Specific Rab GTPase-activating proteins define the Shiga toxin and epidermal growth factor uptake pathways

Rab family guanosine triphosphatases (GTPases) together with their regulators define specific pathways of membrane traffic within eukaryotic cells. In this study, we have investigated which Rab GTPase-activating proteins (GAPs) can interfere with the trafficking of Shiga toxin from the cell surface to the Golgi apparatus and studied transport of the epidermal growth factor (EGF) from the cell surface to endosomes. This screen identifies 6 (EVI5, RN-tre/USP6NL, TBC1D10A–C, and TBC1D17) of 39 predicted human Rab GAPs as specific regulators of Shiga toxin but not EGF uptake. We show that Rab43 is the target of RN-tre and is required for Shiga toxin uptake. In contrast, RabGAP-5, a Rab5 GAP, was unique among the GAPs tested and reduced the uptake of EGF but not Shiga toxin. These results suggest that Shiga toxin trafficking to the Golgi is a multistep process controlled by several Rab GAPs and their target Rabs and that this process is discrete from ligand-induced EGF receptor trafficking

Functional dissection of Rab GTPases involved in primary cilium formation

Primary cilia are sensory structures involved in morphogen signalling during development, liquid flow in the kidney, mechanosensation, sight, and smell (Badano, J.L., N. Mitsuma, P.L. Beales, and N. Katsanis. 2006. Annu. Rev. Genomics Hum. Genet. 7:125–148; Singla, V., and J.F. Reiter. 2006. Science. 313:629–633.). Mutations that affect primary cilia are responsible for several diseases, including neural tube defects, polycystic kidney disease, retinal degeneration, and cancers (Badano et al., 2006; Singla and Reiter, 2006). Primary cilia formation and function requires tight integration of the microtubule cytoskeleton with membrane trafficking (Singla and Reiter, 2006), and this is poorly understood. We show that the Rab GTPase membrane trafficking regulators Rab8a, -17, and -23, and their cognate GTPase-activating proteins (GAPs), XM_037557, TBC1D7, and EVI5like, are involved in primary cilia formation. However, other human Rabs and GAPs are not. Additionally, Rab8a specifically interacts with cenexin/ODF2, a basal body and microtubule binding protein required for cilium biogenesis (Ishikawa, H., A. Kubo, S. Tsukita, and S. Tsukita. 2005. Nat. Cell Biol. 7:517–524), and is the sole Rab enriched at primary cilia. These findings provide a basis for understanding how specific membrane trafficking pathways cooperate with the microtubule cytoskeleton to give rise to the primary cilia

Minocycline fails to modulate cerebrospinal fluid HIV infection or immune activation in chronic untreated HIV-1 infection: results of a pilot study

<p>Abstract</p> <p>Background</p> <p>Minocycline is a tetracycline antibiotic that has been shown to attenuate central nervous system (CNS) lentivirus infection, immune activation, and brain injury in model systems. To initiate assessment of minocycline as an adjuvant therapy in human CNS HIV infection, we conducted an open-labelled pilot study of its effects on cerebrospinal fluid (CSF) and blood biomarkers of infection and immune responses in 7 viremic subjects not taking antiretroviral therapy.</p> <p>Results</p> <p>There were no discernable effects of minocycline on CSF or blood HIV-1 RNA, or biomarkers of immune activation and inflammation including: CSF and blood neopterin, CSF CCL2, CSF white blood cell count, and expression of cell-surface activation markers on CSF and blood T lymphocytes and monocytes.</p> <p>Conclusions</p> <p>This pilot study of biological responses to minocycline suggests little potential for its use as adjunctive antiviral or immunomodulating therapy in chronic untreated HIV infection.</p

Towards a comprehensive understanding of the structural dynamics of a bacterial diterpene synthase during catalysis

Terpenes constitute the largest and structurally most diverse natural product family. Most terpenoids exhibit a stereochemically complex macrocyclic core, which is generated by C–C bond forming of aliphatic oligo-prenyl precursors. This reaction is catalysed by terpene synthases (TPSs), which are capable of chaperoning highly reactive carbocation intermediates through an enzyme-specific reaction. Due to the instability of carbocation intermediates, the proteins’ structural dynamics and enzyme:substrate interactions during TPS catalysis remain elusive. Here, we present the structure of the diterpene synthase CotB2, in complex with an in crystallo cyclised abrupt reaction product and a substrate-derived diphosphate. We captured additional snapshots of the reaction to gain an overview of CotB2’s catalytic mechanism. To enhance insights into catalysis, structural information is augmented with multiscale molecular dynamic simulations. Our data represent fundamental TPS structure dynamics during catalysis, which ultimately enable rational engineering towards tailored terpene macrocycles that are inaccessible by conventional chemical synthesis

