Magnetic Attitude Control System for a Small Satellite. Impact on the Thermal Performance

El principal objetivo de la tesis es estudiar el acoplamiento entre los subsistemas de control de actitud y de control térmico de un pequeño satélite, con el fin de buscar la solución a los problemas relacionados con la determinación de los parámetros de diseño. Se considera la evolución de la actitud y de las temperaturas del satélite bajo la influencia de dos estrategias de orientación diferentes: 1) estabilización magnética pasiva de la orientación (PMAS, passive magnetic attitude stabilization), y 2) control de actitud magnético activo (AMAC, active magnetic attitude control). En primer lugar se presenta el modelo matemático del problema, que incluye la dinámica rotacional y el modelo térmico. En el problema térmico se considera un satélite cúbico modelizado por medio de siete nodos (seis externos y uno interno) aplicando la ecuación del balance térmico. Una vez establecido el modelo matemático del problema, se estudia la evolución que corresponde a las dos estrategias mencionadas. La estrategia PMAS se ha seleccionado por su simplicidad, fiabilidad, bajo coste, ahorrando consumo de potencia, masa coste y complejidad, comparado con otras estrategias. Se ha considerado otra estrategia de control que consigue que el satélite gire a una velocidad requerida alrededor de un eje deseado de giro, pudiendo controlar su dirección en un sistema inercial de referencia, ya que frecuentemente el subsistema térmico establece requisitos de giro alrededor de un eje del satélite orientado en una dirección perpendicular a la radiación solar incidente. En relación con el problema térmico, para estudiar la influencia de la velocidad de giro en la evolución de las temperaturas en diversos puntos del satélite, se ha empleado un modelo térmico linealizado, obtenido a partir de la formulación no lineal aplicando un método de perturbaciones. El resultado del estudio muestra que el tiempo de estabilización de la temperatura y la influencia de las cargas periódicas externas disminuye cuando aumenta la velocidad de giro. Los cambios de temperatura se reducen hasta ser muy pequeños para velocidades de rotación altas. En relación con la estrategia PMAC se ha observado que a pesar de su uso extendido entre los micro y nano satélites todavía presenta problemas que resolver. Estos problemas están relacionados con el dimensionamiento de los parámetros del sistema y la predicción del funcionamiento en órbita. Los problemas aparecen debido a la dificultad en la determinación de las características magnéticas de los cuerpos ferromagnéticos (varillas de histéresis) que se utilizan como amortiguadores de oscilaciones en los satélites. Para estudiar este problema se presenta un modelo analítico que permite estimar la eficiencia del amortiguamiento, y que se ha aplicado al estudio del comportamiento en vuelo de varios satélites, y que se ha empleado para comparar los resultados del modelo con los obtenidos en vuelo, observándose que el modelo permite explicar satisfactoriamente el comportamiento registrado. ABSTRACT The main objective of this thesis is to study the coupling between the attitude control and thermal control subsystems of a small satellite, and address the solution to some existing issues concerning the determination of their parameters. Through the thesis the attitude and temperature evolution of the satellite is studied under the influence of two independent attitude stabilization and control strategies: (1) passive magnetic attitude stabilization (PMAS), and (2) active magnetic attitude control (AMAC). In this regard the mathematical model of the problem is explained and presented. The mathematical model includes both the rotational dynamics and the thermal model. The thermal model is derived for a cubic satellite by solving the heat balance equation for 6 external and 1 internal nodes. Once established the mathematical model of the problem, the above mentioned attitude strategies were applied to the system and the temperature evolution of the 7 nodes of the satellite was studied. The PMAS technique has been selected to be studied due to its prevalent use, simplicity, reliability, and cost, as this strategy significantly saves the overall power, weight, cost, and reduces the complexity of the system compared to other attitude control strategies. In addition to that, another control law that provides the satellite with a desired spin rate along a desired axis of the satellite, whose direction can be controlled with respect to the inertial reference frame is considered, as the thermal subsystem of a satellite usually demands a spin requirement around an axis of the satellite which is positioned perpendicular to the direction of the coming solar radiation. Concerning the thermal problem, to study the influence of spin rate on temperature evolution of the satellite a linear approach of the thermal model is used, which is based on perturbation theory applied to the nonlinear differential equations of the thermal model of a spacecraft moving in a closed orbit. The results of this study showed that the temperature stabilization time and the periodic influence of the external thermal loads decreases by increasing the spin rate. However, the changes become insignificant for higher values of spin rate. Concerning the PMAS strategy, it was observed that in spite of its extended application to micro and nano satellites, still there are some issues to be solved regarding this strategy. These issues are related to the sizing of its system parameters and predicting the in-orbit performance. The problems were found to be rooted in the difficulties that exist in determining the magnetic characteristics of the ferromagnetic bodies (hysteresis rods) that are applied as damping devices on-board satellites. To address these issues an analytic model for estimating their damping efficiency is proposed and applied to several existing satellites in order to compare the results with their respective in-flight data. This model can explain the behavior showed by these satellites.

