Molecular studies on a Dictyostelium homolog of the tafazzin gene, the cause of Barth syndrome in humans

Der eukaryotische Mikroorganismus Dictyostelium discoideum lebt als einzellige Amöbe solange ausreichende Nahrungsressourcen zur Verfügung stehen. Sobald Nahrungsmangel eintritt, entwickeln sich die Zellen von einem einzelligen zu einem mehrzelligen Zustand, der mit einem multizellulären Fruchtkörper abschließt. Dieser Prozess wird durch eine Reihe aufeinanderfolgender Signale organisiert, die eine differentielle Genexpression regulieren. Die Gene der Discoidin I Familie gehören zu den Ersten, die im Laufe des Wachstums-Differenzierungs-Übergangs (engl. GDT) aktiviert werden. Sie eignen sich daher vorzüglich als Marker für den Beginn der Entwicklung. Mit Hilfe einer REMI-Mutagenese und Discoidin I als molekularem Marker sind verschiedene Komponenten des Wachstums-Differenzierungs-Übergangs in unserer Arbeitsgruppe identifiziert worden (Zeng et al., 2000 A und B; Riemann und Nellen, persönliche Mitteilung). Mit demselben Ansatz wurde in der vorliegenden Arbeit eine REMI-Mutante identifiziert, die eine Fehl-Expression von Discoidin zeigte und einen axenischen Wachstumsdefekt bei 15 °C aufwies. Das Gen wurde als Homolog zum humanen Tafazzin-Gen identifiziert. Dieses Gen wurde zur Rekonstruktion des Phänotyps über homologe Rekombination erneut disruptiert, was wie erwartet zu dem zuerst beschriebenen Phänotyp führte. Folgerichtig ergab eine Überexpression des Gens in den Mutanten eine Komplementation des Phänotyps. Immunfluoreszenz-Experimente zeigten eine mitochondriale Lokalisation des Dictyostelium discoideum Taffazzin Proteins. Dass ein mitochondriales Protein in Zusammenhang mit dem Wachstums-Differenzierungs-Übergang steht, ist ein unerwarteter Befund, der aber als Hinweis darauf gewertet werden kann, dass Mitochondrien einen direkten Einfluss auf die entwicklungsspezifische Signaltransduktion ausüben. Die Taffazzin Disruptions-Mutante in Dictyostelium führte zu einem abnormalen Cardiolipin Metabolismus. Dieses Phospholipid ist ein charakteristischer Bestandteil der inneren Mitochondrienmembran und für die Funktion verschiedener Enzyme erforderlich. Unsere vorläufigen Analysen des Phospholipid-Gehalts zeigten Übereinstimmung mit Daten von Patienten mit Barth-Syndrom, einer humanen Erkrankung, bei der das Taffazzin-Gen Mutationen aufweist, und mit Hefe-Mutanten dieses Gens. Dies zeigt den Wert von Dictyostelium discoideum als einen weiteren Modelorganismus zur Untersuchung des Barth-Syndroms und zur Erprobung möglicher Therapieansätze.

Interaktive Netzwerke zur Regulation des Wachstums-Differenzierungs-Übergangs bei Dictyostelium discoideum