Chemical Synthesis of the Fluorescent, Cyclic Dinucleotides c(th)GAMP

The cGAS-STING pathway is known for its role in sensing cytosolic DNA introduced by a viral infection, bacterial invasion or tumorigenesis. Free DNA is recognized by the cyclic GMP-AMP synthase (cGAS) catalyzing the production of 2',3'-cyclic guanosine monophosphate-adenosine monophosphate (2',3'-cGAMP) in mammals. This cyclic dinucleotide acts as a second messenger, activating the stimulator of interferon genes (STING) that finally triggers the transcription of interferon genes and inflammatory cytokines. Due to the therapeutic potential of this pathway, both the production and the detection of cGAMP via fluorescent moieties for assay development is of great importance. Here, we introduce the paralleled synthetic access to the intrinsically fluorescent, cyclic dinucleotides 2'3'-c(th)GAMP and 3'3'-c(th)GAMP based on phosphoramidite and phosphate chemistry, adaptable for large scale synthesis. We examine their binding properties to murine and human STING and confirm biological activity including interferon induction by 2'3'-c(th)GAMP in THP-1 monocytes. Two-photon imaging revealed successful cellular uptake of 2'3'-c(th)GAMP in THP-1 cells

Raltegravir treatment intensification does not alter cerebrospinal fluid HIV-1 infection or immunoactivation in subjects on suppressive therapy

Background. Despite suppression of plasma human immunodeficiency virus type 1 (HIV-1) RNA by antiretroviral therapy to levels below clinical assay detection, infection and immune activation may persist within the central nervous system and possibly lead to continued brain injury. We hypothesized that intensifying therapy would decrease cerebrospinal fluid (CSF) infection and immune activation. Methods. This was a 12-week, randomized, open-label pilot study comparing addition of the integrase inhibitor raltegravir to no treatment augmentation, with an option for rollover to raltegravir. CSF and plasma were analyzed for HIV-1 RNA using a single-copy assay. CSF and blood immune activation was assessed by neopterin concentrations and CD4 1 and CD8 1 T-cell surface antigen expression. Results. Primary analysis compared 14 intensified (including rollovers) to 9 nonintensified subject experiences. Median HIV-1 RNA levels in all samples were lower in CSF (,.3 copies/mL) than in plasma (,.9 copies/mL; P , .0001), and raltegravir did not reduce HIV-1 RNA, CSF neopterin, or CD4 1 and CD8 1 T-cell activation. Conclusions. Raltegravir intensification did not reduce intrathecal immunoactivation or alter CSF HIV-1 RNA levels in subjects with baseline viral suppression. With and without raltegravir intensification, HIV RNA levels in CSF were very low in the enrolled subjects. Clinical Trials Registration. ClinicalTrials.gov (NCT00672932)

Umsetzung der Energiestrategie 2050 - Band II : Neue Ansätze für Staat und Wirtschaft

In diesem, dem zweiten Band von "Energy Governance" befassen sich die Autoren aus vielfältiger Perspektive mit zahlreichen Fragen zur Umsetzung der Energiestrategie 2050. Sie identifizieren jene Fälle von Solaranlagen, bei denen der Ersatz einer Baubewilligung zu Rechtsunsicherheit führt. Sie untersuchen die Wirkung moderner Finanzberichterstattung auf die Politik der kantonalen Beteiligungen an Energieversorgungsunternehmen in einer Marktumgebung tiefer Strompreis. Analysiert wird weiter die staatliche Förderung der erneuerbaren Stromproduktion im Hinblick auf ihre Wirkung. Eine kritische Analyse der "Smart City"-Projekte in der Schweiz und in den Nachbarländern Deutschland und Österreich führt zu Empfehlungen, wie solche Projekte effizienter geplant und umgesetzt werden können. Auch die Beziehung öffentlicher Organisationen zu ihrer "Stakeholder-Umwelt" wird behandelt

