Oscillatory genetic circuits: new designs and mathematical models

Introduction and motivation: A wide variety of organisms have developed in-ternal biomolecular clocks in order to adapt to cyclic changes of the environment. Clock operation involves genetic networks. These genetic networks have to be mod¬eled in order to understand the underlying mechanism of oscillations and to design new synthetic cellular clocks. This doctoral thesis has resulted in two contributions to the fields of genetic clocks and systems and synthetic biology, generally. The first contribution is a new genetic circuit model that exhibits an oscillatory behav¬ior through catalytic RNA molecules. The second and major contribution is a new genetic circuit model demonstrating that a repressor molecule acting on the positive feedback of a self-activating gene produces reliable oscillations. First contribution: A new model of a synthetic genetic oscillator based on a typical two-gene motif with one positive and one negative feedback loop is pre¬sented. The originality is that the repressor is a catalytic RNA molecule rather than a protein or a non-catalytic RNA molecule. This catalytic RNA is a ribozyme that acts post-transcriptionally by binding to and cleaving target mRNA molecules. This genetic clock involves just two genes, a mRNA and an activator protein, apart from the ribozyme. Parameter values that produce a circadian period in both determin¬istic and stochastic simulations have been chosen as an example of clock operation. The effects of the stochastic fluctuations are quantified by a period histogram and autocorrelation function. The conclusion is that catalytic RNA molecules can act as repressor proteins and simplify the design of genetic oscillators. Second and major contribution: It is demonstrated that a self-activating gene in conjunction with a simple negative interaction can easily produce robust matically validated. This model is comprised of two clearly distinct parts. The first is a positive feedback created by a protein that binds to the promoter of its own gene and activates the transcription. The second is a negative interaction in which a repressor molecule prevents this protein from binding to its promoter. A stochastic study shows that the system is robust to noise. A deterministic study identifies that the oscillator dynamics are mainly driven by two types of biomolecules: the protein, and the complex formed by the repressor and this protein. The main conclusion of this study is that a simple and usual negative interaction, such as degradation, se¬questration or inhibition, acting on the positive transcriptional feedback of a single gene is a sufficient condition to produce reliable oscillations. One gene is enough and the positive transcriptional feedback signal does not need to activate a second repressor gene. At the genetic level, this means that an explicit negative feedback loop is not necessary. Unlike many genetic oscillators, this model needs neither cooperative binding reactions nor the formation of protein multimers. Applications and future research directions: Recently, RNA molecules have been found to play many new catalytic roles. The first oscillatory genetic model proposed in this thesis uses ribozymes as repressor molecules. This could provide new synthetic biology design principles and a better understanding of cel¬lular clocks regulated by RNA molecules. The second genetic model proposed here involves only a repression acting on a self-activating gene and produces robust oscil¬lations. Unlike current two-gene oscillators, this model surprisingly does not require a second repressor gene. This result could help to clarify the design principles of cellular clocks and constitute a new efficient tool for engineering synthetic genetic oscillators. Possible follow-on research directions are: validate models in vivo and in vitro, research the potential of second model as a genetic memory, investigate new genetic oscillators regulated by non-coding RNAs and design a biosensor of positive feedbacks in genetic networks based on the operation of the second model Resumen Introduccion y motivacion: Una amplia variedad de organismos han desarro-llado relojes biomoleculares internos con el fin de adaptarse a los cambios ciclicos del entorno. El funcionamiento de estos relojes involucra redes geneticas. El mo delado de estas redes geneticas es esencial tanto para entender los mecanismos que producen las oscilaciones como para diseiiar nuevos circuitos sinteticos en celulas. Esta tesis doctoral ha dado lugar a dos contribuciones dentro de los campos de los circuitos geneticos en particular, y biologia de sistemas y sintetica en general. La primera contribucion es un nuevo modelo de circuito genetico que muestra un comportamiento oscilatorio usando moleculas de ARN cataliticas. La segunda y principal contribucion es un nuevo modelo de circuito genetico que demuestra que una molecula represora actuando sobre el lazo de un gen auto-activado produce oscilaciones robustas. Primera contribucion: Es un nuevo modelo de oscilador genetico sintetico basado en una tipica red genetica compuesta por dos genes con dos lazos de retroa-limentacion, uno positivo y otro negativo. La novedad de este modelo es que el represor es una molecula de ARN catalftica, en lugar de una protefna o una molecula de ARN no-catalitica. Este ARN catalitico es una ribozima que actua despues de la transcription genetica uniendose y cortando moleculas de ARN mensajero (ARNm). Este reloj genetico involucra solo dos genes, un ARNm y una proteina activadora, aparte de la ribozima. Como ejemplo de funcionamiento, se han escogido valores de los parametros que producen oscilaciones con periodo circadiano (24 horas) tanto en simulaciones deterministas como estocasticas. El efecto de las fluctuaciones es-tocasticas ha sido cuantificado mediante un histograma del periodo y la función de auto-correlacion. La conclusion es que las moleculas de ARN con propiedades cataliticas pueden jugar el misnio papel que las protemas represoras, y por lo tanto, simplificar el diseno de los osciladores geneticos. Segunda y principal contribucion: Es un nuevo modelo de oscilador genetico que demuestra que un gen auto-activado junto con una simple interaction negativa puede producir oscilaciones robustas. Este modelo ha sido estudiado y validado matematicamente. El modelo esta compuesto de dos partes bien diferenciadas. La primera parte es un lazo de retroalimentacion positiva creado por una proteina que se une al promotor de su propio gen activando la transcription. La segunda parte es una interaction negativa en la que una molecula represora evita la union de la proteina con el promotor. Un estudio estocastico muestra que el sistema es robusto al ruido. Un estudio determinista muestra que la dinamica del sistema es debida principalmente a dos tipos de biomoleculas: la proteina, y el complejo formado por el represor y esta proteina. La conclusion principal de este estudio es que una simple y usual interaction negativa, tal como una degradation, un secuestro o una inhibition, actuando sobre el lazo de retroalimentacion positiva de un solo gen es una condition suficiente para producir oscilaciones robustas. Un gen es suficiente y el lazo de retroalimentacion positiva no necesita activar a un segundo gen represor, tal y como ocurre en los relojes actuales con dos genes. Esto significa que a nivel genetico un lazo de retroalimentacion negativa no es necesario de forma explicita. Ademas, este modelo no necesita reacciones cooperativas ni la formation de multimeros proteicos, al contrario que en muchos osciladores geneticos. Aplicaciones y futuras lineas de investigacion: En los liltimos anos, se han descubierto muchas moleculas de ARN con capacidad catalitica. El primer modelo de oscilador genetico propuesto en esta tesis usa ribozimas como moleculas repre¬soras. Esto podria proporcionar nuevos principios de diseno en biologia sintetica y una mejor comprension de los relojes celulares regulados por moleculas de ARN. El segundo modelo de oscilador genetico propuesto aqui involucra solo una represion actuando sobre un gen auto-activado y produce oscilaciones robustas. Sorprendente-mente, un segundo gen represor no es necesario al contrario que en los bien conocidos osciladores con dos genes. Este resultado podria ayudar a clarificar los principios de diseno de los relojes celulares naturales y constituir una nueva y eficiente he-rramienta para crear osciladores geneticos sinteticos. Algunas de las futuras lineas de investigation abiertas tras esta tesis son: (1) la validation in vivo e in vitro de ambos modelos, (2) el estudio del potential del segundo modelo como circuito base para la construction de una memoria genetica, (3) el estudio de nuevos osciladores geneticos regulados por ARN no codificante y, por ultimo, (4) el rediseno del se¬gundo modelo de oscilador genetico para su uso como biosensor capaz de detectar genes auto-activados en redes geneticas.