A control theoretic approach to solve a constrained uplink power dynamic game

This paper addresses an uplink power control dynamic game where we assume that each user battery represents the system state that changes with time following a discrete-time version of a differential game. To overcome the complexity of the analysis of a dynamic game approach we focus on the concept of Dynamic Potential Games showing that the game can be solved as an equivalent Multivariate Optimum Control Problem. The solution of this problem is quite interesting because different users split the activity in time, avoiding higher interferences and providing a long term fairness.

Optimization of the Integrated Guidance and Control for a Dual Aerodynamic Control Missile

Las futuras misiones para misiles aire-aire operando dentro de la atmósfera requieren la interceptación de blancos a mayores velocidades y más maniobrables, incluyendo los esperados vehículos aéreos de combate no tripulados. La intercepción tiene que lograrse desde cualquier ángulo de lanzamiento. Una de las principales discusiones en la tecnología de misiles en la actualidad es cómo satisfacer estos nuevos requisitos incrementando la capacidad de maniobra del misil y en paralelo, a través de mejoras en los métodos de guiado y control modernos. Esta Tesis aborda estos dos objetivos simultáneamente, al proponer un diseño integrando el guiado y el control de vuelo (autopiloto) y aplicarlo a misiles con control aerodinámico simultáneo en canard y cola. Un primer avance de los resultados obtenidos ha sido publicado recientemente en el Journal of Aerospace Engineering, en Abril de 2015, [Ibarrondo y Sanz-Aranguez, 2015]. El valor del diseño integrado obtenido es que permite al misil cumplir con los requisitos operacionales mencionados empleando únicamente control aerodinámico. El diseño propuesto se compara favorablemente con esquemas más tradicionales, consiguiendo menores distancias de paso al blanco y necesitando de menores esfuerzos de control incluso en presencia de ruidos. En esta Tesis se demostrará cómo la introducción del doble mando, donde tanto el canard como las aletas de cola son móviles, puede mejorar las actuaciones de un misil existente. Comparado con un misil con control en cola, el doble control requiere sólo introducir dos servos adicionales para accionar los canards también en guiñada y cabeceo. La sección de cola será responsable de controlar el misil en balanceo mediante deflexiones diferenciales de los controles. En el caso del doble mando, la complicación añadida es que los vórtices desprendidos de los canards se propagan corriente abajo y pueden incidir sobre las superficies de cola, alterando sus características de control. Como un primer aporte, se ha desarrollado un modelo analítico completo para la aerodinámica no lineal de un misil con doble control, incluyendo la caracterización de este efecto de acoplamiento aerodinámico. Hay dos modos de funcionamiento en picado y guiñada para un misil de doble mando: ”desviación” y ”opuesto”. En modo ”desviación”, los controles actúan en la misma dirección, generando un cambio inmediato en la sustentación y produciendo un movimiento de translación en el misil. La respuesta es rápida, pero en el modo ”desviación” los misiles con doble control pueden tener dificultades para alcanzar grandes ángulos de ataque y altas aceleraciones laterales. Cuando los controles actúan en direcciones opuestas, el misil rota y el ángulo de ataque del fuselaje se incrementa para generar mayores aceleraciones en estado estacionario, aunque el tiempo de respuesta es mayor. Con el modelo aerodinámico completo, es posible obtener una parametrización dependiente de los estados de la dinámica de corto periodo del misil. Debido al efecto de acoplamiento entre los controles, la respuesta en bucle abierto no depende linealmente de los controles. El autopiloto se optimiza para obtener la maniobra requerida por la ley de guiado sin exceder ninguno de los límites aerodinámicos o mecánicos del misil. Una segunda contribución de la tesis es el desarrollo de un autopiloto con múltiples entradas de control y que integra la aerodinámica no lineal, controlando los tres canales de picado, guiñada y cabeceo de forma simultánea. Las ganancias del autopiloto dependen de los estados del misil y se calculan a cada paso de integración mediante la resolución de una ecuación de Riccati de orden 21x21. Las ganancias obtenidas son sub-óptimas, debido a que una solución completa de la ecuación de Hamilton-Jacobi-Bellman no puede obtenerse de manera práctica, y se asumen ciertas simplificaciones. Se incorpora asimismo un mecanismo que permite acelerar la respuesta en caso necesario. Como parte del autopiloto, se define una estrategia para repartir el esfuerzo de control entre el canard y la cola. Esto se consigue mediante un controlador aumentado situado antes del bucle de optimización, que minimiza el esfuerzo total de control para maniobrar. Esta ley de alimentación directa mantiene al misil cerca de sus condiciones de equilibrio, garantizando una respuesta transitoria adecuada. El controlador no lineal elimina la respuesta de fase no-mínima característica de la cola. En esta Tesis se consideran dos diseños para el guiado y control, el control en Doble-Lazo y el control Integrado. En la aproximación de Doble-Lazo, el autopiloto se sitúa dentro de un bucle interior y se diseña independientemente del guiado, que conforma el bucle más exterior del control. Esta estructura asume que existe separación espectral entre los dos, esto es, que los tiempos de respuesta del autopiloto son mucho mayores que los tiempos característicos del guiado. En el estudio se combina el autopiloto desarrollado con una ley de guiado óptimo. Los resultados obtenidos demuestran que se consiguen aumentos muy importantes en las actuaciones frente a misiles con control canard o control en cola, y que la interceptación, cuando se lanza cerca del curso de colisión, se consigue desde cualquier ángulo alrededor del blanco. Para el misil de doble mando, la estrategia óptima resulta en utilizar el modo de control opuesto en la aproximación al blanco y utilizar el modo de desviación justo antes del impacto. Sin embargo la lógica de doble bucle no consigue el impacto cuando hay desviaciones importantes con respecto al curso de colisión. Una de las razones es que parte de la demanda de guiado se pierde, ya que el misil solo es capaz de modificar su aceleración lateral, y no tiene control sobre su aceleración axial, a no ser que incorpore un motor de empuje regulable. La hipótesis de separación mencionada, y que constituye la base del Doble-Bucle, puede no ser aplicable cuando la dinámica del misil es muy alta en las proximidades del blanco. Si se combinan el guiado y el autopiloto en un único bucle, la información de los estados del misil está disponible para el cálculo de la ley de guiado, y puede calcularse la estrategia optima de guiado considerando las capacidades y la actitud del misil. Una tercera contribución de la Tesis es la resolución de este segundo diseño, la