tRNA-linked molecular beacons for imaging mRNAs in the cytoplasm of living cells

When oligonucleotide probes are microinjected into cells to image the distribution of RNAs, they are rapidly sequestered into the nucleus. As a result, it is difficult to detect mRNAs in the cytoplasm of living cells. We were able to overcome this process by attaching tRNA transcripts to the probes. We show that when fluorescently labeled tRNAs, tRNAs with extensions at their 5′ end, or chimeric molecules in which a molecular beacon possessing a 2′-O-methylribonucleotide backbone is linked to a tRNA, are injected into the nucleus of HeLa cells, they are exported into the cytoplasm. When these constructs are introduced into the cytoplasm, they remain cytoplasmic. These constructs allow the distribution of both the general mRNA population and specific mRNAs to be imaged in living cells. This strategy should also be useful for enhancing the efficacy of antisense oligonucleotides by keeping them in the cytoplasm. Our observations show that the fidelity of the tRNA export system is relaxed for unnatural tRNA variants when they are introduced into the nucleus in large amounts

Stochastic mRNA Synthesis in Mammalian Cells

Individual cells in genetically homogeneous populations have been found to express different numbers of molecules of specific proteins. We investigated the origins of these variations in mammalian cells by counting individual molecules of mRNA produced from a reporter gene that was stably integrated into the cell's genome. We found that there are massive variations in the number of mRNA molecules present in each cell. These variations occur because mRNAs are synthesized in short but intense bursts of transcription beginning when the gene transitions from an inactive to an active state and ending when they transition back to the inactive state. We show that these transitions are intrinsically random and not due to global, extrinsic factors such as the levels of transcriptional activators. Moreover, the gene activation causes burst-like expression of all genes within a wider genomic locus. We further found that bursts are also exhibited in the synthesis of natural genes. The bursts of mRNA expression can be buffered at the protein level by slow protein degradation rates. A stochastic model of gene activation and inactivation was developed to explain the statistical properties of the bursts. The model showed that increasing the level of transcription factors increases the average size of the bursts rather than their frequency. These results demonstrate that gene expression in mammalian cells is subject to large, intrinsically random fluctuations and raise questions about how cells are able to function in the face of such noise

Identificación y caracterización de las brechas entre educación básica e instituciones públicas de educación superior

La Pontificia Universidad Católica del Perú (PUCP), a través INNOVAPUCP y el CISE, llevó a cabo el servicio de consultoría para el Programa para la Mejora de la Calidad y Pertinencia de los servicios de Educación Superior Universitaria y Tecnológica (PMESUT) entre los meses de julio y diciembre de 2020. Cabe mencionar que participaron, como consultores, especialistas pertenecientes a diversas unidades, departamentos y centros de nuestra casa superior de estudios. Los términos de referencia contemplaron dos grandes objetivos:• Determinar en qué medida una prueba de acceso a la educación superior podría contribuir a superar la problemática de las brechas existentes entre estos dos niveles comprendidos: educación básica regular (EBR) y educación superior. Esto en atención al tipo de prueba, sus referentes o insumos, sus implicancias, los impactos favorables e indeseados que pudieran producirse en desmedro del currículo de la educación básica, las posibles consecuencias en la educación superior al contar con un mejor instrumento de selección y los aspectos legales asociados a una prueba o conjunto de pruebas de esta naturaleza.• Determinar el potencial de una prueba o conjunto de pruebas de acceso a la educación superior para ofrecer insumos de relevancia para reflexionar y tomar decisiones acercade la ejecución curricular en la EBR. En otras palabras, esta información, tal como ocurreen otros países, bien podría servir como referente importante para los procesos de estudio, análisis y balances respecto a las competencias que se logran al terminar el quinto de secundaria en este país.Desde el punto de vista operativo, se avanzó en relación con cada uno de los siguientes objetivos específicos, sobre los que se estructuró el resumen del informe final:• Caracterizar los estudios nacionales e internacionales relacionados con el análisis de brechas entre la educación básica y la educación superior.• Comparar las competencias de egreso de la educación básica con los perfiles de ingreso de las instituciones públicas de educación superior (universidades e institutos).• Analizar las pruebas de admisión a las instituciones públicas de educación superior desde la óptica de sus contenidos y condicionamientos a la educación básica. • Caracterizar las percepciones de los diferentes actores (estudiantes, profesores, autoridades e investigadores) relevantes en estos dos niveles acerca de las diferentes brechas y las posibles formas de enfrentar esta situación. • Estimar la pertinencia técnica, legal, pedagógica y social de una prueba estandarizada para el acceso a la educación superior en el Perú. Cristalizar estos objetivos específicos ha supuesto desplegar una serie de técnicas y procedimientos propios de la investigación en el campo de las ciencias sociales, como la revisión de literatura especializada, la sistematización de experiencias previas, los estudios cuantitativos (cuestionarios online) y cualitativos (grupos focales y entrevistas semiestructuradas). Así, de manera específica, el trabajo empírico comprendió a actores relevantes en la problemática mencionada. En cuanto a los estudiantes que participaron de los grupos focales y entrevistas semiestructuradas, se contó con quienes estaban cursando el último año de educación básica, el tercer ciclo o módulo en institutos superiores tecnológicos (IST) y el tercer ciclo o semestre en universidades, todos ellos del sector público. Es relevante acotar que, en cuanto a la educación superior, no se consideró a los que ingresaron en el marco de la pandemia por medio de exámenes virtuales, pues ya se conocían los problemas técnicos y de concepción que acusaban los funcionarios de estas casas de estudios. Además, la situación de excepcionalidad y el reciente aprendizaje configuraron un escenario poco adecuado para estudiar el fenómeno en su complejidad real y en el que actúan, además, de manera más nítida, las academias o centros preuniversitarios. Los profesores de educación básica, de IST y de universidades participaron en grupos focales y entrevistas semiestructuradas. Asimismo, se consideró a los directivos de las instituciones mencionadas y a líderes educativos de cada una de las regiones priorizadas, quienes mantuvieron entrevistas semiestructuradas. De manera adicional, se desarrollaron grupos focales de directivos más profesores de las academias preuniversitarias. Por último, es preciso acotar que todos los despliegues empíricos o que demandaron un trabajo de campo (grupos focales y entrevistas semiestructuradas) transcurrieron de manera virtual, a raíz de la actual coyuntura por el COVID-19 y gracias al accionar de monitores responsables de cada región priorizada

