NOVEL THERAPEUTIC TARGETS IDENTIFIED BY HIGH-THROUGHPUT TECHNOLOGIES FOR TRIPLE-NEGATIVE BREAST CANCER

Triple-negative breast cancers (TNBC) are characterized by the lack of or reduced expression of the estrogen and progesterone receptors, and normal expression of the human epidermal growth factor receptor 2. The lack of a well-characterized target for treatment leaves only systemic chemotherapy as the mainstay of treatment. Approximately 60-70% of patients are chemosensitive, while the remaining majority does not respond. Targeted therapies that take advantage of the unique molecular perturbations found in triple-negative breast cancer are needed. The genes that are frequently amplified or overexpressed represent potential therapeutic targets for triple-negative breast cancer. The purpose of this study was to identify and validate novel therapeutic targets for triple-negative breast cancers. 681 genes showed consistent and highly significant overexpression in TNBC compared to receptor-positive cancers in 2 data sets. For two genes, 3 of the 4 siRNAs showed preferential growth inhibition in TNBC cells. These two genes were the low density lipoprotein receptor-related protein 8 (LRP8) and very low-density lipoprotein receptor (VLDLR). Exposure to their cognate ligands, reelin and apolipoprotein E isoform 4 (ApoE4), stimulated the growth of TNBC cells in vitro. Suppression of the expression of either LRP8 or VLDLR or exposure to RAP (an inhibitor of ligand binding to LRP8 and VLDLR) abolished this ligand-induced proliferation. High-throughput protein and metabolic arrays revealed that ApoE4 stimulation rescued TNBC cells from serum-starvation induced up-regulation of genes involved in lipid biosynthesis, increased protein expression of oncogenes involved in the MAPK/ERK and DNA repair pathways, and reduced the serum-starvation induction of biochemicals involved in oxidative stress response and glycolytic metabolism. shLRP8 MDA-MB-231 xenografts had reduced tumor volume, in comparison to parental and shCON xenografts. These results indicate that LRP8-APOE signaling confers survival advantages to TNBC tumors under reduced nutrient conditions and during cellular environmental stress. We revealed that the LRP8-APOE receptor-ligand system is overexpressed in human TNBC. We also demonstrated that this receptor system mediates a strong growth promoting and survival function in TNBC cells in vitro and helps to sustain the growth of MDA-MD-231 xenografts. We propose that inhibitors of LRP8-APOE signaling may be clinically useful therapeutic agents for triple-negative breast cancer.