An abstract machine based execution model for computer architecture design and efficient implementation of logic programs in parallel.

The term "Logic Programming" refers to a variety of computer languages and execution models which are based on the traditional concept of Symbolic Logic. The expressive power of these languages offers promise to be of great assistance in facing the programming challenges of present and future symbolic processing applications in Artificial Intelligence, Knowledge-based systems, and many other areas of computing. The sequential execution speed of logic programs has been greatly improved since the advent of the first interpreters. However, higher inference speeds are still required in order to meet the demands of applications such as those contemplated for next generation computer systems. The execution of logic programs in parallel is currently considered a promising strategy for attaining such inference speeds. Logic Programming in turn appears as a suitable programming paradigm for parallel architectures because of the many opportunities for parallel execution present in the implementation of logic programs. This dissertation presents an efficient parallel execution model for logic programs. The model is described from the source language level down to an "Abstract Machine" level suitable for direct implementation on existing parallel systems or for the design of special purpose parallel architectures. Few assumptions are made at the source language level and therefore the techniques developed and the general Abstract Machine design are applicable to a variety of logic (and also functional) languages. These techniques offer efficient solutions to several areas of parallel Logic Programming implementation previously considered problematic or a source of considerable overhead, such as the detection and handling of variable binding conflicts in AND-Parallelism, the specification of control and management of the execution tree, the treatment of distributed backtracking, and goal scheduling and memory management issues, etc. A parallel Abstract Machine design is offered, specifying data areas, operation, and a suitable instruction set. This design is based on extending to a parallel environment the techniques introduced by the Warren Abstract Machine, which have already made very fast and space efficient sequential systems a reality. Therefore, the model herein presented is capable of retaining sequential execution speed similar to that of high performance sequential systems, while extracting additional gains in speed by efficiently implementing parallel execution. These claims are supported by simulations of the Abstract Machine on sample programs.