Umsetzung der Energiestrategie 2050: Herausforderungen und Chancen für Staat und Wirtschaft

Sammelband der Reihe "Energy Governance Working Paper" Nr. 1 bis 7Die Energiestrategie 2050 des Bundes definiert anspruchsvolle Ziele. Für deren Erreichung hat der Bundesrat daher unter anderem den Aktionsplan Energieforschung ins Leben gerufen. Dazu wurden acht sogenannte SCCERs, Swiss Competence Center for Energy Research, initiiert, in denen hochschulübergreifend angewandte Energie-Forschung betrieben wird. Die Zürcher Hochschule für Angewandte Wissenschaften (ZHAW) ist an vier dieser acht SCCERs aktiv beteiligt. Die ZHAW hat diese Aufgabe zum Anlass genommen, Energieforschung zum strategischen Schwerpunkt der gesamten Fachhochschule zu erklären und in allen Departementen Kompetenzaufbauprojekte zu starten. Der vorliegende Sammelband präsentiert die ersten Ergebnisse dieser Kompetenzaufbauprojekte an der School of Management and Law, wobei zwei dieser Projekte in Zusammenarbeit mit Forschern aus den Departementen Angewandte Linguistik und School of Engineering erfolgten. Dabei wurden die Herausforderungen und Chancen, die sich für Staat und Wirtschaft aus der Umsetzung der Energiestrategie 2050 ableiten, auf verschiedenen Ebenen betrachtet: die Schweiz im internationalen Vergleich, Besonderheiten der Führung von EVUs, rechtliche und ökonomische Rahmenbedingungen und die Gestaltung der Energie-Zukunft in Schweizer Städten

Defectos del desarrollo del esmalte en dentición temporaria: Su relación con caries y el estado nutricional en niños menores de 6 años de edad en Mendoza, República Argentina

Objective: to establish the relationships between the presence of Enamel development defects (EDD) in the temporary dentition, the Early Childhood Caries (ECC) and the nutritional status of children attending two centers for the prevention and treatment of child malnutrition, in Mendoza, Rep. Argentina. Materials and method: On a total of 307 children between 12 and 71 months of age (151 eutrophic and 156 with child malnutrition), with prior parental consent, dmft and dmfs were recorded according to ICDAS II categories of active caries 2 to 6, and presence/absence of EDD. Frequency distributions and nonparametric association tests with p≤ 0.05 were determined. Results: 47.6% of the children studied presented EDD, being more prevalent in children with malnutrition (X2=13.063; p= 0.00). A statistically significant association was recorded between the presence of DDE and dmfs (Mann Whitney U= 9945, p=0.015) and dmfs (Mann Whitney U= 10044.5, p=0.021). In the children who presented DDE (n=146), the malnourished group presented dmft values higher than the eutrophic group (Mann Whitney U=2034, p= 0.044). Conclusions: EDD were more prevalent in children with malnutrition, and were positively related in this study group with ECC.Objetivo: establecer las relaciones entre la presencia Defectos del Desarrollo del Esmalte (DDE) en dentición temporaria, la Caries de Infancia Temprana (CIT) y el estado nutricional en niños asistentes a dos centros de prevención y tratamiento de la desnutrición infantil, en Mendoza, Rep. Argentina. Materiales y método: Sobre un total de 307 niños entre 12 y 71 meses de edad (151 eutróficos y 156 con desnutrición infantil), previo consentimiento parental, se registraron ceod y ceos según categorías de caries activas ICDAS II 2 a 6, y presencia/ausencia de DDE. Se determinaron las distribuciones de frecuencias y pruebas de asociación no paramétricas con p≤ 0,05. Resultados: el 47.6% de los niños estudiados presentó DDE, siendo más prevalente en niños con desnutrición (X2=13.063; p= 0,00). Se registró una asociación estadísticamente significativa entre la presencia de DDE y ceod (U de Mann Whitney= 9945, p=0,015) y ceos (U de Mann Whitney= 10044,5, p=0,021). En los niños que presentaron DDE (n=146), el grupo con desnutrición presentó valores de ceod mayores que el eutrófico (U de Mann Whitney=2034, p= 0.044). Conclusiones: los DDE fueron más prevalentes en los niños con desnutrición, y se relacionaron positivamente en este grupo de estudio con CIT