integración no lineal del guiado y del autopiloto (IGA) para el misil de doble control. Aproximaciones anteriores en la literatura han planteado este sistema en ejes cuerpo, resultando en un sistema muy inestable debido al bajo amortiguamiento del misil en cabeceo y guiñada. Las simplificaciones que se tomaron también causan que el misil se deslice alrededor del blanco y no consiga la intercepción. En nuestra aproximación el problema se plantea en ejes inerciales y se recurre a la dinámica de los cuaterniones, eliminado estos inconvenientes. No se limita a la dinámica de corto periodo del misil, porque se construye incluyendo de modo explícito la velocidad dentro del bucle de optimización. La formulación resultante en el IGA es independiente de la maniobra del blanco, que sin embargo se ha de incluir en el cálculo del modelo en Doble-bucle. Un típico inconveniente de los sistemas integrados con controlador proporcional, es el problema de las escalas. Los errores de guiado dominan sobre los errores de posición del misil y saturan el controlador, provocando la pérdida del misil. Este problema se ha tratado aquí con un controlador aumentado previo al bucle de optimización, que define un estado de equilibrio local para el sistema integrado, que pasa a actuar como un regulador. Los criterios de actuaciones para el IGA son los mismos que para el sistema de Doble-Bucle. Sin embargo el problema matemático resultante es muy complejo. El problema óptimo para tiempo finito resulta en una ecuación diferencial de Riccati con condiciones terminales, que no puede resolverse. Mediante un cambio de variable y la introducción de una matriz de transición, este problema se transforma en una ecuación diferencial de Lyapunov que puede resolverse mediante métodos numéricos. La solución resultante solo es aplicable en un entorno cercano del blanco. Cuando la distancia entre misil y blanco es mayor, se desarrolla una solución aproximada basada en la solución de una ecuación algebraica de Riccati para cada paso de integración. Los resultados que se han obtenido demuestran, a través de análisis numéricos en distintos escenarios, que la solución integrada es mejor que el sistema de Doble-Bucle. Las trayectorias resultantes son muy distintas. El IGA preserva el guiado del misil y consigue maximizar el uso de la propulsión, consiguiendo la interceptación del blanco en menores tiempos de vuelo. El sistema es capaz de lograr el impacto donde el Doble-Bucle falla, y además requiere un orden menos de magnitud en la cantidad de cálculos necesarios. El efecto de los ruidos radar, datos discretos y errores del radomo se investigan. El IGA es más robusto, resultando menos afectado por perturbaciones que el Doble- Bucle, especialmente porque el núcleo de optimización en el IGA es independiente de la maniobra del blanco. La estimación de la maniobra del blanco es siempre imprecisa y contaminada por ruido, y degrada la precisión de la solución de Doble-Bucle. Finalmente, como una cuarta contribución, se demuestra que el misil con guiado IGA es capaz de realizar una maniobra de defensa contra un blanco que ataque por su cola, sólo con control aerodinámico. Las trayectorias estudiadas consideran una fase pre-programada de alta velocidad de giro, manteniendo siempre el misil dentro de su envuelta de vuelo. Este procedimiento no necesita recurrir a soluciones técnicamente más complejas como el control vectorial del empuje o control por chorro para ejecutar esta maniobra. En todas las demostraciones matemáticas se utiliza el producto de Kronecker como una herramienta practica para manejar las parametrizaciones dependientes de variables, que resultan en matrices de grandes dimensiones. ABSTRACT Future missions for air to air endo-atmospheric missiles require the interception of targets with higher speeds and more maneuverable, including forthcoming unmanned supersonic combat vehicles. The interception will need to be achieved from any angle and off-boresight launch conditions. One of the most significant discussions in missile technology today is how to satisfy these new operational requirements by increasing missile maneuvering capabilities and in parallel, through the development of more advanced guidance and control methods. This Thesis addresses these two objectives by proposing a novel optimal integrated guidance and autopilot design scheme, applicable to more maneuverable missiles with forward and rearward aerodynamic controls. A first insight of these results have been recently published in the Journal of Aerospace Engineering in April 2015, [Ibarrondo and Sanz-Aránguez, 2015]. The value of this integrated solution is that it allows the missile to comply with the aforementioned requirements only by applying aerodynamic control. The proposed design is compared against more traditional guidance and control approaches with positive results, achieving reduced control efforts and lower miss distances with the integrated logic even in the presence of noises. In this Thesis it will be demonstrated how the dual control missile, where canard and tail fins are both movable, can enhance the capabilities of an existing missile airframe. Compared to a tail missile, dual control only requires two additional servos to actuate the canards in pitch and yaw. The tail section will be responsible to maintain the missile stabilized in roll, like in a classic tail missile. The additional complexity is that the vortices shed from the canard propagate downstream where they interact with the tail surfaces, altering the tail expected control characteristics. These aerodynamic phenomena must be properly described, as a preliminary step, with high enough precision for advanced guidance and control studies. As a first contribution we have developed a full analytical model of the nonlinear aerodynamics of a missile with dual control, including the characterization of this cross-control coupling effect. This development has been produced from a theoretical model validated with reliable practical data obtained from wind tunnel experiments available in the scientific literature, complement with computer fluid dynamics and semi-experimental methods. There are two modes of operating a missile with forward and rear controls, ”divert” and ”opposite” modes. In divert mode, controls are deflected in the same direction, generating an increment in direct lift and missile translation. Response is fast, but in this mode, dual control missiles may have difficulties in achieving large angles of attack and high level of lateral accelerations. When controls are deflected in opposite directions (opposite mode) the missile airframe rotates and the body angle of attack is increased to generate greater accelerations in steady-state, although the response time is larger. With the aero-model, a state dependent parametrization of the dual control missile short term dynamics can be obtained. Due to the cross-coupling effect, the open loop dynamics for the dual control missile is not linearly dependent of the fin positions. The short term missile dynamics are blended with the servo