traffic measurements taken in an operational GPRS network. The data measurements reported...... here are done at the Gi interface. The aim of this paper is to reveal some key statistics of GPRS data applications and to validate if the existing traffic models can adequately describe traffic volume and inter-arrival time distribution for different services. Additionally, we present a method of user...

DATA ANALYSIS OF TRAFFIC ACCIDENTS AND THEIR CAUSES IN GOMEL FOR 2013 AND 2014

Directory of Open Access Journals (Sweden)

S. A. Azemsha

2015-01-01

Full Text Available Measures undertaken for improvement of road traffic safety presuppose clampdown on violation of road traffic regulations but no attention is paid to the fact that there is no possibility to ensure road traffic safety due to inconformity of roads to the modern safety requirements. Therefore reduction in accident rate is connected with some changes in approaches to designing, construction and maintenance of roads. Nowadays when the number of automobiles is extremely large and their number is increasing with every passing year driver’s professionalism becomes the most significant factor. In these circumstances the professionalism is demonstrated not so much while driving in bad road conditions as it was previously but the professionalism is more revealed in the case when it is necessary to drive in conditions of large workloads and high manoeuvring rate when a special important role is given to the capability to forecast a situation, in other words the capability to read the road. Moral climate on the road is no less important as well and it practically fully depends on a driver.The paper contains an analysis of the Gomel traffic police data on quantitative distribution of road traffic accidents and their victims according to the time of day and month, lighting conditions, weather conditions, age and sex factors, types of violations. Situations of traffic behavior, motivations of drivers and road users, drivers training have been shown in terms of impact on the road traffic accident risk. The paper considers a human factor which rather often causes an accident. An evaluation has been given to such factor as road speed of transport facilities which exerts a significant influence on an accident risk and its severity.

Enhancing the Quality of Service for Real Time Traffic over Optical Burst Switching (OBS Networks with Ensuring the Fairness for Other Traffics.

Directory of Open Access Journals (Sweden)

Mohammed A Al-Shargabi

Full Text Available Optical burst switching (OBS networks have been attracting much consideration as a promising approach to build the next generation optical Internet. A solution for enhancing the Quality of Service (QoS for high priority real time traffic over OBS with the fairness among the traffic types is absent in current OBS' QoS schemes. In this paper we present a novel Real Time Quality of Service with Fairness Ratio (RT-QoSFR scheme that can adapt the burst assembly parameters according to the traffic QoS needs in order to enhance the real time traffic QoS requirements and to ensure the fairness for other traffic. The results show that RT-QoSFR scheme is able to fulfill the real time traffic requirements (end to end delay, and loss rate ensuring the fairness for other traffics under various conditions such as the type of real time traffic and traffic load. RT-QoSFR can guarantee that the delay of the real time traffic packets does not exceed the maximum packets transfer delay value. Furthermore, it can reduce the real time traffic packets loss, at the same time guarantee the fairness for non real time traffic packets by determining the ratio of real time traffic inside the burst to be 50-60%, 30-40%, and 10-20% for high, normal, and low traffic loads respectively.

Fostering Self-Regulation in Training Complex Cognitive Tasks

Science.gov (United States)

van Meeuwen, Ludo W.; Brand-Gruwel, Saskia; Kirschner, Paul A.; de Bock, Jeano J. P. R.; van Merriënboer, Jeroen J. G.

