Study of the functions of mammalian PrimPol protein in vivo

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 18-11-2016Esta tesis tiene embargado el acceso al texto completo hasta el 18-05-2018During eukaryotic DNA replication, the replisome machinery slows down when the template DNA is damaged by endogenous or exogenous elements. To minimize the risk of fork collapse, “DNA damage tolerance” (DDT) mechanisms facilitate progression of DNA replication through damaged templates. A classical DDT response involves the use of DNA polymerases specialized in “translesion synthesis”. An alternative DDT mechanism consists in the reinitiation of DNA synthesis downstream of the damaged DNA, leaving a short unreplicated gap to be repaired post-replicatively. This mechanism requires the repositioning of the replisome and a new priming event. In 2013, mammalian PrimPol protein was identified as the enzyme that mediates fork restart after different types of DNA lesions, including thymine photodimers caused by UV light. PrimPol is a member of the archaeal-eukaryotic primase (AEP) superfamily and is included in a clade of AEP proteins that contain both primase and polymerase activities. Recent studies confirmed a similar role for PrimPol at natural obstacles or difficult-to-replicate sites. Despite these antecedents, the contribution of PrimPol to maintaining genomic stability and its possible protective function against UV-induced DNA damage remained to be investigated. In this thesis we have studied the functions of mouse PrimPol in vivo. In a first approach, we participated in a structure-function study designed to understand the functional importance of several conserved amino acids in the PrimPol family, including two variants presumably associated to pathology: Y89D and Y100H. This study led to the identification of two amino acids located within a conserved WFYY motif that are indispensable for PrimPol functionality. In the second part of the thesis, we have studied the physiological consequences of PrimPol ablation in mammals using a KO mouse model. PrimPol-/- mice are viable but hypersensitive to UV-induced DNA damage. Without PrimPol, skin healing after UV irradiation is compromised due to inefficient cell proliferation in the epidermis, which is necessary to replace cells with damaged DNA. The absence of PrimPol also caused a higher incidence of UV-induced papillomas, gender-specific alopecia, obesity and the development of aggressive tumors. Our results underscore the importance of mammalian PrimPol for the maintenance of genomic stability and strongly suggest a tumor suppression function in vivo.Durante la replicación del DNA eucariótico, la maquinaria replicativa se ralentiza cuando el DNA molde ha sido dañado por causas exógenas o endógenas. Para minimizar el riesgo de colapso en las horquillas de replicación, los mecanismos de tolerancia al daño en el DNA (DDT) facilitan el proceso de replicación a través de moldes dañados. Un mecanismo clásico de DDT implica el uso de DNA polimerasas especializadas en síntesis “translesiva”. Un mecanismo alternativo consiste en la reiniciación de la síntesis de DNA tras la lesión dejando un segmento de DNA sin replicar que será reparado post-replicativamente. Este mecanismo requiere el reposicionamiento del replisoma y un nuevo evento de “primado”. En 2013, la proteína PrimPol humana fue identificada como el enzima responsable del reinicio de las horquillas ante diferentes tipos de lesiones en el DNA, incluyendo fotodímeros de timina causados por la luz ultravioleta (UV). PrimPol pertenece a la superfamilia de primasas de archaea y eukarya (AEP) y se incluye en un clado de proteínas AEP que contienen actividades primasa y polimerasa. Estudios recientes han confirmado una función similar para PrimPol ante obstáculos naturales en el genoma o puntos difíciles de replicar. A pesar de estos antecedentes, la contribución de PrimPol al mantenimiento de la estabilidad genómica y su posible función protectora ante el daño inducido por UV todavía no habían sido investigadas. En esta tesis hemos estudiado las funciones de la proteína PrimPol in vivo. En primer lugar, hemos participado en un estudio de estructura-función diseñado para comprender la relevancia funcional de varios aminoácidos conservados en la familia PrimPol, inlcuyendo dos variantes presuntamente asociadas a patologías humanas: Y89D y Y100H. Este estudio condujo a la identificación de dos aminoácidos, localizados en el motivo conservado WFYY, que son indispensables para la funcionalidad de PrimPol. En la segunda parte de la tesis, hemos estudiado las consecuencias fisiológicas de la pérdida de PrimPol en mamíferos utilizando un modelo KO murino. Los ratones PrimPol-/- son viables pero hipersensibles al daño inducido por UV. Sin PrimPol, el proceso de curación de la piel tras haber recibido una dosis alta de UV se ve comprometido debido a la ineficiente proliferación celular en la epidermis, que es necesaria para reemplazar las células dañadas. La ausencia de PrimPol también causó una incidencia mayor de papilomas, alopecia, obesidad y desarrollo de tumores agresivos. Nuestros resultados muestran la importancia de PrimPol en el mantenimiento de la estabilidad genómica y sugieren una función supresora de tumores in vivo