Regulation of v-Mos kinase activity by phosphorylation

In this thesis, I investigated the effect of cylic AMP-dependent protein kinase (PKA) on v-Mos kinase activity. Increase in PKA activity in vivo brought about either by forskolin treatment or by overexpression of the PKA catalytic subunit resulted in a significant inhibition of v-Mos kinase activity. The purified PKA catalytic subunit was able to phosphorylate recombinant p37$\rm\sp{v-mos}$ in vitro, suggesting that the mechanism of in vivo inhibition of v-Mos kinase involves direct phosphorylation by PKA. Ser-263 was identified as a residue that is normally phosphorylated at a very low level but whose phosphorylation is dramatically increased upon forskolin treatment. Consistent with the inhibitory role of Ser-263 phosphorylation, the Ala-263 mutant of v-Mos was not inhibited by forskolin treatment. Based on our results, we propose that the known inhibitory role of PKA in the initiation of oocyte maturation could be explained at least in part by its inhibition of Mos kinase.^ Combining tryptic phosphopeptide two-dimensional mapping analysis and in vitro mutagenesis studies, I identified Ser-56 as the major in vivo phosphorylation site on v-Mos. I studied the interrelationship between Ser-34 and Ser-56 phosphorylation in regulating v-Mos function. After site-directed mutagenesis to substitute serine residues with alanine or glutamic acid in different combinations to mimick unphosphorylated and phosphorylated serines respectively, various v-Mos mutants were expressed in COS-1 cells. As expected, Ala-34 mutant of v-Mos had very low (less 5% of wild type) kinase activity. The Ala-56 mutant had kinase activity 50% that of wild type. Surprisingly, the Ala-34 Ala-56 double mutant and the Ala-56 mutant exhibited identical kinase activity. On the other hand, Ala-34 Glu-56 double mutant had reduced kinase activity comparable to Ala-34 mutant. These results suggest that the phosphorylation at Ser-56 may serve to inhibit the activation of newly synthesized Mos protein. As predicted from Xenopus c-Mos studies, Glu-34 mutant of v-Mos was highly active (125% that of wild type). Interestingly, consistant with the model involving an inhibitory role of Ser-56 phosphorylation, the Glu-34 Glu-56 double mutant was totally inactive as a kinase. Moreover in my experiments, there was a perfect correlation between the level of v-Mos kinase activity of various mutants and their transforming activity. The latter is dependent upon MEK1 phosphorylation/ activation in v-mos transformed cells. Residues corresponding to both v-Mos Ser-34 and Ser-56 are evolutionarily conserved in c-Mos. Therefore, the cytostatic factor function of c-Mos may be regulated in the same manner as v-Mos kinase activity.^ It has been known that v-mos transforms cells by affecting G1 phase progression of the cell cycle. Here I showed that mos induces cyclin D1 expression in mos transformed NIH 3T3 cells and NRK 6m2 cells, and this induced level was found to be unaffected by serum starvation. Consequently, cyclin D1-Cdk4 and cyclin E-Cdk2 activities increase, and retinoblastoma protein is hyperphosphorylated. Based on studies from several laboratories, these findings suggest that increased amount of cyclin D1-Cdk4 complexes ties up the limited amount of cyclin E-Cdk2 inhibitors (e.g. p27), causing the activation of cyclin E-Cdk2. My results indicate that activation of key cell cycle regulators of G1 phase may be important for cellular transformation by mos. (Abstract shortened by UMI.) ^

The SasS -SasR two-component signal transduction system required for early Myxococcus xanthus multicellular *development

Initiation of Myxococcus xanthus multicellular development requires both nutrient limitation and high cell density. The extracellular signal, A signal, which consists of a set of amino acids at specific concentrations, serves as a cell density signal in M. xanthus early development. A reporter gene, designated 4521, that requires both starvation and A signal for developmental expression was used to identify mutations in the signal transduction pathways. A group of point mutations located in the chromosomal sasB locus that bypasses both requirements was previously isolated. One of these point mutations, sasB7, was mapped to the sasS gene, which is predicted to encode a transmembrane histidine protein kinase required for normal development. SasS is a positive regulator of 4521 and a candidate A signal sensor. This dissertation continues the characterization of the sasB locus, focusing on the sasR gene and the functional relationship of SasS and SasR. ^ The sasR gene is located 2.2-kb downstream of sasS. It is predicted to encode an NtrC-like response regulator, which belongs to the family of sigma54 transcriptional activators. SasR is a positive regulator of 4521 gene and is required for normal development. The sasR mutant displays phenotypes similar to that of sasS mutant. Both SasS and SasR are required for the A-signal-dependent 4521 expression. Genetic epistasis analysis indicates that SasR functions downstream of SasS. Biochemical studies show that SasS has autokinase activity, and phosphorylated SasS is able to transfer its phosphate to SasR. We propose that SasS and SasR form a two-component signal transduction system in the A signal transduction pathway. ^ To search for the genes regulated by SasS and SasR, expression patterns of a group of developmental genes were compared in wild-type and sasS null mutant backgrounds. SasS and SasR were found to positively regulate sasN and 4521. The sasN gene was previously identified as a negative regulator of 4521, located at about 170-bp downstream of sasR. It is required for normal fruiting body development. Based on the above data, a regulatory network consisting of sasS, sasR, sasN, and 4521 is hypothesized, and the interactions of the components in this network can now be further studied. ^