2018-01-01

In complex cognitive domains such as air traffic control, professionals must be able to adapt to and act upon continuing changes in a highly advanced technological work environment. To function optimally in such an environment, the controllers must be able to regulate their learning. Although these regulation skills should be part of their…

Structuring of Road Traffic Flows

Directory of Open Access Journals (Sweden)

Planko Rožić

2005-09-01

Full Text Available Systemic traffic count on the Croatian road network hasbeen carried out for more than three decades in different ways.During this period a large number of automatic traffic countershave been installed, and they operate on different principles.The traffic count has been analyzed from the aspect of vehicleclassification. The count results can be only partly comparedsince they yield different structures of traffic flows. Special analysisrefers to the classification of vehicles by automatic trafficcounters.During the research, a database has been formed with physicalelements of vehicles of over five thousand vehicle types. Theresearch results prove that the vehicle length only is not sufficientfor the classification of vehicles, the way it is used in thepresent automatic traffic counts, but rather the number of axles,the wheelbase as well as the front and rear overhangs needto be considered as well. Therefore, the detector system shouldapply also the detector of axles.The results have been presented that were obtained as partof the program TEST- Technological, research, developmentproject supported by the Minist1y of Science, Education andSport.

Visual Analysis in Traffic & Re-identification

DEFF Research Database (Denmark)

Møgelmose, Andreas

and analysis, and person re-identification. In traffic sign detection, the work comprises a thorough survey of the state of the art, assembly of the worlds largest public dataset with U.S. traffic signs, and work in machine learning based detection algorithms. It was shown that detection of U.S. traffic signs...

Some random models in traffic science

Energy Technology Data Exchange (ETDEWEB)

Hjorth, U.

1996-06-01

We give an overview of stochastic models for the following traffic phenomena. Models for traffic flow including gaps and capacities for lanes, crossings and roundabouts. Models for wanted and achieved speed distributions. Mode selection models including dispersed equilibrium models and traffic accident models. Also some statistical questions are discussed. 60 refs, 1 tab

Autosolitons in applied physics and traffic flow

International Nuclear Information System (INIS)

Kerner, B.S.

1996-01-01

A review of investigations of autosolitons in nonlinear systems which are of interest for the applied physics and for the transportation research is presented. Autosolitons are solitary intrinsic states which can be formed in a broad class of physical, chemical, biological dissipative distributed media and in traffic flow. Properties of autosolitons which are general for physical systems and for traffic flow will be discussed. Based on results of recent investigations of traffic jams in traffic flow, a comparison of nonlinear characteristics of traffic jams and with nonlinear properties of autosolitons which can be formed in active systems with diffusion will be given. Forms, properties, processes of evolution of autosolitons in traffic flow, in semiconductors and in gas discharge plasma are considered. copyright 1996 American Institute of Physics

Distraction-related road traffic collisions

African Journals Online (AJOL)

drivers involved in road traffic collisions (RTC) were using mobile phones. Our study supports ... while driving. Keywords: Distraction, prevention, road traffic collision, mobile phone. ..... keeps us connected with others with great advantages.

Prenatal and childhood traffic-related air pollution exposure and childhood executive function and behavior.

Science.gov (United States)

Harris, Maria H; Gold, Diane R; Rifas-Shiman, Sheryl L; Melly, Steven J; Zanobetti, Antonella; Coull, Brent A; Schwartz, Joel D; Gryparis, Alexandros; Kloog, Itai; Koutrakis, Petros; Bellinger, David C; Belfort, Mandy B; Webster, Thomas F; White, Roberta F; Sagiv, Sharon K; Oken, Emily