Reporte de Mercados Financieros - tercer trimestre de 2013

El Banco de la República (BR) genera información para la toma de decisiones, la rendición de cuentas y la difusión al público. En particular, el Reporte de Mercados Financieros está enmarcado dentro del principio de difusión al público y contribuye a cumplir con el servicio que presta el Banco de ofrecer información e investigación económica de calidad

Reporte de Mercados Financieros - primer trimestre de 2013

Este reporte hace una descripción de la dinámica del mercado financiero a nivel local, apoyándose en comparaciones internacionales. El entendimiento del comportamiento de los diferentes mercados es esencial para las entidades encargadas de la supervisión, para los participantes de dichos mercados y para las autoridades encargadas de establecer el marco regulatorio de los mismos

Reporte de Mercados Financieros - segundo trimestre de 2013

Este reporte hace una descripción de la dinámica del mercado financiero a nivel local, apoyándose en comparaciones internacionales. El entendimiento del comportamiento de los diferentes mercados es esencial para las entidades encargadas de la supervisión, para los participantes de dichos mercados y para las autoridades encargadas de establecer el marco regulatorio de los mismos

Reporte de Mercados Financieros - cuarto trimestre de 2013

El Banco de la República (BR) genera información para la toma de decisiones, la rendición de cuentas y la difusión al público. En particular, el Reporte de Mercados Financieros está enmarcado dentro del principio de difusión al público y contribuye a cumplir con el servicio que presta el Banco de ofrecer información e investigación económica de calidad

Antimicrobial resistance among migrants in Europe: a systematic review and meta-analysis