Die Signaltransduktion in niederen und höheren Zellen gehört zu einem der intensivst beforschten, molekularen Mechanismen. Wie gelangt ein externer Stimulus in die Zelle, bzw. wie wird das entsprechende Signal von der Zelloberfläche in das Zellinnere übertragen? Welche Proteine, die in die Signaltransduktion involviert sind, benötigt die Zelle um auf diesen Stimulus zu reagieren – und wie reagiert die Zelle letztendlich auf dieses extrazelluläre Signal? In den letzten Jahren wurde deutlich, dass diese interaktiven Netzwerke hochkomplex sind und für die molekularbiologische Forschung nur dann einsehbar werden, wenn gezielt Mutanten hergestellt werden, die z.B. Rezeptoren oder interzelluläre Komponenten nicht mehr vorweisen können. Die Erforschung der Signaltransduktionsprozesse ist mittlerweile aus den Laboren der Grundlagenforschung auch in die molekularbiologischen Labors der pharmazeutischen Forschung übertragen worden. Aktuell wurden in den letzten Jahren mehrere Substanzen entwickelt, die z.B. für die Bekämpfung von bösartigen Tumoren geeignet sind, und diese Substanzen zeichnen sich dadurch aus, dass sie Komponenten der Signaltransduktion blockieren, bzw. Botenstoffe der Neoangiogenese aus dem Serum entfernen und so den Tumor „aushungern“. In Dictyostelium discoideum sind bereits zahlreiche Signaltransduktionskomponenten beschrieben worden und es finden sich die bekannten Systeme, wie z.B. Transmembranrezeptoren, G-Proteine oder ras-Proteine, die man aus anderen Organismen kennt wieder. Auch MAP-Kinase-Kaskaden sind vorhanden und verschiedene extrazelluläre Signalstoffe, wie z.B. cAMP, PSF oder CMF sind bekannt und teilweise charakterisiert. Dictyostelium discoideum eignet sich aus diesen Gründen und aus Gründen der biochemischen und zellbiologischen Verfügbarkeit dazu, Prozesse der Signalerkennung und deren Weiterleitung zu den Effektorproteinen zu erforschen. Das Primärziel dieser Arbeit war es, möglichst eine neue Komponente in einem der bereits bekannten Signalwege der Discoidin-Regulation durch Mutagenesen zu zerstören, um diese anschließend beschreiben und charakterisieren zu können.Dazu wurde die sog. REMI-Mutagenese am neu gegründeten Labor der Universität Kassel etabliert und ferner die Zellkulturtechnik von D. discoideum für den Routineeinsatz eingearbeitet. Eine weitere Aufgabe der vorliegenden Arbeit war das Screening bereits bekannter Zellinien, die durch einen auffälligen Phänotyp nach einer Mutagenese isoliert worden waren. Dieses Screening sollte mit Western-blot-Analysen des sog. Discoidin-Proteins durchgeführt werden. Zusätzlich sollten neue Methoden entwickelt werden, die es möglich machen die Interaktionen während des vegetativen Wachstums vom Dictyostelium in Klebsiella-Suspension zu beschreiben.

Cirugía de epilepsia

Capítulo sobre Cirugía de Epilepsia del libro decisiones en neurología en su segunda edición de la editorial de la universidad del rosario. El capitulo pretende guiar al lector en que momentos los pacientes con epilepsia refractaria son buenos candidatos para manejo quirúrgico.

Convergencia de vías de señalización en un modelo de apoptosis

La sepsis es un evento inflamatorio generalizado del organismo inducido por un daño causado generalmente por un agente infeccioso. El patógeno más frecuentemente asociado con esta entidad es el Staphylococcus aureus, responsable de la inducción de apoptosis en células endoteliales debida a la producción de ceramida. Se ha descrito el efecto protector de la proteína C activada (PCA) en sepsis y su relación con la disminución de la apoptosis de las células endoteliales. En este trabajo se analizó la activación de las quinasas AKT, ASK1, SAPK/JNK y p38 en un modelo de apoptosis endotelial usando las técnicas de Western Blotting y ELISA. Las células endoteliales (EA.hy926), se trataron con C2-ceramida (130μM) en presencia de inhibidores químicos de cada una de estas quinasas y PCA. La supervivencia de las células en presencia de inhibidores químicos y PCA fue evaluada por medio de ensayos de activación de las caspasas 3, 7 y 9, que verificaban la muerte celular por apoptosis. Los resultados evidencian que la ceramida reduce la activación de AKT y aumenta la activación de las quinasas ASK, SAPK/JNK y p38, en tanto que PCA ejerce el efecto contrario. Adicionalmente se encontró que la tiorredoxina incrementa la activación/fosforilación de AKT, mientras que la quinasa p38 induce la defosforilación de AKT.

La cooperación hacia Mauritania y la externalización del control migratorio español como mecanismos de securitización de la política migratoria española hacia Mauritania (1996 - 2013)

La llegada masiva de migrantes irregulares provenientes del África sub-sahariana ha sido un fenómeno que ha afectado en términos económicos, sociales, políticos y culturales, a los países que conforman la Unión Europea. Es así como los países más afectados por este tipo de migración (España, Grecia e Italia) han pedido a los demás miembros de la Unión adoptar una política conjunta y armonizada con el fin de solucionar la problemática migratoria irregular. No obstante, la cooperación, como acción unilateral, ha resultado ser el mecanismo más efectivo, en especial el caso español, para contener y externalizar los controles del movimiento migratorio irregular subsahariano en países como Mauritania, como resultado de un proceso de securitización de la migración irregular al interior de España a comienzos del siglo XX.