Traffic-related air pollution exposure may influence brain development and function and thus be related to neurobehavioral problems in children, but little is known about windows of susceptibility. Examine associations of gestational and childhood exposure to traffic-related pollution with executive function and behavior problems in children. We studied associations of pre- and postnatal pollution exposures with neurobehavioral outcomes in 1212 children in the Project Viva pre-birth cohort followed to mid-childhood (median age 7.7years). Parents and classroom teachers completed the Behavior Rating Inventory of Executive Function (BRIEF) and the Strengths and Difficulties Questionnaire (SDQ). Using validated spatiotemporal models, we estimated exposure to black carbon (BC) and fine particulate matter (PM 2.5 ) in the third trimester of pregnancy, from birth to 3years, from birth to 6years, and in the year before behavioral ratings. We also measured residential distance to major roadways and near-residence traffic density at birth and in mid-childhood. We estimated associations of BC, PM 2.5 , and other traffic exposure measures with BRIEF and SDQ scores, adjusted for potential confounders. Higher childhood BC exposure was associated with higher teacher-rated BRIEF Behavioral Regulation Index (BRI) scores, indicating greater problems: 1.0 points (95% confidence interval (CI): 0.0, 2.1) per interquartile range (IQR) increase in birth-age 6BC, and 1.7 points (95% CI: 0.6, 2.8) for BC in the year prior to behavioral ratings. Mid-childhood residential traffic density was also associated with BRI score (0.6, 95% CI: 0.1, 1.1). Birth-age 3BC was not associated with BRIEF or SDQ scores. Third trimester BC exposure was not associated with teacher-rated BRI scores (-0.2, 95% CI: -1.1, 0.8), and predicted lower scores (fewer problems) on the BRIEF Metacognition Index (-1.2, 95% CI: -2.2, -0.2) and SDQ total difficulties (-0.9, 95% CI: -1.4, -0.4). PM 2.5 exposure was

Countermeasures in the field of human factors in relation, to pedestrian behaviour, regulations, and law enforcement.

NARCIS (Netherlands)

Kraay, J.H.

1974-01-01

Existing measures and theoretical studies on pedestrian behavior, regulations, and enforcement are reviewed. Specific consideration is given to: types of crossings; traffic signal effectiveness; bridges and subways; traffic training areas for children; characteristic movements of pedestrian

Eps homology domain endosomal transport proteins differentially localize to the neuromuscular junction

Directory of Open Access Journals (Sweden)

Mate Suzanne E

2012-09-01

Full Text Available Abstract Background Recycling of endosomes is important for trafficking and maintenance of proteins at the neuromuscular junction (NMJ. We have previously shown high expression of the endocytic recycling regulator Eps15 homology domain-containing (EHD1 proteinin the Torpedo californica electric organ, a model tissue for investigating a cholinergic synapse. In this study, we investigated the localization of EHD1 and its paralogs EHD2, EHD3, and EHD4 in mouse skeletal muscle, and assessed the morphological changes in EHD1−/− NMJs. Methods Localization of the candidate NMJ protein EHD1 was assessed by confocal microscopy analysis of whole-mount mouse skeletal muscle fibers after direct gene transfer and immunolabeling. The potential function of EHD1 was assessed by specific force measurement and α-bungarotoxin-based endplate morphology mapping in EHD1−/− mouse skeletal muscle. Results Endogenous EHD1 localized to primary synaptic clefts of murine NMJ, and this localization was confirmed by expression of recombinant green fluorescent protein labeled-EHD1 in murine skeletal muscle in vivo. EHD1−/− mouse skeletal muscle had normal histology and NMJ morphology, and normal specific force generation during muscle contraction. The EHD 1–4 proteins showed differential localization in skeletal muscle: EHD2 to muscle vasculature, EHD3 to perisynaptic regions, and EHD4 to perinuclear regions and to primary synaptic clefts, but at lower levels than EHD1. Additionally, specific antibodies raised against mammalian EHD1-4 recognized proteins of the expected mass in the T. californica electric organ. Finally, we found that EHD4 expression was more abundant in EHD1−/− mouse skeletal muscle than in wild-type skeletal muscle. Conclusion EHD1 and EHD4 localize to the primary synaptic clefts of the NMJ. Lack of obvious defects in NMJ structure and muscle function in EHD1−/− muscle may be due to functional compensation by other EHD paralogs.