3-Phase Rectifier System with very demanding dynamic load: Architecture analysis and control strategy

A distributed power architecture for aerospace application with very restrictive specifications is analyzed. Parameters as volume, weight and losses are analyzed for the considered power architectures. In order to protect the 3 phase generator against high load steps, an intermediate bus (based in a high capacitance) to provide energy to the loads during the high load steps is included. Prototypes of the selected architecture for the rectifier and EMI filter are built and the energy control is validated.

Naturaleza, ética, y arquitectura: Autenticidad y criterios éticos que integran el desarrollo de una arquitectura más sostenible = Nature, ethics and architecture: Authenticity and ethics criteria that integrate the development of a more sustainable architecture

La ecología no solamente ha puesto de manifiesto problemas ambientales, sino que ha confirmado la necesidad de una nueva armonía entre los propios seres humanos y de éstos con la naturaleza y con todos los seres que la habitan. Es necesario un nuevo contrato que determine nuestras relaciones con la Naturaleza (Serrs1), y una nueva Ética para nuestras vidas (Guattari2). La ética medioambiental nos ha dado una visión universal y supra-generacional de la gestión de la naturaleza y como consecuencia, una nueva forma de construir nuestra ‘segunda’ Naturaleza, que es la arquitectura. ¿Qué es lo esencial que esta nueva ética exige para la arquitectura? Este es un momento crucial para reconsiderar los objetivos de la arquitectura, porque lo ‘eco’ está produciendo grandes cambios. ¿Implica esta era post-ecológica una particular ética, es decir, referida a sus fines y medios? ¿Porqué, para qué, para quién, cómo debemos hacer la arquitectura de nuestro tiempo? Es momento de afrontar críticamente el discurso de la eco-arquitectura, e incluso de repensar los propios límites de la arquitectura. El desarrollo actual del conocimiento medioambiental es esencialmente técnico y utilitario, pero ¿es el reto solamente técnico?¿Es suficiente la suma de lo medioambiental-social-económico-cultural para definirla? ¿Hay claves que nos puedan dar la dimensión ética de esta aproximación técnica-empírica? ¿Sabemos lo que estamos haciendo cuando aplicamos este conocimiento? Y, sobre todo, ¿cuál es el sentido de lo que estamos haciendo? La tesis que se propone puede resumirse: De acuerdo con el actual conocimiento que tenemos de la Naturaleza, la Arquitectura de nuestro tiempo deber reconsiderar sus fines y medios, puesto que la ética medioambiental está definiendo nuevos objetivos. Para fundamentar y profundizar en esta afirmación la tesis analiza cómo son hoy día las relaciones entre Ética-Naturaleza-Arquitectura (Fig.1), lo que facilitará las claves de cuáles son los criterios éticos (en cuanto a fines y medios) que deben definir la arquitectura del tiempo de la ecología. ABSTRACT Ecology shows us not only environmental problems; it shows that we need a new balance and harmony between individuals, beings, communities and Nature. We need a new contract with Nature according to Serres576, and a new Ethics for our lives according to Guattari577. Environmental ethics have given us a universal and supra-generational vision of the management of our Nature and, as a consequence, a new way to construct our ‘second’ nature, which is architecture. What is essential for this new architecture that the new ethics demand? This is a critical moment to reconsider the object of architecture, because the ‘eco’ is making significant changes in it. Are there any specifically ethical concerns (ends and means) in the post-ecological era? Why, for what, for whom, how should we make architecture in our times? This is the time to approach the eco-architectural discourse critically and to question the current boundaries of architecture itself: Where is eco-architecture going? The current development of environmental knowledge is essentially technical and utilitarian, but it is its technical aspect the only challenge? Is the sum of environmental-social-economic aspects enough to define it? Are there any clues which can give an ethical sense to this technical-empirical approach? Do we know what we are doing when we apply this knowledge? And overall, what is the meaning of what we are doing? Exploring this subject, this thesis makes a statement: In accordance with the actual knowledge of Nature, Architecture of our time must reconsider its ends and means, since the environmental ethics is defining new objectives. To support that, the thesis analyzes what the relationships between Ethics –Nature- Architecture (Fig. 53) are like nowadays, this will provide the clues of which ethical criteria (ends and means) must architecture of an ecological era define.