On the energetics of stratified turbulent mixing, irreversible thermodynamics, Boussinesq models and the ocean heat engine controversy

In this paper, the available potential energy (APE) framework of Winters et al. (J. Fluid Mech., vol. 289, 1995, p. 115) is extended to the fully compressible Navier– Stokes equations, with the aims of clarifying (i) the nature of the energy conversions taking place in turbulent thermally stratified fluids; and (ii) the role of surface buoyancy fluxes in the Munk & Wunsch (Deep-Sea Res., vol. 45, 1998, p. 1977) constraint on the mechanical energy sources of stirring required to maintain diapycnal mixing in the oceans. The new framework reveals that the observed turbulent rate of increase in the background gravitational potential energy GPEr , commonly thought to occur at the expense of the diffusively dissipated APE, actually occurs at the expense of internal energy, as in the laminar case. The APE dissipated by molecular diffusion, on the other hand, is found to be converted into internal energy (IE), similar to the viscously dissipated kinetic energy KE. Turbulent stirring, therefore, does not introduce a new APE/GPEr mechanical-to-mechanical energy conversion, but simply enhances the existing IE/GPEr conversion rate, in addition to enhancing the viscous dissipation and the entropy production rates. This, in turn, implies that molecular diffusion contributes to the dissipation of the available mechanical energy ME =APE +KE, along with viscous dissipation. This result has important implications for the interpretation of the concepts of mixing efficiency γmixing and flux Richardson number Rf , for which new physically based definitions are proposed and contrasted with previous definitions. The new framework allows for a more rigorous and general re-derivation from the first principles of Munk & Wunsch (1998, hereafter MW98)’s constraint, also valid for a non-Boussinesq ocean: G(KE) ≈ 1 − ξ Rf ξ Rf Wr, forcing = 1 + (1 − ξ )γmixing ξ γmixing Wr, forcing , where G(KE) is the work rate done by the mechanical forcing, Wr, forcing is the rate of loss of GPEr due to high-latitude cooling and ξ is a nonlinearity parameter such that ξ =1 for a linear equation of state (as considered by MW98), but ξ <1 otherwise. The most important result is that G(APE), the work rate done by the surface buoyancy fluxes, must be numerically as large as Wr, forcing and, therefore, as important as the mechanical forcing in stirring and driving the oceans. As a consequence, the overall mixing efficiency of the oceans is likely to be larger than the value γmixing =0.2 presently used, thereby possibly eliminating the apparent shortfall in mechanical stirring energy that results from using γmixing =0.2 in the above formula.

Understanding mixing efficiency in the oceans. Do the nonlinearities of the equation of state matter?

There exist two central measures of turbulent mixing in turbulent stratified fluids that are both caused by molecular diffusion: 1) the dissipation rate D(APE) of available potential energy APE; 2) the turbulent rate of change Wr, turbulent of background gravitational potential energy GPEr. So far, these two quantities have often been regarded as the same energy conversion, namely the irreversible conversion of APE into GPEr, owing to the well known exact equality D(APE)=Wr, turbulent for a Boussinesq fluid with a linear equation of state. Recently, however, Tailleux (2009) pointed out that the above equality no longer holds for a thermally-stratified compressible, with the ratio ξ=Wr, turbulent/D(APE) being generally lower than unity and sometimes even negative for water or seawater, and argued that D(APE) and Wr, turbulent actually represent two distinct types of energy conversion, respectively the dissipation of APE into one particular subcomponent of internal energy called the "dead" internal energy IE0, and the conversion between GPEr and a different subcomponent of internal energy called "exergy" IEexergy. In this paper, the behaviour of the ratio ξ is examined for different stratifications having all the same buoyancy frequency N vertical profile, but different vertical profiles of the parameter Υ=α P/(ρCp), where α is the thermal expansion coefficient, P the hydrostatic pressure, ρ the density, and Cp the specific heat capacity at constant pressure, the equation of state being that for seawater for different particular constant values of salinity. It is found that ξ and Wr, turbulent depend critically on the sign and magnitude of dΥ/dz, in contrast with D(APE), which appears largely unaffected by the latter. These results have important consequences for how the mixing efficiency should be defined and measured in practice, which are discussed.