TBC-8, a Putative RAB-2 GAP, Regulates Dense Core Vesicle Maturation in Caenorhabditis elegans

Science.gov (United States)

Hannemann, Mandy; Sasidharan, Nikhil; Hegermann, Jan; Kutscher, Lena M.; Koenig, Sabine; Eimer, Stefan

2012-01-01

Dense core vesicles (DCVs) are thought to be generated at the late Golgi apparatus as immature DCVs, which subsequently undergo a maturation process through clathrin-mediated membrane remodeling events. This maturation process is required for efficient processing of neuropeptides within DCVs and for removal of factors that would otherwise interfere with DCV release. Previously, we have shown that the GTPase, RAB-2, and its effector, RIC-19, are involved in DCV maturation in Caenorhabditis elegans motoneurons. In rab-2 mutants, specific cargo is lost from maturing DCVs and missorted into the endosomal/lysosomal degradation route. Cargo loss could be prevented by blocking endosomal delivery. This suggests that RAB-2 is involved in retention of DCV components during the sorting process at the Golgi-endosomal interface. To understand how RAB-2 activity is regulated at the Golgi, we screened for RAB-2–specific GTPase activating proteins (GAPs). We identified a potential RAB-2 GAP, TBC-8, which is exclusively expressed in neurons and which, when depleted, shows similar DCV maturation defects as rab-2 mutants. We could demonstrate that RAB-2 binds to its putative GAP, TBC-8. Interestingly, TBC-8 also binds to the RAB-2 effector, RIC-19. This interaction appears to be conserved as TBC-8 also interacted with the human ortholog of RIC-19, ICA69. Therefore, we propose that a dynamic ON/OFF cycling of RAB-2 at the Golgi induced by the GAP/effector complex is required for proper DCV maturation. PMID:22654674

Acid sphingomyelinase activity is regulated by membrane lipids and facilitates cholesterol transfer by NPC2.

Science.gov (United States)

Oninla, Vincent O; Breiden, Bernadette; Babalola, Jonathan O; Sandhoff, Konrad

2014-12-01

During endocytosis, membrane components move to intraluminal vesicles of the endolysosomal compartment for digestion. At the late endosomes, cholesterol is sorted out mainly by two sterol-binding proteins, Niemann-Pick protein type C (NPC)1 and NPC2. To study the NPC2-mediated intervesicular cholesterol transfer, we developed a liposomal assay system. (Abdul-Hammed, M., B. Breiden, M. A. Adebayo, J. O. Babalola, G. Schwarzmann, and K. Sandhoff. 2010. Role of endosomal membrane lipids and NPC2 in cholesterol transfer and membrane fusion. J. Lipid Res. 51: 1747-1760.) Anionic lipids stimulate cholesterol transfer between liposomes while SM inhibits it, even in the presence of anionic bis(monoacylglycero)phosphate (BMP). Preincubation of vesicles containing SM with acid sphingomyelinase (ASM) (SM phosphodiesterase, EC 3.1.4.12) results in hydrolysis of SM to ceramide (Cer), which enhances cholesterol transfer. Besides SM, ASM also cleaves liposomal phosphatidylcholine. Anionic phospholipids derived from the plasma membrane (phosphatidylglycerol and phosphatidic acid) stimulate SM and phosphatidylcholine hydrolysis by ASM more effectively than BMP, which is generated during endocytosis. ASM-mediated hydrolysis of liposomal SM was also stimulated by incorporation of diacylglycerol (DAG), Cer, and free fatty acids into the liposomal membranes. Conversely, phosphatidylcholine hydrolysis was inhibited by incorporation of cholesterol, Cer, DAG, monoacylglycerol, and fatty acids. Our data suggest that SM degradation by ASM is required for physiological secretion of cholesterol from the late endosomal compartment, and is a key regulator of endolysosomal lipid digestion. Copyright © 2014 by the American Society for Biochemistry and Molecular Biology, Inc.

TBC-8, a putative RAB-2 GAP, regulates dense core vesicle maturation in Caenorhabditis elegans.