system to obtain an extended autopilot model, where the response is linear with the control fins turning rates, that will be the control variables. The flight control loop is optimized to achieve the maneuver required by the guidance law without exceeding any of the missile aerodynamic or mechanical limitations. The specific aero-limitations and relevant performance indicators for the dual control are set as part of the analysis. A second contribution of this Thesis is the development of a step-tracking multi-input autopilot that integrates non-linear aerodynamics. The designed dual control missile autopilot is a full three dimensional autopilot, where roll, pitch and yaw are integrated, calculating command inputs simultaneously. The autopilot control gains are state dependent, and calculated at each integration step solving a matrix Riccati equation of order 21x21. The resulting gains are sub-optimal as a full solution for the Hamilton-Jacobi-Bellman equation cannot be resolved in practical terms and some simplifications are taken. Acceleration mechanisms with an λ-shift is incorporated in the design. As part of the autopilot, a strategy is defined for proper allocation of control effort between canard and tail channels. This is achieved with an augmented feed forward controller that minimizes the total control effort of the missile to maneuver. The feedforward law also maintains the missile near trim conditions, obtaining a well manner response of the missile. The nonlinear controller proves to eliminate the non-minimum phase effect of the tail. Two guidance and control designs have been considered in this Thesis: the Two- Loop and the Integrated approaches. In the Two-Loop approach, the autopilot is placed in an inner loop and designed separately from an outer guidance loop. This structure assumes that spectral separation holds, meaning that the autopilot response times are much higher than the guidance command updates. The developed nonlinear autopilot is linked in the study to an optimal guidance law. Simulations are carried on launching close to collision course against supersonic and highly maneuver targets. Results demonstrate a large boost in performance provided by the dual control versus more traditional canard and tail missiles, where interception with the dual control close to collision course is achieved form 365deg all around the target. It is shown that for the dual control missile the optimal flight strategy results in using opposite control in its approach to target and quick corrections with divert just before impact. However the Two-Loop logic fails to achieve target interception when there are large deviations initially from collision course. One of the reasons is that part of the guidance command is not followed, because the missile is not able to control its axial acceleration without a throttleable engine. Also the separation hypothesis may not be applicable for a high dynamic vehicle like a dual control missile approaching a maneuvering target. If the guidance and autopilot are combined into a single loop, the guidance law will have information of the missile states and could calculate the most optimal approach to the target considering the actual capabilities and attitude of the missile. A third contribution of this Thesis is the resolution of the mentioned second design, the non-linear integrated guidance and autopilot (IGA) problem for the dual control missile. Previous approaches in the literature have posed the problem in body axes, resulting in high unstable behavior due to the low damping of the missile, and have also caused the missile to slide around the target and not actually hitting it. The IGA system is posed here in inertial axes and quaternion dynamics, eliminating these inconveniences. It is not restricted to the missile short term dynamic, and we have explicitly included the missile speed as a state variable. The IGA formulation is also independent of the target maneuver model that is explicitly included in the Two-loop optimal guidance law model. A typical problem of the integrated systems with a proportional control law is the problem of scales. The guidance errors are larger than missile state errors during most of the flight and result in high gains, control saturation and loss of control. It has been addressed here with an integrated feedforward controller that defines a local equilibrium state at each flight point and the controller acts as a regulator to minimize the IGA states excursions versus the defined feedforward state. The performance criteria for the IGA are the same as in the Two-Loop case. However the resulting optimization problem is mathematically very complex. The optimal problem in a finite-time horizon results in an irresoluble state dependent differential Riccati equation with terminal conditions. With a change of variable and the introduction of a transition matrix, the equation is transformed into a time differential Lyapunov equation that can be solved with known numerical methods in real time. This solution results range limited, and applicable when the missile is in a close neighborhood of the target. For larger ranges, an approximate solution is used, obtained from solution of an algebraic matrix Riccati equation at each integration step. The results obtained show, by mean of several comparative numerical tests in diverse homing scenarios, than the integrated approach is a better solution that the Two- Loop scheme. Trajectories obtained are very different in the two cases. The IGA fully preserves the guidance command and it is able to maximize the utilization of the missile propulsion system, achieving interception with lower miss distances and in lower flight times. The IGA can achieve interception against off-boresight targets where the Two- Loop was not able to success. As an additional advantage, the IGA also requires one order of magnitude less calculations than the Two-Loop solution. The effects of radar noises, discrete radar data and radome errors are investigated. IGA solution is robust, and less affected by radar than the Two-Loop, especially because the target maneuvers are not part of the IGA core optimization loop. Estimation of target acceleration is always imprecise and noisy and degrade the performance of the two-Loop solution. The IGA trajectories are such that minimize the impact of radome errors in the guidance loop. Finally, as a fourth contribution, it is demonstrated that the missile with IGA guidance is capable of performing a defense against attacks from its rear hemisphere, as a tail attack, only with aerodynamic control. The studied trajectories have a preprogrammed high rate turn maneuver, maintaining the missile within its controllable envelope. This solution does not recur to more complex features in service today, like vector control of the missile thrust or side thrusters. In all the mathematical treatments and demonstrations, the Kronecker product has been introduced as a practical tool to handle the state dependent parametrizations that have resulted in very high order matrix equations.

