The T=1 capsid protein of Penicillium chrysogenum virus is formed by a repeated helix-rich core indicative of gene duplication

et al.Penicillium chrysogenum virus (PcV), a member of the Chrysoviridae family, is a double-stranded RNA (dsRNA) fungal virus with a multipartite genome, with each RNA molecule encapsidated in a separate particle. Chrysoviruses lack an extracellular route and are transmitted during sporogenesis and cell fusion. The PcV capsid, based on a T=1 lattice containing 60 subunits of the 982-amino-acid capsid protein, remains structurally undisturbed throughout the viral cycle, participates in genome metabolism, and isolates the virus genome from host defense mechanisms. Using three-dimensional cryoelectron microscopy, we determined the structure of the PcV virion at 8.0 Å resolution. The capsid protein has a high content of rod-like densities characteristic of α-helices, forming a repeated α-helical core indicative of gene duplication. Whereas the PcV capsid protein has two motifs with the same fold, most dsRNA virus capsid subunits consist of dimers of a single protein with similar folds. The spatial arrangement of the α-helical core resembles that found in the capsid protein of the L-A virus, a fungal totivirus with an undivided genome, suggesting a conserved basic fold. The encapsidated genome is organized in concentric shells; whereas the inner dsRNA shells are well defined, the outermost layer is dense due to numerous interactions with the inner capsid surface, specifically, six interacting areas per monomer. The outermost genome layer is arranged in an icosahedral cage, sufficiently well ordered to allow for modeling of an A-form dsRNA. The genome ordering might constitute a framework for dsRNA transcription at the capsid interior and/or have a structural role for capsid stability. Copyright © 2010, American Society for Microbiology. All Rights Reserved.This work was supported by grants from the Spanish Ministry of Science and Innovation (BFU 2008-02328/BMC and S-0505-Mat-0238 to J.L.C. and BIO2008-02361 to J.R.C.) and the NIH Intramural Research Program with support from the Center for Information Technology.Peer Reviewe

Trapping and patterning of biological objects using photovoltaic tweezers

Photovoltaic tweezers are a recently proposed technique for manipulation and patterning of micro- and nano-objects. It is based in the dielectrophoretic forces associated to the electric fields induced by illumination of certain ferroelectrics due to the bulk photovoltaic effect. The technique has been applied to the patterning of dielectric and metal micro- and nano-particles. In this work, we report the use of photovoltaic tweezers to pattern biological objects on LiNbO3:Fe. Specifically, spores and pollen grains and their nanometric fragments have been trapped and patterned. 1D and 2D arrangements have been achieved by deposition in air or from a hexane suspension. The quality of patterns obtained with nanometric fragments is even better than previous results using photovoltaic tweezers with inorganic micro- and nano-particles. In fact, 1D patterns with a period of 2 μm, almost half of the minimum reported period achieved with photovoltaic tweezers, have been obtained with pollen fragmentsThis work was supported by Spanish projects MAT2011-28379-C03 and MAT2014-57704-C0

Ultrastructural and functional analyses of recombinant influenza virus ribonucleoproteins suggest dimerization of nucleoprotein during virus amplification

8 pages, 7 figures.-- PMID: 10590102 [PubMed].-- PMCID: PMC111524.Full-text version available Open Access at PebMed Central: http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=10590102Influenza virus ribonucleoproteins (RNPs) were reconstituted in vivo from cloned cDNAs expressing the three polymerase subunits, the nucleoprotein (NP), and short template RNAs. The structure of purified RNPs was studied by electron microscopy and image processing. Circular and elliptic structures were obtained in which the NP and the polymerase complex could be defined. Comparison of the structure of RNPs of various lengths indicated that each NP monomer interacts with approximately 24 nucleotides. The analysis of the amplification of RNPs with different lengths showed that those with the highest replication efficiency contained an even number of NP monomers, suggesting that the NP is incorporated as dimers into newly synthesized RNPs.J. Ortega was a fellow of Instituto de Estudios Turolenses. This work was supported by Programa Sectorial de Promoción General del Conocimiento (grants PB97-1160 and PB96-0818).Peer reviewe

The Structure of a Biologically Active Influenza Virus Ribonucleoprotein Complex

The influenza viruses contain a segmented, single-stranded RNA genome of negative polarity. Each RNA segment is encapsidated by the nucleoprotein and the polymerase complex into ribonucleoprotein particles (RNPs), which are responsible for virus transcription and replication. Despite their importance, information about the structure of these RNPs is scarce. We have determined the three-dimensional structure of a biologically active recombinant RNP by cryo-electron microscopy. The structure shows a nonameric nucleoprotein ring (at 12 Å resolution) with two monomers connected to the polymerase complex (at 18 Å resolution). Docking the atomic structures of the nucleoprotein and polymerase domains, as well as mutational analyses, has allowed us to define the interactions between the functional elements of the RNP and to propose the location of the viral RNA. Our results provide the first model for a functional negative-stranded RNA virus ribonucleoprotein complex. The structure reported here will serve as a framework to generate a quasi-atomic model of the molecular machine responsible for viral RNA synthesis and to test new models for virus RNA replication and transcription