Directory of Open Access Journals (Sweden)

Mandy Hannemann

Full Text Available Dense core vesicles (DCVs are thought to be generated at the late Golgi apparatus as immature DCVs, which subsequently undergo a maturation process through clathrin-mediated membrane remodeling events. This maturation process is required for efficient processing of neuropeptides within DCVs and for removal of factors that would otherwise interfere with DCV release. Previously, we have shown that the GTPase, RAB-2, and its effector, RIC-19, are involved in DCV maturation in Caenorhabditis elegans motoneurons. In rab-2 mutants, specific cargo is lost from maturing DCVs and missorted into the endosomal/lysosomal degradation route. Cargo loss could be prevented by blocking endosomal delivery. This suggests that RAB-2 is involved in retention of DCV components during the sorting process at the Golgi-endosomal interface. To understand how RAB-2 activity is regulated at the Golgi, we screened for RAB-2-specific GTPase activating proteins (GAPs. We identified a potential RAB-2 GAP, TBC-8, which is exclusively expressed in neurons and which, when depleted, shows similar DCV maturation defects as rab-2 mutants. We could demonstrate that RAB-2 binds to its putative GAP, TBC-8. Interestingly, TBC-8 also binds to the RAB-2 effector, RIC-19. This interaction appears to be conserved as TBC-8 also interacted with the human ortholog of RIC-19, ICA69. Therefore, we propose that a dynamic ON/OFF cycling of RAB-2 at the Golgi induced by the GAP/effector complex is required for proper DCV maturation.

Modeling the Short-Term Effect of Traffic and Meteorology on Air Pollution in Turin with Generalized Additive Models

Directory of Open Access Journals (Sweden)

Pancrazio Bertaccini

2012-01-01

Full Text Available Vehicular traffic plays an important role in atmospheric pollution and can be used as one of the key predictors in air-quality forecasting models. The models that can account for the role of traffic are especially valuable in urban areas, where high pollutant concentrations are often observed during particular times of day (rush hour and year (winter. In this paper, we develop a generalized additive models approach to analyze the behavior of concentrations of nitrogen dioxide (NO2, and particulate matter (PM10, collected at the environmental monitoring stations distributed throughout the city of Turin, Italy, from December 2003 to April 2005. We describe nonlinear relationships between predictors and pollutants, that are adjusted for unobserved time-varying confounders. We examine several functional forms for the traffic variable and find that a simple form can often provide adequate modeling power. Our analysis shows that there is a saturation effect of traffic on NO2, while such saturation is less evident in models linking traffic to PM10 behavior, having adjusted for meteorological covariates. Moreover, we consider the proposed models separately by seasons and highlight similarities and differences in the predictors’ partial effects. Finally, we show how forecasting can help in evaluating traffic regulation policies.

Peroxisomes, lipid droplets, and endoplasmic reticulum "hitchhike" on motile early endosomes.

Science.gov (United States)

Guimaraes, Sofia C; Schuster, Martin; Bielska, Ewa; Dagdas, Gulay; Kilaru, Sreedhar; Meadows, Ben R A; Schrader, Michael; Steinberg, Gero

2015-12-07

Intracellular transport is mediated by molecular motors that bind cargo to be transported along the cytoskeleton. Here, we report, for the first time, that peroxisomes (POs), lipid droplets (LDs), and the endoplasmic reticulum (ER) rely on early endosomes (EEs) for intracellular movement in a fungal model system. We show that POs undergo kinesin-3- and dynein-dependent transport along microtubules. Surprisingly, kinesin-3 does not colocalize with POs. Instead, the motor moves EEs that drag the POs through the cell. PO motility is abolished when EE motility is blocked in various mutants. Most LD and ER motility also depends on EE motility, whereas mitochondria move independently of EEs. Covisualization studies show that EE-mediated ER motility is not required for PO or LD movement, suggesting that the organelles interact with EEs independently. In the absence of EE motility, POs and LDs cluster at the growing tip, whereas ER is partially retracted to subapical regions. Collectively, our results show that moving EEs interact transiently with other organelles, thereby mediating their directed transport and distribution in the cell. © 2015 Guimaraes et al.