Characterization of genes modulated during pheomelanogenesis using differential display

Molecular and biochemical mechanisms that modulate the production of eumelanin or pheomelanin pigments involve the opposing effects of two intercellular signaling molecules, α-melanocyte stimulating hormone (MSH) and agouti signal protein (ASP). ASP is an antagonist of MSH signaling through the melanocyte-specific MSH receptor, although its mechanism(s) of action is controversial. We previously have reported significant down-regulation of all known melanogenic genes during the eumelanin to pheomelanin switch in murine hair follicle melanocytes and in cultured melanocytes treated with recombinant ASP. To identify factors that might be involved in the switch to pheomelanogenesis, we screened ASP-treated melanocytes by using differential display and identified three up-regulated genes: a DNA replication control protein, a basic helix–loop–helix transcription factor, and a novel gene. We have simultaneously identified six down-regulated genes in ASP-treated melanocytes; two of those encode tyrosinase and TRP2, melanogenic genes known to be down-regulated during pheomelanogenesis, which provide good internal controls for this approach. These results suggest that there are complex mechanisms involved in the switch to pheomelanin production, and that these modulated genes might be involved in the pleiotropic changes seen in yellow mice, including the change in coat color.

Effects of Age on the Posttranscriptional Regulation of CCAAT/Enhancer Binding Protein α and CCAAT/Enhancer Binding Protein β Isoform Synthesis in Control and LPS-Treated Livers

The CCAAT/enhancer binding protein α (C/EBPα) and CCAAT/enhancer binding protein β (C/EBPβ) mRNAs are templates for the differential translation of several isoforms. Immunoblotting detects C/EBPαs with molecular masses of 42, 38, 30, and 20 kDa and C/EBPβs of 35, 20, and ∼8.5 kDa. The DNA-binding activities and pool levels of p42C/EBPα and p30C/EBPα in control nuclear extracts decrease significantly whereas the binding activity and protein levels of the 20-kDa isoforms increase dramatically with LPS treatment. Our studies suggest that the LPS response involves alternative translational initiation at specific in-frame AUGs, producing specific C/EBPα and C/EBPβ isoform patterns. We propose that alternative translational initiation occurs by a leaky ribosomal scanning mechanism. We find that nuclear extracts from normal aged mouse livers have decreased p42C/EBPα levels and binding activity, whereas those of p20C/EBPα and p20C/EBPβ are increased. However, translation of 42-kDa C/EBPα is not down-regulated on polysomes, suggesting that aging may affect its nuclear translocation. Furthermore, recovery of the C/EBPα- and C/EBPβ-binding activities and pool levels from an LPS challenge is delayed significantly in aged mouse livers. Thus, aged livers have altered steady-state levels of C/EBPα and C/EBPβ isoforms. This result suggests that normal aging liver exhibits characteristics of chronic stress and a severe inability to recover from an inflammatory challenge.