Constitutively active cAMP receptor mutants dominantly inhibit receptor-mediated signaling pathways in Dictyostelium discoideum

Dictyostelium, a soil amoeba, is able to develop from free-living cells to multicellular fruiting bodies upon starvation using extracellular cAMP to mediate cell-cell communication, chemotaxis and developmental gene expression. The seven transmembrane G protein-coupled cAMP receptor-1 (cAR1) mediated responses, such as the activation of adenylyl cyclase and guanylyl cyclase, are transient, due to the existence of poorly understood adaptation mechanisms. For this dissertation, the powerful genetics of the Dictyostelium system was employed to study the adaptation mechanism of cAR1-mediated cAMP signaling as well as mechanisms intrinsic to cAR1 that regulate its activation. ^ We proposed that constitutively active cAR1 would cause constant adaptation, thus inhibiting downstream pathways that are essential for aggregation and development. Therefore, a screen for dominant negative cAR1 mutants was undertaken to identify constitutively active receptor mutants. Three dominant negative cAR1 mutants were identified. All appear to be constitutively active receptor mutants because they are constitutively phosphorylated and possess high affinity for cAMP. Biochemical studies showed that these mutant receptors prevented the activation of downstream effectors, including adenylyl and guanylyl cyclases. In addition, these cells also were defective in cAMP chemotaxis and cAR1-mediated gene expression. These findings suggest that the mutant receptors block development by constantly activating multiple adaptation pathways. ^ Sequence analysis revealed that these mutations (I104N, L100H) are clustered in a conserved region of the third transmembrane helix (TM3) of cAR1. To investigate the role of this region in receptor activation, one of these residues, I104, was mutated to all the other 19 possible amino acids. We found that all but the most conservative substitutions increase the receptor's affinity about 20- to 70-fold. However, only highly polar substitutions of I104, particularly basic residues, resulted in receptors that are constitutively phosphorylated and dominantly inhibit development, suggesting that highly polar substitutions not only disrupt an interaction constraining the receptor in its low-affinity, inactive state but also promote an additional conformational change that resembles the ligand-bound conformation. Our findings suggest that I104 plays a specific role in constraining the receptor in its inactive state and that substituting it with highly polar residues results in constitutive activation. ^

YKL -40, a vascular -associated serum marker for glioblastoma, correlates with chronic activation of AKT

YKL-40 is a secreted glycoprotein that has been reported to be expressed in pathologic conditions of extracellular matrix degradation and angiogenesis, such as rheumatoid arthritis, severe osteoarthritis, primary colorectal cancer, metastatic breast cancer, and recurrent ovarian cancer (Dehn, Hogdall et al. 2003). ^ We have identified YKL-40 as a serum marker for glioblastoma multiforme (GBM) using microarray analysis from samples of GBM. We compared the gene expression profile of 19 gliomas to pooled normal brain tissue using the Incyte 10,000 gene expression array. The most differentially expressed gene in this analysis was YKL-40; it was detected in GBM samples with a range of 3 to 62-fold elevation over normal brain. Western blot analysis of glioma samples for YKL-40 protein levels revealed substantial elevation in approximately 65% of GBMs, and undetectable levels in lower-grade gliomas and normal brain tissue. ELISA analysis on serum samples of glioma patients showed that YKL-40 levels were substantially elevated in many of the GBM patients. Statistical analysis indicated that in patients with glioma, serum YKL-40 levels correlate with tumor grade and potentially tumor burden in GBM. ^ Furthermore, we found that YKL-40 expression by in-situ hybridization on a brain tumor tissue array was limited to GBM's and gliosarcomas (GSA), and that YKL-40 expression was specific to the GBM component of GSA. Additional in-situ hybridization analysis, found it to be regionally associated with tumor vasculature as well as activated AKT expression in both human and mouse GBM's. Correlation of elevated YKL-40 with phospho-AKT was confirmed by Western blot analysis on a series of glioblastoma tumors, and inhibition of PI3 Kinase signaling by addition of LY294002 also decreased secretion of YKL-40 over a 7-day period in U87 glioblastoma cell tine. Lastly, YKL-40 expression was induced in response to serum starvation and altered by interaction with specific extracellular matrix (ECM) modules. In summary, we have identified the first accurate serum marker for high-grade gliomas. Furthermore, our findings indicate that YKL-40 is a highly expressed vascular-related glycoprotein in human GBM tissue and that it is affected by the AKT signaling pathway and interaction with components of brain ECM proteins. ^