Taxonomía de la evolución arquitectónica 'Architecture 2000 and beyond'

Architecture 2000, publicado por Charles Jencks originalmente en el año 1971, es un libro cuyo objetivo explícito es predecir el futuro de la arquitectura hasta el fi nal del siglo XX y este hecho ha favorecido que se haya editado por segunda vez precisamente ese emblemático año 2000, manteniendo fundamentalmente el contenido original, aunque añadiendo una introducción y un capítulo final en los que se ofrece una evaluación de lo dicho treinta años antes. En este nuevo Architecture 2000, Jencks añade al texto principal sus comentarios sobre lo acertado o erróneo de sus predicciones anteriores, juzgadas ahora a la luz de los hechos acontecidos y actualiza su mapa evolutivo con una nueva versión de su anterior Evolutionary Tree to the year 2000, que ahora pasará a denominarse Evolutionary Tree of the 20th century architecture. No hay duda de que lo más infl uyente y duradero de la obra de Jencks ha sido la representación gráfica de la evolución de la arquitectura a través de sus evolutionary trees. Pero, aunque Architecture 2000 debe ser valorada sobre todo como una aportación imprescindible para entender la arquitectura de la segunda mitad del siglo XX, tanto por su identifi cación de las tendencias a través de los que se han conducido las experiencias de los profesionales como por su concepción dinámica de la arquitectura plasmada gráfi camente a través de sus evolutionary trees, sería un error considerarla como mera historia y dar por amortizados los métodos prospectivos o los juicios y expectativas del autor sobre lo acontecido en la arquitectura a lo largo de las últimas décadas.

Survey on integration of Applied Geology and Geomorphology in Civil Engineering curricula in the framework of the European Higher Education Area (EHEA) by using practical trips

The engineer must have sufficient theoretical knowledge to be applied to solve specific problems, with the necessary capacity to simplify these approaches, and taking into account factors such as speed, simplicity, quality and economy. In Geology, its ultimate goal is the exploration of the history of the geological events through observation, deduction, reasoning and, in exceptional cases by the direct underground exploration or experimentation. Experimentation is very limited in Geology. Reproduction laboratory of certain phenomena or geological processes is difficult because both time and space become a large scale. For this reason, some Earth Sciences are in a nearly descriptive stage whereas others closest to the experimental, Geophysics and Geochemistry, have assimilated progress experienced by the physics and chemistry. Thus, Anglo-Saxon countries clearly separate Engineering Geology from Geological Engineering, i.e. Applied Geology to the Geological Engineering concepts. Although there is a big professional overlap, the first one corresponds to scientific approach, while the last one corresponds to a technological one. Applied Geology to Engineering could be defined as the Science and Applied Geology to the design, construction and performance of engineering infrastructures in and field geology discipline. There has been much discussion on the primacy of theory over practice. Today prevails the exaggeration of practice, but you get good workers and routine and mediocre teachers. This idea forgets too that teaching problem is a problem of right balance. The approach of the action lines on the European Higher Education Area (EHEA) framework provides for such balance. Applied Geology subject represents the first real contact with the physical environment with the practice profession and works. Besides, the situation of the topic in the first trace of Study Plans for many students implies the link to other subjects and topics of the career (tunnels, dams, groundwater, roads, etc). This work analyses in depth the justification of such practical trips. It shows the criteria and methods of planning and the result which manifests itself in pupils. Once practical trips experience developed, the objective work tries to know about results and changes on student’s motivation in learning perspective. This is done regardless of the outcome of their knowledge achievements assessed properly and they are not subject to such work. For this objective, it has been designed a survey about their motivation before and after trip. Survey was made by the Unidad Docente de Geología Aplicada of the Departamento de Ingeniería y Morfología del Terreno (Escuela Técnica Superior de Ingenieros de Caminos, Canales y Puertos, Universidad Politécnica de Madrid). It was completely anonymous. Its objective was to collect the opinion of the student as a key agent of learning and teaching of the subject. All the work takes place under new teaching/learning criteria approach at the European framework in Higher Education. The results are exceptionally good with 90% of student’s participation and with very high scores in a number of questions as the itineraries, teachers and visited places (range of 4.5 to 4.2 in a 5 points scale). The majority of students are very satisfied (average of 4.5 in a 5 points scale).