Fomentar las vocaciones STEM entre las alumnas de educación secundaria a través del aprendizaje-servicio

En este artículo se describe una experiencia de Aprendizaje-Servicio llevada a cabo en la ETS de Ingenieros de Caminos, Canales y Puertos de la Universidad Politécnica de Madrid. El proyecto ha permitido poner en práctica actividades de apoyo a la educación, en concreto a los últimos años de educación secundaria y bachillerato y fomentar las vocaciones en titulaciones STEM (Ciencias, Tecnología, Ingeniería y Matemáticas) especialmente entre las estudiantes mujeres de educación secundaria y bachillerato situados en una zona con un nivel de renta inferior a la media. Para ello, alumnas de grado y máster presentaron sus trabajos fin de titulación a alumnos de educación secundaria. De esta forma, los alumnos de los centros de educación secundaria comprendieron qué es la Ingeniería Civil y qué tipos de proyectos se desarrollan bajo el ámbito de la misma. El objetivo es hacer la ingeniería un camino vital posible para las mujeres y que la decisión que ellas tomen sobre sus estudios no se vea influenciada por prejuicios y falsos sentimientos de no valía

A Component-Based Approach for Securing Indoor Home Care Applications

eHealth systems have adopted recent advances on sensing technologies together with advances in information and communication technologies (ICT) in order to provide people-centered services that improve the quality of life of an increasingly elderly population. As these eHealth services are founded on the acquisition and processing of sensitive data (e.g., personal details, diagnosis, treatments and medical history), any security threat would damage the public's confidence in them. This paper proposes a solution for the design and runtime management of indoor eHealth applications with security requirements. The proposal allows applications definition customized to patient particularities, including the early detection of health deterioration and suitable reaction (events) as well as security needs. At runtime, security support is twofold. A secured component-based platform supervises applications execution and provides events management, whilst the security of the communications among application components is also guaranteed. Additionally, the proposed event management scheme adopts the fog computing paradigm to enable local event related data storage and processing, thus saving communication bandwidth when communicating with the cloud. As a proof of concept, this proposal has been validated through the monitoring of the health status in diabetic patients at a nursing home.This work was financed under project DPI2015-68602-R (MINECO/FEDER, UE), UPV/EHU under project PPG17/56 and GV/EJ under recognized research group IT914-16

Flexibility Support for Homecare Applications Based on Models and Multi-Agent Technology

In developed countries, public health systems are under pressure due to the increasing percentage of population over 65. In this context, homecare based on ambient intelligence technology seems to be a suitable solution to allow elderly people to continue to enjoy the comforts of home and help optimize medical resources. Thus, current technological developments make it possible to build complex homecare applications that demand, among others, flexibility mechanisms for being able to evolve as context does (adaptability), as well as avoiding service disruptions in the case of node failure (availability). The solution proposed in this paper copes with these flexibility requirements through the whole life-cycle of the target applications: from design phase to runtime. The proposed domain modeling approach allows medical staff to design customized applications, taking into account the adaptability needs. It also guides software developers during system implementation. The application execution is managed by a multi-agent based middleware, making it possible to meet adaptation requirements, assuring at the same time the availability of the system even for stateful applications.This work was financed in part by the University of the Basque Country (UPV/EHU) under project UFI 11/28, by the Regional Government of the Basque Country under Project IT719-13, and by the MCYT&FEDER under project DPI 2012-37806-C02-01