Nd isotope data for planktonic foraminifera from within and around sapropels S1 and S5 from ODP Site 160-967

The development of widespread anoxic conditions in the deep oceans is evidenced by the accumulation and preservation of organic-carbon-rich sediments, but its precise cause remains controversial. The two most popular hypotheses involve (1) circulation-induced increased stratification resulting in reduced oxygenation of deep waters or (2) enhanced productivity in the surface ocean, increasing the raining down of organic matter and overwhelming the oxic remineralization potential of the deep ocean. In the periodic development of deep-water anoxia in the Pliocene-Pleistocene Mediterranean Sea, increased riverine runoff has been implicated both as a source for nutrients that fuel enhanced photic-zone productivity and a source of a less dense freshwater cap leading to reduced circulation, basin-wide stagnation, and deep-water oxygen starvation. Monsoon-driven increases in Nile River discharge and increased regional precipitation due to enhanced westerly activity-two mechanisms that represent fundamentally different climatic driving forces-have both been suggested as causes of the altered freshwater balance. Here we present data that confirm a distinctive neodymium (Nd) isotope signature for the Nile River relative to the Eastern Mediterranean-providing a new tracer of enhanced Nile outflow into the Mediterranean in the past. We further present Nd isotope data for planktonic foraminifera that suggest a clear increase in Nile discharge during the central intense period of two recent anoxic events. Our data also suggest, however, that other regional freshwater sources were more important at the beginning and end of the anoxic events. Taken at face value, the data appear to imply a temporal link between peaks in Nile discharge and enhanced westerly activity.

(Table 2) Production characteristics of phytoplankton and selected related factors at bottle sampling stations in the subtropical and tropical Atlantic Ocean

In October and November 2002, high and relatively high values of chlorophyll a concentration at the sea surface (Cchl) were observed in the English Channel (0.47 mg/m**3), in waters of the North Atlantic Current (0.25 mg/m**3 ), in the tropical and subtropical anticyclonic gyres (0.07-0.42 mg/m**3), and also in the southwestern region of the southern subtropical anticyclonic gyre (usually 0.11-0.23 mg/m**3). The central regions of the southern subtropical anticyclonic gyre (SATG) and the North Atlantic tropical gyre (NATR) were characterized by lower values of Cchl (0.02-0.08 mg/m**3 for the SATG and 0.07-0.14 mg/m**3 for the NATR). At most of the SATG stations, values of surface primary production (Cphs) varied from 2.5 to 5.5 mg C/m**3 per day and were mainly defined by fluctuations of Cchl (r = +0.78) rather than by those of the assimilation number (r = +0.54). Low assimilation activity of phytoplankton in these waters (1.3-4.6 mg chl a per hour) pointed to a lack of nutrients. Analysis of variability of their concentration and composition of photosynthetic pigments showed that, in waters north of 30°N, the growth of phytoplankton was mostly restricted by deficiency of nitrogen, while, in more southern areas, at the majority of stations (about 60%), phosphorus concentrations were minimal. At low concentrations of nitrates and nitrites, ammonium represented itself as a buffer that prevented planktonic algae from extreme degrees of nitric starvation. In tropical waters and in waters of the SATG, primary production throughout the water column varied from 240 to 380 mg C/m**2 30° per day. This level of productivity at stations with low values of C chl (<0.08 mg/m**3) was provided by a well-developed deep chlorophyll maximum and high transparency of water. Light curves of photosynthesis based on in situ measurements point to high efficiency of utilizing penetrating solar radiation by phytoplankton on cloudy days.