A protein with simultaneous capsid scaffolding and dsRNA-binding activities enhances the birnavirus capsid mechanical stability

Viral capsids are metastable structures that perform many essential processes; they also act as robust cages during the extracellular phase. Viruses can use multifunctional proteins to optimize resources (e.g., VP3 in avian infectious bursal disease virus, IBDV). The IBDV genome is organized as ribonucleoproteins (RNP) of dsRNA with VP3, which also acts as a scaffold during capsid assembly. We characterized mechanical properties of IBDV populations with different RNP content (ranging from none to four RNP). The IBDV population with the greatest RNP number (and best fitness) showed greatest capsid rigidity. When bound to dsRNA, VP3 reinforces virus stiffness. These contacts involve interactions with capsid structural subunits that differ from the initial interactions during capsid assembly. Our results suggest that RNP dimers are the basic stabilization units of the virion, provide better understanding of multifunctional proteins, and highlight the duality of RNP as capsidstabilizing and genetic information platformsThis work was supported by grants from the Spanish Ministry of Economy and Competitivity (FIS2011-29493 to PJP, BFU2011-29038 to JLC and BFU2014-55475R to JRC) and Comunidad Autónoma de Madrid (S2013/MIT-2850 to JLC and S2013/MIT-2807 to JRC

Manejo perioperatorio de fármacos anticoagulantes y antiagregantes en el paciente con fractura de cadera

La fractura de cadera eleva de manera significativa en los estudios publicados la morbilidad y la mortalidad en los pacientes ancianos que la padecen. La participación en el proceso de fármacos que alteran la hemostasia se ha convertido en uno de los aspectos más importantes y controvertidos del manejo perioperatorio del paciente con fractura de cadera. La anticoagulación o antiagregación plaquetaria de estos pacientes tiene unas implicaciones tanto en la práctica anestésica como quirúrgica que hacen necesaria la búsqueda de un equilibrio entre el riesgo hemorrágico y trombótico de los mismos. Con la elaboración del presente protocolo, en línea con las últimas evidencias científicas publicadas, se sientan las bases para avanzar en el tratamiento de este grupo de pacientes. Una atención multidisciplinar especializada, una fijación precoz de la fractura antes de las 48 horas, y un efectivo programa de rehabilitación pueden disminuir las complicaciones del proceso.Hip fracture rises significantly in published studies morbidity and mortality in elderly patients. Participation in the process of hemostasis-altering drugs has become one of the most important and controversial perioperative management of patients with hip fracture aspects. Anticoagulation or antiplatelet of these patients have some implications as a surgical anesthetic practice that make it necessary to seek a balance between bleeding and thrombotic risk thereof. With the development of this protocol, in line with the latest scientific evidence published, the stage is set to advance the treatment of this patient group. A specialized multidisciplinary care, early fracture fixation within 48 hours and an effective rehabilitation program can reduce complications of that process

Characterization of interaction of magnetic nanoparticles with breast cancer cells

Background: Different superparamagnetic iron oxide nanoparticles have been tested for their potential use in cancer treatment, as they enter into cells with high effectiveness, do not induce cytotoxicity, and are retained for relatively long periods of time inside the cells. We have analyzed the interaction, internalization and biocompatibility of dimercaptosuccinic acid-coated superparamagnetic iron oxide nanoparticles with an average diameter of 15 nm and negative surface charge in MCF-7 breast cancer cells. Results: Cells were incubated with dimercaptosuccinic acid-coated superparamagnetic iron oxide nanoparticles for different time intervals, ranging from 0.5 to 72 h. These nanoparticles showed efficient internalization and relatively slow clearance. Time-dependent uptake studies demonstrated the maximum accumulation of dimercaptosuccinic acid-coated superparamagnetic iron oxide nanoparticles after 24 h of incubation, and afterwards they were slowly removed from cells. Superparamagnetic iron oxide nanoparticles were internalized by energy dependent endocytosis and localized in endosomes. Transmission electron microscopy studies showed macropinocytosis uptake and clathrin-mediated internalization depending on the nanoparticles aggregate size. MCF-7 cells accumulated these nanoparticles without any significant effect on cell morphology, cytoskeleton organization, cell cycle distribution, reactive oxygen species generation and cell viability, showing a similar behavior to untreated control cells. Conclusions: All these findings indicate that dimercaptosuccinic acid-coated superparamagnetic iron oxide nanoparticles have excellent properties in terms of efficiency and biocompatibility for application to target breast cancer cellsThe research leading to these results have received partial funding from the European Seventh Framework Programme (FP7/2007-2013) under the project MULTIFUN grant agreement no. 262943, and the project Nanofrontmag-CM (S2013/MIT-2850) from the Comunidad de Madrid. Additional grants were obtained from BFU 2011–29038 and CTQ2013-48767-C3-3-R from the Ministerio de Economia y Competitividad and S2009/Mat 1507 from the Comunidad de Madrid (to JLC), from EU FP7 project NAMDIATREAM (ref 246479) and from “la Caixa” / CNB International PhD Programme Fellowship