Two gonadotropin-releasing hormone receptor subtypes with distinct ligand selectivity and differential distribution in brain and pituitary in the goldfish (Carassius auratus)

In the goldfish (Carassius auratus) the two endogenous forms of gonadotropin-releasing hormone (GnRH), namely chicken GnRH II ([His5,Trp7,Tyr8]GnRH) and salmon GnRH ([Trp7,Leu8]GnRH), stimulate the release of both gonadotropins and growth hormone from the pituitary. This control is thought to occur by means of the stimulation of distinct GnRH receptors. These receptors can be distinguished on the basis of differential gonadotropin and growth hormone releasing activities of naturally occurring GnRHs and GnRHs with variant amino acids in position 8. We have cloned the cDNAs of two GnRH receptors, GfA and GfB, from goldfish brain and pituitary. Although the receptors share 71% identity, there are marked differences in their ligand selectivity. Both receptors are expressed in the pituitary but are differentially expressed in the brain, ovary, and liver. Thus we have found and cloned two full-length cDNAs that appear to correspond to different forms of GnRH receptor, with distinct pharmacological characteristics and tissue distribution, in a single species.

Combinatorial roles for pRB, p107, and p130 in E2F-mediated cell cycle control

Numerous studies have implicated the pRB family of nuclear proteins in the control of cell cycle progression. Although over-expression experiments have revealed that each of these proteins, pRB, p107, and p130, can induce a G1 cell cycle arrest, mouse knockouts demonstrated distinct developmental requirements for these proteins, as well as partial functional redundancy between family members. To study the mechanism by which the closely related pRB family proteins contribute to cell cycle progression, we generated 3T3 fibroblasts derived from embryos that lack one or more of these proteins (pRB−/−, p107−/−, p130−/−, pRB−/−/p107−/−, pRB−/−/p130−/−, and p107−/−/p130−/−). By comparing the growth and cell cycle characteristics of these cells, we have observed clear differences in the manner in which they transit through the G1 and S phases as well as exit from the cell cycle. Deletion of Rb, or more than one of the family members, results in a shortening of G1 and a lengthening of S phase, as well as a reduction in growth factor requirements. In addition, the individual cell lines showed differential regulation of a subset of E2F-dependent gene promoters, as well as differences in cell cycle-dependent kinase activity. Taken together, these observations suggest that the closely related pRB family proteins affect cell cycle progression through distinct biochemical mechanisms and that their coordinated action may contribute to their diverse functions in various physiological settings.

Differential gene expression in p53-mediated apoptosis-resistant vs. apoptosis-sensitive tumor cell lines

Induction of wild-type p53 in the ECV-304 bladder carcinoma cell line by infection with a p53 recombinant adenovirus (Ad5CMV-p53) resulted in extensive apoptosis and eventual death of nearly all of the cells. As a strategy to determine the molecular events important to p53-mediated apoptosis in these transformed cells, ECV-304 cells were selected for resistance to p53 by repeated infections with Ad5CMV-p53. We compared the expression of 5,730 genes in p53-resistant (DECV) and p53-sensitive ECV-304 cells by reverse transcription–PCR, Northern blotting, and DNA microarray analysis. The expression of 480 genes differed by 2-fold or more between the two p53-infected cell lines. A number of potential targets for p53 were identified that play roles in cell cycle regulation, DNA repair, redox control, cell adhesion, apoptosis, and differentiation. Proline oxidase, a mitochondrial enzyme involved in the proline/pyrroline-5-carboxylate redox cycle, was up-regulated by p53 in ECV but not in DECV cells. Pyrroline-5-carboxylate (P5C), a proline-derived metabolite generated by proline oxidase, inhibited the proliferation and survival of ECV-304 and DECV cells and induced apoptosis in both cell lines. A recombinant proline oxidase protein tagged with a green fluorescent protein at the amino terminus localized to mitochondria and induced apoptosis in p53-null H1299 non-small cell lung carcinoma cells. The results directly implicate proline oxidase and the proline/P5C pathway in p53-induced growth suppression and apoptosis.