The effect of mechanical stirring on horizontal convection

An important experimental result, as yet poorly understood, is that mechanical stirring can significantly enhance the strength of horizontal convection. A contentious issue is whether this necessarily implies that the mechanical stirring replaces the buoyancy forcing as the main source of energy driving the observed overturning circulation, as has been suggested for the Atlantic meridional overturning circulation (AMOC). In this paper, rigorous energetics considerations and idealized numerical experiments reveal that the rate at which the surface buoyancy forcing supplies energy to the fluid, as measured by the production rate of available potential energy G(APE), does not solely depend upon the buoyancy forcing, as is often implicitly assumed, but also upon the vertical stratification, such that the deeper the thermocline depth, the larger G(APE). This suggests that mechanical stirring enhances horizontal convection because it causes more energy to be extracted from the buoyancy forcing. It does so by enhancing turbulent mixing, which allows surface heating to reach greater depths, which increases the thermocline depth and hence G(APE). This paper therefore proposes a new hypothesis, namely that mechanically stirred horizontal convection and the AMOC are best described as mechanically controlled heat engines.

Climate entropy budget of the HadCM3 atmosphere-ocean general circulation model and of FAMOUS, its low resolution version

The entropy budget is calculated of the coupled atmosphere–ocean general circulation model HadCM3. Estimates of the different entropy sources and sinks of the climate system are obtained directly from the diabatic heating terms, and an approximate estimate of the planetary entropy production is also provided. The rate of material entropy production of the climate system is found to be ∼50 mW m−2 K−1, a value intermediate in the range 30–70 mW m−2 K−1 previously reported from different models. The largest part of this is due to sensible and latent heat transport (∼38 mW m−2 K−1). Another 13 mW m−2 K−1 is due to dissipation of kinetic energy in the atmosphere by friction and Reynolds stresses. Numerical entropy production in the atmosphere dynamical core is found to be about 0.7 mW m−2 K−1. The material entropy production within the ocean due to turbulent mixing is ∼1 mW m−2 K−1, a very small contribution to the material entropy production of the climate system. The rate of change of entropy of the model climate system is about 1 mW m−2 K−1 or less, which is comparable with the typical size of the fluctuations of the entropy sources due to interannual variability, and a more accurate closure of the budget than achieved by previous analyses. Results are similar for FAMOUS, which has a lower spatial resolution but similar formulation to HadCM3, while more substantial differences are found with respect to other models, suggesting that the formulation of the model has an important influence on the climate entropy budget. Since this is the first diagnosis of the entropy budget in a climate model of the type and complexity used for projection of twenty-first century climate change, it would be valuable if similar analyses were carried out for other such models.

Entropy versus APE production: on the buoyancy power input in the oceans energy cycle

This letter argues that the current controversy about whether Wbuoyancy, the power input due to the surface buoyancy fluxes, is large or small in the oceans stems from two distinct and incompatible views on how Wbuoyancy relates to the volume-integrated work of expansion/contraction B. The current prevailing view is that Wbuoyancy should be identified with the net value of B, which current theories estimate to be small. The alternative view, defended here, is that only the positive part of B, i.e., the one converting internal energy into mechanical energy, should enter the definition of Wbuoyancy, since the negative part of B is associated with the non-viscous dissipation of mechanical energy. Two indirect methods suggest that by contrast, the positive part of B is potentially large.