Sesión científica celebrada el 30 de noviembre de 2017 para conmemorar los premios Nobel 2017 en Fisiología o Medicina y en Química

Un año más, la Real Academia Nacional de Farmacia se reúne en Sesión Científica para conmemorar los Premios Nobel correspondientes a las especialidades de Fisiología o Medicin

Depresión del sistema mononuclear-fagocítico provocada por altas dosis de morfínico

We evaluated in human monocytes the effect of high doses of alfentanyl on the expression of vimentin filaments, the phagocytic activity and the membrane display of HLA-DR molecules in the subjects undergoing surgery. The study was performed on 30 patients, ASAI-II. The patients received 100 mcg/kg i.v. of Alfentanil and the maintenance of anaesthesia was made with Alfentanil (2-3 mcg/kg/min.). The patients were randomized in two groups. The patients were ventilated with N2O:O2 (1:1) (Group I) or air: O2 (1:1) (Group II). After surgery, all patients of the Group II received Naloxone (0.2-0.4 mg). Central venous blood samples were obtained before induction, one and two hours after induction of anaesthesia and at the end of surgery. Separation of monocytes was performed according to Boyum technique. CD35 and HLA-DR molecules and vimentin filaments were studied by indirect immunofluorescence method using monoclonal antibodies. Percentage of positive cells were read with a cytofluorometer. The phagocytic function of monocytes was determined by ingestion of latex particles. Cortisol and ACTH plasma levels were determined by RIA. High doses of Alfentanyl depress phagocytic function and membrane display of CD35 and HLA-DR molecules in monocyte and induce marked changes in the organization of vimentin filaments in these cells in patients undergoing surgery. This monocytic depression was more marked in the patients ventilated with N2O. In our results there was uninhibition of ACTH and cortisol plasma levels responses to surgical stress by Alfentanil administration. Since the effects of Alfentanil were reversed by Naloxone, an opioid receptor mechanism seems to mediate these event

Study of antihypertensive drugs prescribed in old people

Introducción: El control de la hipertensión arterial enlentece o evita la aparición de lesiones en órganos diana. Hay pocos estudios que muestren si los tratamientos antihipertensivos cumplen las normas de las Guías, en los Centros de Salud. Pacientes y Métodos: Se realizó un estudio descriptivo transversal, sobre la prescripción de antihipertensivos en mayores de un Centro de Salud, con población de referencia de 25000 habitantes. Se seleccionaron al azar 220 historias, recogiendo los datos clínicos desde el 14 al 25 de noviembre de 2011. Se analizó si los antihipertensivos utilizados concordaban con las indicaciones de las Guías Terapéuticas. Resultados:Lo más utilizado es la combinación de diuréticos tiazídicos con IECAs, seguido de IECAs en monoterapia. Muchas de las asociaciones no tienen evidencias. Los registros en la historia son insuficientes. Discusión: En muchos casos no se puede conocer la idoneidad del fármaco por falta de registros en la historia clínica. Falta un cronograma de seguimiento protocolizado para el control del paciente hipertenso.Introduction: The control of hypertension slows or prevents the onset of target organ damage. There are few studies that show whether antihypertensive treatments meet the standards of the Guidelines.Objectives: The objective of this project is to estimate the validity of this initiative in the Centres where it has already started, and also to introduce this initiative to the students of the Master degree of Secondary School, through a training workshop. Materials and methods: We carried-out a descriptive, cross-sectional study on the prescription of anti-hypertensive drugs to aged people. This study was carried out in a Community Health Centre with a reference population of 25,000 inhabitants. Cases: 220 randomly selected stories, collecting clinical data from 14 to 25 November 2011. We analyzed whether antihypertensive drugs used were consistent with the indications of the Guidelines. Results: The most used is the combination of thiazide diuretics with ACE inhibitors, followed by ACE inhibitors alone. Many of the associations have no evidence. The history records are inadequate. Conclusions: In many cases we can not know the suitability of the drug due to lack of records in the medical record. Missing protocolized follow-up schedule for the control of hypertensive patients