Model-Driven Design and Development of Flexible Automated Production Control Configurations for Industry 4.0

The continuous changes of the market and customer demands have forced modern automation systems to provide stricter Quality of service (QoS) requirements. This work is centered in automation production system flexibility, understood as the ability to shift from one controller configuration to a different one, in the most quick and cost-effective way, without disrupting its normal operation. In the manufacturing field, this allows to deal with non-functional requirements such as assuring control system availability or workload balancing, even in the case of failure of a machine, components, network or controllers. Concretely, this work focuses on flexible applications at production level, using Programmable Logic Controllers (PLCs) as primary controllers. The reconfiguration of the control system is not always possible as it depends on the process state. Thus, an analysis of the system state is necessary to make a decision. In this sense, architectures based on industrial Multi Agent Systems (MAS) have been used to provide this support at runtime. Additionally, the introduction of these mechanisms makes the design and the implementation of the control system more complex. This work aims at supporting the design and development of such flexible automation production systems, through the proposed model-based framework. The framework consists of a set of tools that, based on models, automate the generation of control code extensions that add flexibility to the automation production system, according to industry 4.0 paradigm.This work was financed by MCIU/AEI/FEDER, UE (grant number RTI2018-096116-B-I00) and by GV/EJ (grant number IT1324-19)

Supporting Diversity in Science through Social Networking

In this Community Page, we learn how a scientific community leverages social networking tools to connect a group of dispersed scientific researchers in Ciencia Puerto Rico; this effort fosters innovative research and educational collaborations and changes the way scientists interact with the public

Estudio de la carga interna en pádel amateur mediante la frecuencia cardíaca

Los objetivos de este estudio fueron evaluar el nivel de condición física de 8 sujetos amateur de pádel, definir su perfil energético de esfuerzo en juego real mediante registro de frecuencia cardíaca (FC), y proponer un conjunto de variables justificadas de esta, que permita analizar el perfil de carga interna en pádel. Mediante prueba de esfuerzo incremental máxima en cinta de correr se obtuvieron parámetros ergoespirométricos, respiratorios y sus equivalentes cardíacos, introducidos en el sistema de registro de la FC Polar Team. Los sujetos disputaron 7 partidos de entrenamiento de 1 hora de duración y 72-96 horas de separación entre ellos, obteniéndose como variables: consumo máximo de oxígeno (VO2 máx) y porcentaje (%) de VO2 máx en el umbral anaeróbico; en prueba de esfuerzo, FC máx., media y mín., y zonas de trabajo metabólicas (rangos de FC). Los resultados presentan VO2 máx de 51,15 ± 5,73 ml · kg–1 · min–1, FC máx. durante el juego de 154,75 ± 7,25 ppm, FC med de 130,0 ± 10,4 ppm para tiempo de juego y 89,75 % del tiempo de juego en zona de trabajo aeróbica. Como conclusión, el esfuerzo al que son sometidos durante el juego a nivel cardiorrespiratorio los jugadores amateur de pádel se basa casi exclusivamente en metabolismos aeróbicos. Además, las variables máx., mín. y media de FC para tiempos de juego y descanso y el establecimiento de zonas de trabajo de FC pueden aportarnos información importante sobre lo que ocurre en el juego a nivel cardiorrespiratorio

Aspect-ratio and lateral-resolution enhancement in force microscopy by attaching nanoclusters generated by an ion cluster source at the end of a silicon tip