Avian Influenza Virus Infection of Immortalized Human Respiratory Epithelial Cells Depends upon a Delicate Balance between Hemagglutinin Acid Stability and Endosomal pH.

Science.gov (United States)

Daidoji, Tomo; Watanabe, Yohei; Ibrahim, Madiha S; Yasugi, Mayo; Maruyama, Hisataka; Masuda, Taisuke; Arai, Fumihito; Ohba, Tomoyuki; Honda, Ayae; Ikuta, Kazuyoshi; Nakaya, Takaaki

2015-04-24

The highly pathogenic avian influenza (AI) virus, H5N1, is a serious threat to public health worldwide. Both the currently circulating H5N1 and previously circulating AI viruses recognize avian-type receptors; however, only the H5N1 is highly infectious and virulent in humans. The mechanism(s) underlying this difference in infectivity remains unclear. The aim of this study was to clarify the mechanisms responsible for the difference in infectivity between the current and previously circulating strains. Primary human small airway epithelial cells (SAECs) were transformed with the SV40 large T-antigen to establish a series of clones (SAEC-Ts). These clones were then used to test the infectivity of AI strains. Human SAEC-Ts could be broadly categorized into two different types based on their susceptibility (high or low) to the viruses. SAEC-T clones were poorly susceptible to previously circulating AI but were completely susceptible to the currently circulating H5N1. The hemagglutinin (HA) of the current H5N1 virus showed greater membrane fusion activity at higher pH levels than that of previous AI viruses, resulting in broader cell tropism. Moreover, the endosomal pH was lower in high susceptibility SAEC-T clones than that in low susceptibility SAEC-T clones. Taken together, the results of this study suggest that the infectivity of AI viruses, including H5N1, depends upon a delicate balance between the acid sensitivity of the viral HA and the pH within the endosomes of the target cell. Thus, one of the mechanisms underlying H5N1 pathogenesis in humans relies on its ability to fuse efficiently with the endosomes in human airway epithelial cells. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Aggregation of nanoparticles in endosomes and lysosomes produces surface-enhanced Raman spectroscopy

Science.gov (United States)

Lucas, Leanne J.; Chen, Xiaoke K.; Smith, Aaron J.; Korbelik, Mladen; Zeng, Haishan; Lee, Patrick W. K.; Hewitt, Kevin Cecil

2015-01-01

The purpose of this study was to explore the use of surface-enhanced Raman spectroscopy (SERS) to image the distribution of epidermal growth factor receptor (EGFR) in cells. To accomplish this task, 30-nm gold nanoparticles (AuNPs) tagged with antibodies to EGFR (1012 per mL) were incubated with cells (106 per mL) of the A431 human epidermoid carcinoma and normal human bronchial epithelial cell lines. Using the 632.8-nm excitation line of a He-Ne laser, Raman spectroscopy measurements were performed using a point mapping scheme. Normal cells show little to no enhancement. SERS signals were observed inside the cytoplasm of A431 cells with an overall enhancement of 4 to 7 orders of magnitude. Raman intensity maps of the 1450 and 1583 cm-1 peaks correlate well with the expected distribution of EGFR and AuNPs, aggregated following uptake by endosomes and lysosomes. Spectral features from tyrosine and tryptophan residues dominate the SERS signals.

Traffic noise in Hyderabad city. part I: road traffic noise

International Nuclear Information System (INIS)

Shaikh, G.H.; Shaikh, Q.

2000-01-01

Traffic noise survey was conducted at 20 sites in different areas and localities in Hyderabad city and each site noise data was collected continuously from 0800 to 2000 h. The data was analyzed for L/sub A99/, L/sub A90/, L/sub A50/, L/sub 10/ and L/sub A1/, and approximate values of L/sub Aeq12h/ were evaluated for each site. The results are discussed with reference to some criteria for community annoyance and means and ways to limit high-level traffic noise are suggested. (author)

An MHC-I cytoplasmic domain/HIV-1 Nef fusion protein binds directly to the mu subunit of the AP-1 endosomal coat complex.