Alkaline-mediated differential interaction (AMDI): A simple automatable single-nucleotide polymorphism assay

The key requirements for high-throughput single-nucleotide polymorphism (SNP) typing of DNA samples in large-scale disease case-control studies are automatability, simplicity, and robustness, coupled with minimal cost. In this paper we describe a fluorescence technique for the detection of SNPs that have been amplified by using the amplification refractory mutation system (ARMS)-PCR procedure. Its performance was evaluated using 32 sequence-specific primer mixes to assign the HLA-DRB alleles to 80 lymphoblastoid cell line DNAs chosen from our database for their diversity. All had been typed previously by alternative methods, either direct sequencing or gel electrophoresis. We believe the detection system that we call AMDI (alkaline-mediated differential interaction) satisfies the above criteria and is suitable for general high-throughput SNP typing.

Feedback Control and Diurnal Regulation of Gibberellin 20-Oxidase Transcript Levels in Potato1

Tuber formation in potato (Solanum tuberosum) is promoted by short photoperiods and is inhibited by gibberellins (GAs). Endogenous levels of GA1 were shown to decrease in stolons and leaves of potato plants induced to tuberize, which suggests that photoperiodic regulation of GA biosynthesis may play a role in tuber induction. We report the isolation of three potato cDNA clones (StGA20ox1–3) encoding GA 20-oxidase, a key regulatory enzyme in the GA-biosynthetic pathway. Using northern analysis, we detected a differential pattern of tissue-specific expression of the mRNAs corresponding to these clones. StGA20ox mRNAs were also very abundant in leaves of the potato ga1 mutant, which is blocked in the 13-hydroxylation step, and were strongly down-regulated by gibberellic acid, suggesting a feedback regulation of these genes. In plants grown in short-day (inductive) conditions, levels of the StGA20ox transcripts in leaves fluctuated during a 24-h period, with a peak of accumulation observed about 4 h after the lights were turned off. Interruption of the night with a 30-min “night break” of light (noninductive conditions) did not have a marked effect on the levels of accumulation of the three GA 20-oxidase mRNAs during the day, but it induced a second peak of expression of StGA20ox1 and StGA20ox3 transcripts late in the night. This observation, together with the finding that StGA20ox1 mRNA is expressed at high levels in leaves, suggests that night-break induction of this gene might play a role in the control of tuberization by regulating endogenous levels of GAs in response to daylength conditions.

Coordinating DNA replication initiation with cell growth: differential roles for DnaA and SeqA proteins.

We describe here the development of a new approach to the analysis of Escherichia coli replication control. Cells were grown at low growth rates, in which case the bacterial cell cycle approximates that of eukaryotic cells with G1, S, and G2 phases: cell division is followed sequentially by a gap period without DNA replication, replication of the single chromosome, another gap period, and finally the next cell division. Flow cytometry of such slowly growing cells reveals the timing of replication initiation as a function of cell mass. The data show that initiation is normally coupled to cell physiology extremely tightly: the distribution of individual cell masses at the time of initiation in wild-type cells is very narrow, with a coefficient of variation of less than 9%. Furthermore, a comparison between wild-type and seqA mutant cells shows that initiation occurs at a 10-20% lower mass in the seqA mutant, providing direct evidence that SeqA is a bona fide negative regulator of replication initiation. In dnaA (Ts) mutants the opposite is found: the mass at initiation is dramatically increased and the variability in cell mass at initiation is much higher than that for wild-type cells. In contrast to wild-type and dnaA(Ts) cells, seqA mutant cells frequently go through two initiation events per cell division cycle, and all the origins present in each cell are not initiated in synchrony. The implications for the complex interplay amongst growth, cell division, and DNA replication are discussed.

Locust adipokinetic hormones: carrier-independent transport and differential inactivation at physiological concentrations during rest and flight.

Since concomitant release of structurally related peptide hormones with apparently similar functions seems to be a general concept in endocrinology, we have studied the dynamics of the lifetime of the three known adipokinetic hormones (AKHs) of the migratory locust, which control flight-directed mobilization of carbohydrate and lipid from fat body stores. Although the structure of the first member of the AKHs has been known for 20 years, until now, reliable data on their inactivation and removal from the hemolymph are lacking, because measurement requires AKHs with high specific radioactivity. Employing tritiated AKHs with high specific radioactivity, obtained by catalytic reduction with tritium gas of the dehydroLeu2 analogues of the AKHs synthesized by the solid-phase procedure, studies with physiological doses of as low as 1.0 pmol per locust could be conducted. The AKHs appear to be transported in the hemolymph in their free forms and not associated with a carrier protein, despite their strong hydrophobicity. Application of AKHs in their free form in in vivo and in vitro studies therefore now has been justified. We have studied the degradation of the three AKHs during rest and flight. The first cleavage step by an endopeptidase is crucial, since the resulting degradation products lack any adipokinetic activity. Half-lives for AKH-I, -II and -III were 51, 40, and 5 min, respectively, for rest conditions and 35, 37, and 3 min, respectively, during flight. The rapid and differential degradation of structurally related hormones leads to changes in the ratio in which they are released and therefore will have important consequences for concerted hormone action at the level of the target organ or organs, suggesting that each of the known AKHs may play its own biological role in the overall syndrome of insect flight.