One of the factors that limit the spatial resolution in atomic force microscopy (AFM) is the physical size of the probe. This limitation is particularly severe when the imaged structures are comparable in size to the tip’s apex. The resolution in the AFM is usually enhanced by using sharp tips with high aspect ratios. In the present paper we propose an approach to modify AFM tips that consists of depositing nanoclusters on standard silicon tips. We show that the use of those tips leads to atomic force microscopy images of higher aspect ratios and spatial resolution. The present approach has two major properties. It provides higher aspect-ratio images of nanoscale objects and, at the same time, enables to functionalize the AFM tips by depositing nanoparticles with well-controlled chemical composition.The authors acknowledge the Spanish Ministerio de Ciencia e Innovación and Comisión Interministerial para la Ciencia Y la Tecnología—CICYT under Contract Nos. MAT2008-06765-C02-02, MAT2009-08650, and CSD2007 00041 (Nanoselect) and through the FPI and “Juan de La Cierva” programs for financial support.Peer reviewe

A Customizable Architecture for Application-Centric Management of Context-Aware Applications

[EN] Context-aware applications present common requirements (e.g., heterogeneity, scalability, adaptability, availability) in a variety of domains (e.g., healthcare, natural disaster prevention, smart factories). Besides, they do also present domain specific requirements, among which the application concept itself is included. Therefore, a platform in charge of managing their execution must be generic enough to cover common requirements, but it must also be adaptable enough to consider the domain aspects to meet the demands at application-level. Several approaches in the literature tackle some of these demands, but not all of them, and without considering the applications concept and the customization demands in different domains. This work proposes a generic and customizable management architecture that covers both types of requirements based on multi-agent technology and model-driven development. Multi-agent technology is used to enable the distributed intelligence needed to address many common requirements, whereas model-driven development allows to address domain specific particularities. On top of that, a customization methodology to develop specific platforms from this generic architecture is also presented. This methodology is assessed by means of a case study in the domain of eHealthCare. Finally, the performance of MAS-RECON is compared with the most popular tool for the orchestration of containerized applications.This work was supported in part by the Ministerio de Ciencia, Innovacion y Universidades (MCIU)/Agencia Estatal de Investigacion (AEI)/Fondo Europeo de Desarrollo Regional (FEDER), Union Europea (UE), under Grant RTI2018-096116-B-I00; and in part by the Gobierno Vasco (GV)/Eusko Jaurlaritza (EJ) under Grant IT1324-19

Validation of an instrument on tactical-offensive knowledge for players with intellectual disability

El objetivo de este trabajo se centra en construir y validar una herramienta que permita, de forma fiable, evaluar el conocimiento de base de jugadores con discapacidad intelectual leve en aspectos de táctica ofensiva en FS, permitiendo usarlo como evaluación inicial complementaria que guíe el proceso de entrenamiento. El tamaño muestral fue de 68 jugadores (Medad=27; SDedad=9.06 y Mexperiencia=11.78; SDexperiencia=1,29). Los resultados muestran valores adecuados de consistencia interna y de fiabilidad (α=0.64, ω=0.74, ICC=0.64 y test-retest). De manera complementaria, se realizó un análisis cualitativo, mediante una entrevista colectiva a un grupo de expertos, sobre la utilidad del instrumento en esta población de deportistas. Los resultados permiten establecer una sólida base para el empleo de este instrumento en futuros estudiosThe objective of this work is to build and validate a tool that allows, reliably, evaluate the base knowledge of players with mild intellectual impairment in aspects of offensive tactical in FS, so that it can be used as an additional initial evaluation to guide the training process of coaches. The sample size is constituted by 68 subjects (Mage=27; SDage=9.06 y Mexperience=11.78; SDexperience=1,29). The results show satisfactory internal consistency and reliability values (α=0.64, ω=0.74, ICC=0.64 and test-retest). In a complementary manner, a qualitative analysis was carried out, through a collective interview with a group of experts, about the utility of the instrument in this population of athletes. The results allow to establish a solid base for the use of this instrument in future studie