Directory of Open Access Journals (Sweden)

Rajendra Kumar Singh

2009-12-01

Full Text Available The down-regulation of the major histocompatibility complex class I (MHC-I from the surface of infected cells by the Nef proteins of primate immunodeficiency viruses likely contributes to pathogenesis by providing evasion of cell-mediated immunity. HIV-1 Nef-induced down-regulation involves endosomal trafficking and a cooperative interaction between the cytoplasmic domain (CD of MHC-I, Nef, and the clathrin adaptor protein complex-1 (AP-1. The CD of MHC-I contains a key tyrosine within the sequence YSQA that is required for down-regulation by Nef, but this sequence does not conform to the canonical AP-binding tyrosine-based motif Yxxphi, which mediates binding to the medium (micro subunits of AP complexes. We previously proposed that Nef allows the MHC-I CD to bind the mu subunit of AP-1 (micro1 as if it contained a Yxxphimotif.Here, we show that a direct interaction between the MHC-I CD/Nef and micro1 plays a primary role in the down-regulation of MHC-I: GST pulldown assays using recombinant proteins indicated that most of the MHC-I CD and Nef residues that are required for the down-regulation in human cells contribute to direct interactions with a truncated version of micro1. Specifically, the tyrosine residue of the YSQA sequence in the MHC-I CD as well as Nef residues E62-65 and P78 each contributed to the interaction between MHC-I CD/Nef and micro1 in vitro, whereas Nef M20 had little to no role. Conversely, residues F172/D174 and V392/L395 of the binding pocket on micro1 for Yxxphi motifs were required for a robust interaction.These data indicate that the MHC-I cytoplasmic domain, Nef, and the C-terminal two thirds of the mu subunit of AP-1 are sufficient to constitute a biologically relevant interaction. The data also reveal an unexpected role for a hydrophobic pocket in micro1 for interaction with MHC-I CD/Nef.

Proactive Traffic Information Control in Emergency Evacuation Network

Directory of Open Access Journals (Sweden)

Zhengfeng Huang

2015-01-01

Full Text Available Traffic demand in emergency evacuation is usually too large to be effectively managed with reactive traffic information control methods. These methods adapt to the road traffic passively by publishing real-time information without consideration of the routing behavior feedback produced by evacuees. Other remedy measures have to be prepared in case of nonrecurring congestion under these methods. To use the network capacity fully to mitigate near-future evacuation traffic congestion, we propose proactive traffic information control (PTIC model. Based on the mechanism between information and routing behavior feedback, this model can change the route choice of evacuees in advance by dissipating strategic traffic information. Generally, the near-future traffic condition is difficult to accurately predict because it is uncertain in evacuation. Assume that the value of traffic information obeys certain distribution within a range, and then real-time traffic information may reflect the most-likely near-future traffic condition. Unlike the real-time information, the proactive traffic information is a selection within the range to achieve a desired level of the road network performance index (total system travel time. In the aspect of the solution algorithm, differential equilibrium decomposed optimization (D-EDO is proposed to compare with other heuristic methods. A field study on a road network around a large stadium is used to validate the PTIC.

Modeling Network Traffic in Wavelet Domain

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Sheng Ma

2004-12-01

Full Text Available This work discovers that although network traffic has the complicated short- and long-range temporal dependence, the corresponding wavelet coefficients are no longer long-range dependent. Therefore, a "short-range" dependent process can be used to model network traffic in the wavelet domain. Both independent and Markov models are investigated. Theoretical analysis shows that the independent wavelet model is sufficiently accurate in terms of the buffer overflow probability for Fractional Gaussian Noise traffic. Any model, which captures additional correlations in the wavelet domain, only improves the performance marginally. The independent wavelet model is then used as a unified approach to model network traffic including VBR MPEG video and Ethernet data. The computational complexity is O(N for developing such wavelet models and generating synthesized traffic of length N, which is among the lowest attained.