Mast cells express a peripheral cannabinoid receptor with differential sensitivity to anandamide and palmitoylethanolamide.

Mast cells are multifunctional bone marrow-derived cells found in mucosal and connective tissues and in the nervous system, where they play important roles in tissue inflammation and in neuroimmune interactions. Very little is known about endogenous molecules and mechanisms capable of modulating mast cell activation. Palmitoylethanolamide, found in peripheral tissues, has been proposed to behave as a local autacoid capable of downregulating mast cell activation and inflammation. A cognate N-acylamide, anandamide, the ethanolamide of arachidonic acid, occurs in brain and is a candidate endogenous agonist for the central cannabinoid receptor (CB1). As a second cannabinoid receptor (CB2) has been found in peripheral tissues, the possible presence of CB2 receptors on mast cells and their interaction with N-acylamides was investigated. Here we report that mast cells express both the gene and a functional CB2 receptor protein with negative regulatory effects on mast cell activation. Although both palmitoylethanolamide and anandamide bind to the CB2 receptor, only the former downmodulates mast cell activation in vitro. Further, the functional effect of palmitoylethanolamide, as well as that of the active cannabinoids, was efficiently antagonized by anandamide. The results suggest that (i) peripheral cannabinoid CB2 receptors control, upon agonist binding, mast cell activation and therefore inflammation; (ii) palmitoylethanolamide, unlike anandamide, behaves as an endogenous agonist for the CB2 receptor on mast cells; (iii) modulatory activities on mast cells exerted by the naturally occurring molecule strengthen a proposed autacoid local inflammation antagonism (ALIA) mechanism; and (iv) palmitoylethanolamide and its derivatives may provide antiinflammatory therapeutic strategies specifically targeted to mast cells ("ALIAmides").

Differential modulation of Th1- and Th2-related cytokine mRNA expression by a synthetic peptide homologous to a conserved domain within retroviral envelope protein.

The influence of a synthetic retroviral peptide, CKS-17, on T helper type 1 (Th1)- or Th2-related cytokines was investigated in human blood mononuclear cells. Cells were stimulated with staphylococcal enterotoxin A, anti-CD3 plus anti-CD28 monoclonal antibodies, or lipopolysaccharide to induce cytokine mRNA. mRNA was detected by a reverse transcription-polymerase chain reaction or Northern blot analysis. CKS-17 down-regulated stimulant-induced mRNA accumulation for interferon gamma (IFN-gamma), interleukin (IL)-2, and p40 heavy and p35 light chains of IL-12, a cytokine that mediates development of Th1 response. CKS-17 up-regulated stimulant-induced mRNA accumulation of IL-10 and did not suppress Th2-related cytokine (IL-4, IL-5, IL-6, or IL-13) mRNA expression. A reverse sequence of CKS-17 peptide, used as a control, showed no such action. Anti-human IL-10 monoclonal antibody blocked ability of CKS-17 to inhibit mRNA accumulation for IFN-gamma but not the CKS-17 suppressive activity of IL-12 p40 heavy chain mRNA. Thus, CKS-17-mediated suppression of IFN-gamma mRNA expression is dependent upon augmentation of IL-10 production by CKS-17. This conserved component of several retroviral envelope proteins, CKS-17, may act as an immunomodulatory epitope responsible for cytokine dysregulation that leads to suppression of cellular immunity.

Control and navigation of a rover for agricultural purposes

L’obiettivo di questa tesi è di descrivere e implementare via software un modello di rover autonomo per uso in ambito agricolo. La scelta di questo argomento deriva dal fatto che al laboratorio CASY dell’Università di Bologna è stato commissionato un robot che possa aiutare piccoli imprenditori agricoli a essere competitivi con i più grandi. Le funzionalità che il robot avrà, una volta ultimato, andranno dal tagliare l’erba allo spruzzare fertilizzante sugli alberi da frutto. Questa tesi si interessa del progetto del sistema di navigazione. Inizialmente viene introdotto il modello cinematico e in particolare la configurazione differential drive in cui il rover rientra. Successivamente viene elaborato un sistema di controllo basato sulla linearizzazione statica del feedback. Una volta completati il modello e il sistema di controllo si procede con la generazione di traiettoria: vengono analizzati e confrontati alcuni algoritmi per l’inseguimento di una traiettoria definita tramite waypoint. Infine è presentato un algoritmo per la navigazione all’interno di un campo di filari di alberi da frutto. Le uniche informazioni esterne disponibili in questo contesto sono le rilevazioni di sensori di distanza frontali e laterali, in quanto un GPS sarebbe troppo impreciso per gli scopi. Questa tesi costituisce la base per ulteriori sviluppi del progetto. In particolare la realizzazione di un programma di supervisione che stabilisca la modalità di moto da attuare e programmi specifici per le varie funzionalità agricole del rover.