BACKGROUND: Rates of antimicrobial resistance (AMR) are rising globally and there is concern that increased migration is contributing to the burden of antibiotic resistance in Europe. However, the effect of migration on the burden of AMR in Europe has not yet been comprehensively examined. Therefore, we did a systematic review and meta-analysis to identify and synthesise data for AMR carriage or infection in migrants to Europe to examine differences in patterns of AMR across migrant groups and in different settings. METHODS: For this systematic review and meta-analysis, we searched MEDLINE, Embase, PubMed, and Scopus with no language restrictions from Jan 1, 2000, to Jan 18, 2017, for primary data from observational studies reporting antibacterial resistance in common bacterial pathogens among migrants to 21 European Union-15 and European Economic Area countries. To be eligible for inclusion, studies had to report data on carriage or infection with laboratory-confirmed antibiotic-resistant organisms in migrant populations. We extracted data from eligible studies and assessed quality using piloted, standardised forms. We did not examine drug resistance in tuberculosis and excluded articles solely reporting on this parameter. We also excluded articles in which migrant status was determined by ethnicity, country of birth of participants' parents, or was not defined, and articles in which data were not disaggregated by migrant status. Outcomes were carriage of or infection with antibiotic-resistant organisms. We used random-effects models to calculate the pooled prevalence of each outcome. The study protocol is registered with PROSPERO, number CRD42016043681. FINDINGS: We identified 2274 articles, of which 23 observational studies reporting on antibiotic resistance in 2319 migrants were included. The pooled prevalence of any AMR carriage or AMR infection in migrants was 25·4% (95% CI 19·1-31·8; I2 =98%), including meticillin-resistant Staphylococcus aureus (7·8%, 4·8-10·7; I2 =92%) and antibiotic-resistant Gram-negative bacteria (27·2%, 17·6-36·8; I2 =94%). The pooled prevalence of any AMR carriage or infection was higher in refugees and asylum seekers (33·0%, 18·3-47·6; I2 =98%) than in other migrant groups (6·6%, 1·8-11·3; I2 =92%). The pooled prevalence of antibiotic-resistant organisms was slightly higher in high-migrant community settings (33·1%, 11·1-55·1; I2 =96%) than in migrants in hospitals (24·3%, 16·1-32·6; I2 =98%). We did not find evidence of high rates of transmission of AMR from migrant to host populations. INTERPRETATION: Migrants are exposed to conditions favouring the emergence of drug resistance during transit and in host countries in Europe. Increased antibiotic resistance among refugees and asylum seekers and in high-migrant community settings (such as refugee camps and detention facilities) highlights the need for improved living conditions, access to health care, and initiatives to facilitate detection of and appropriate high-quality treatment for antibiotic-resistant infections during transit and in host countries. Protocols for the prevention and control of infection and for antibiotic surveillance need to be integrated in all aspects of health care, which should be accessible for all migrant groups, and should target determinants of AMR before, during, and after migration. FUNDING: UK National Institute for Health Research Imperial Biomedical Research Centre, Imperial College Healthcare Charity, the Wellcome Trust, and UK National Institute for Health Research Health Protection Research Unit in Healthcare-associated Infections and Antimictobial Resistance at Imperial College London

Structural Alterations in a Component of Cytochrome c Oxidase and Molecular Evolution of Pathogenic Neisseria in Humans

Three closely related bacterial species within the genus Neisseria are of importance to human disease and health. Neisseria meningitidis is a major cause of meningitis, while Neisseria gonorrhoeae is the agent of the sexually transmitted disease gonorrhea and Neisseria lactamica is a common, harmless commensal of children. Comparative genomics have yet to yield clear insights into which factors dictate the unique host-parasite relationships exhibited by each since, as a group, they display remarkable conservation at the levels of nucleotide sequence, gene content and synteny. Here, we discovered two rare alterations in the gene encoding the CcoP protein component of cytochrome cbb3 oxidase that are phylogenetically informative. One is a single nucleotide polymorphism resulting in CcoP truncation that acts as a molecular signature for the species N. meningitidis. We go on to show that the ancestral ccoP gene arose by a unique gene duplication and fusion event and is specifically and completely distributed within species of the genus Neisseria. Surprisingly, we found that strains engineered to express either of the two CcoP forms conditionally differed in their capacity to support nitrite-dependent, microaerobic growth mediated by NirK, a nitrite reductase. Thus, we propose that changes in CcoP domain architecture and ensuing alterations in function are key traits in successive, adaptive radiations within these metapopulations. These findings provide a dramatic example of how rare changes in core metabolic proteins can be connected to significant macroevolutionary shifts. They also show how evolutionary change at the molecular level can be linked to metabolic innovation and its reversal as well as demonstrating how genotype can be used to infer alterations of the fitness landscape within a single host

Genome Stability of Lyme Disease Spirochetes: Comparative Genomics of Borrelia burgdorferi Plasmids

Lyme disease is the most common tick-borne human illness in North America. In order to understand the molecular pathogenesis, natural diversity, population structure and epizootic spread of the North American Lyme agent, Borrelia burgdorferi sensu stricto, a much better understanding of the natural diversity of its genome will be required. Towards this end we present a comparative analysis of the nucleotide sequences of the numerous plasmids of B. burgdorferi isolates B31, N40, JD1 and 297. These strains were chosen because they include the three most commonly studied laboratory strains, and because they represent different major genetic lineages and so are informative regarding the genetic diversity and evolution of this organism. A unique feature of Borrelia genomes is that they carry a large number of linear and circular plasmids, and this work shows that strains N40, JD1, 297 and B31 carry related but non-identical sets of 16, 20, 19 and 21 plasmids, respectively, that comprise 33–40% of their genomes. We deduce that there are at least 28 plasmid compatibility types among the four strains. The B. burgdorferi ∼900 Kbp linear chromosomes are evolutionarily exceptionally stable, except for a short ≤20 Kbp plasmid-like section at the right end. A few of the plasmids, including the linear lp54 and circular cp26, are also very stable. We show here that the other plasmids, especially the linear ones, are considerably more variable. Nearly all of the linear plasmids have undergone one or more substantial inter-plasmid rearrangements since their last common ancestor. In spite of these rearrangements and differences in plasmid contents, the overall gene complement of the different isolates has remained relatively constant