High frequency variability of the Atlantic meridional overturning circulation

We compare the variability of the Atlantic meridional overturning circulation (AMOC) as simulated by the coupled climate models of the RAPID project, which cover a wide range of resolution and complexity, and observed by the RAPID/MOCHA array at about 26N. We analyse variability on a range of timescales, from five-daily to interannual. In models of all resolutions there is substantial variability on timescales of a few days; in most AOGCMs the amplitude of the variability is of somewhat larger magnitude than that observed by the RAPID array, while the time-mean is within about 10% of the observational estimate. The amplitude of the simulated annual cycle is similar to observations, but the shape of the annual cycle shows a spread among the models. A dynamical decomposition shows that in the models, as in observations, the AMOC is predominantly geostrophic (driven by pressure and sea-level gradients), with both geostrophic and Ekman contributions to variability, the latter being exaggerated and the former underrepresented in models. Other ageostrophic terms, neglected in the observational estimate, are small but not negligible. The time-mean of the western boundary current near the latitude of the RAPID/MOCHA array has a much wider model spread than the AMOC does, indicating large differences among models in the simulation of the wind-driven gyre circulation, and its variability is unrealistically small in the models. In many RAPID models and in models of the Coupled Model Intercomparison Project Phase 3 (CMIP3), interannual variability of the maximum of the AMOC wherever it lies, which is a commonly used model index, is similar to interannual variability in the AMOC at 26N. Annual volume and heat transport timeseries at the same latitude are well-correlated within 15--45N, indicating the climatic importance of the AMOC. In the RAPID and CMIP3 models, we show that the AMOC is correlated over considerable distances in latitude, but not the whole extent of the north Atlantic; consequently interannual variability of the AMOC at 50N, where it is particularly relevant to European climate, is not well-correlated with that of the AMOC at 26N, where it is monitored by the RAPID/MOCHA array.

The key role of the western boundary in linking the AMOC strength to the north-south pressure gradient

A key idea in the study of the Atlantic meridional overturning circulation (AMOC) is that its strength is proportional to the meridional density gradient, or more precisely, to the strength of the meridional pressure gradient. A physical basis that would tell us how to estimate the relevant meridional pressure gradient locally from the density distribution in numerical ocean models to test such an idea, has been lacking however. Recently, studies of ocean energetics have suggested that the AMOC is driven by the release of available potential energy (APE) into kinetic energy (KE), and that such a conversion takes place primarily in the deep western boundary currents. In this paper, we develop an analytical description linking the western boundary current circulation below the interface separating the North Atlantic Deep Water (NADW) and Antarctic Intermediate Water (AAIW) to the shape of this interface. The simple analytical model also shows how available potential energy is converted into kinetic energy at each location, and that the strength of the transport within the western boundary current is proportional to the local meridional pressure gradient at low latitudes. The present results suggest, therefore, that the conversion rate of potential energy may provide the necessary physical basis for linking the strength of the AMOC to the meridional pressure gradient, and that this could be achieved by a detailed study of the APE to KE conversion in the western boundary current.

Vertical and horizontal processes in the global atmosphere and the maximum entropy production conjecture

The objective of this paper is to reconsider the Maximum Entropy Production conjecture (MEP) in the context of a very simple two-dimensional zonal-vertical climate model able to represent the total material entropy production due at the same time to both horizontal and vertical heat fluxes. MEP is applied first to a simple four-box model of climate which accounts for both horizontal and vertical material heat fluxes. It is shown that, under condition of fixed insolation, a MEP solution is found with reasonably realistic temperature and heat fluxes, thus generalising results from independent two-box horizontal or vertical models. It is also shown that the meridional and the vertical entropy production terms are independently involved in the maximisation and thus MEP can be applied to each subsystem with fixed boundary conditions. We then extend the four-box model by increasing its resolution, and compare it with GCM output. A MEP solution is found which is fairly realistic as far as the horizontal large scale organisation of the climate is concerned whereas the vertical structure looks to be unrealistic and presents seriously unstable features. This study suggest that the thermal meridional structure of the atmosphere is predicted fairly well by MEP once the insolation is given but the vertical structure of the atmosphere cannot be predicted satisfactorily by MEP unless constraints are imposed to represent the determination of longwave absorption by water vapour and clouds as a function of the state of the climate. Furthermore an order-of-magnitude estimate of contributions to the material entropy production due to horizontal and vertical processes within the climate system is provided by using two different methods. In both cases we found that approximately 40 mW m−2 K−1 of material entropy production is due to vertical heat transport and 5–7 mW m−2 K−1 to horizontal heat transport