Radar profiles from South Shetland Islands/Western Antarctic Peninsula

The sedimentary architecture of polar gravel-beach ridges is presented and it is shown that ridge internal geometries reflect past wave-climate conditions. Ground-penetrating radar (GPR) data obtained along the coasts of Potter Peninsula (King George Island) show that beach ridges unconformably overlie the prograding strand plain. Development of individual ridges is seen to result from multiple storms in periods of increased storm-wave impact on the coast. Strand-plain progradation, by contrast, is the result of swash sedimentation at the beach-face under persistent calm conditions. The sedimentary architecture of beach ridges in sheltered parts of the coast is characterized by seaward-dipping prograding beds, being the result of swash deposition under stormy conditions, or aggrading beds formed by wave overtopping. By contrast, ridges exposed to high-energy waves are composed of seaward- as well as landward-dipping strata, bundled by numerous erosional unconformities. These erosional unconformities are the result of sediment starvation or partial reworking of ridge material during exceptional strong storms. The number of individual ridges which are preserved from a given time interval varies along the coast depending on the morphodynamic setting: sheltered coasts are characterized by numerous small ridges, whereas fewer but larger ridges develop on exposed beaches. The frequency of ridge building ranges from decades in the low-energy settings up to 1600 years under high-energy conditions. Beach ridges in the study area cluster at 9.5, 7.5, 5.5, and below 3.5 m above the present-day storm beach. Based on radiocarbon data, this is interpreted to reflect distinct periods of increased storminess and/or shortened annual sea-ice coverage in the area of the South Shetland Islands for the times around 4.3, c. 3.1, 1.9 ka cal BP, and after 0.65 ka cal BP. Ages further indicate that even ridges at higher elevations can be subject to later reactivation and reworking. A careful investigation of the stratigraphic architecture is therefore essential prior to sampling for dating purposes.

Dioxin-like and non-dioxin like contaminants in the herring Clupea harengus in the Norwegian and North Sea

The Norwegian spring spawning (NSS) herring is an ecologically important fish stock in the Norwegian Sea, and with a catch volume exceeding one million tons a year it is also economically important and a valuable food source. In order to provide a baseline of the levels of contaminants in this fish stock, the levels of organohalogen compounds were determined in 800 individual herring sampled at 29 positions in the Norwegian Sea and off the coast of Norway. Due to seasonal migration, the herring were sampled where they were located during the different seasons. Concentrations of dioxins and dioxin-like PCBs, non-dioxin-like PCBs (PCB7) and PBDEs were determined in fillet samples of individual herring, and found to be relatively low, with means (min-max) of 0.77 (0.24-3.5) ngTEQ/kg wet weight (ww), 5.0 (1.4-24) µg/kg ww and 0.47 (0.091-3.1) µg/kg ww, respectively. The concentrations varied throughout the year due to the feeding- and spawning cycle: Starved, pre-spawning herring caught off the Norwegian coast in January-February had the highest levels and those caught in the Norwegian Sea in April-June, after further starvation and spawning, had the lowest levels. These results show that the concentrations of organohalogen compounds in NSS herring are relatively low and closely tied to their physiological condition, and that in the future regular monitoring of NSS herring should be made in the spawning areas off the Norwegian coast in late winter.

Shell condition and survival of Puget Sound pteropods are impaired by ocean acidification conditions

We tested whether the thecosome pteropod Limacina helicina from Puget Sound, an urbanized estuary in the northwest continental US, experiences shell dissolution and altered mortality rates when exposed to the high CO2, low aragonite saturation state (Omega a) conditions that occur in Puget Sound and the northeast Pacific Ocean. Five, week-long experiments were conducted in which we incubated pteropods collected from Puget Sound in four carbon chemistry conditions: current summer surface (460-500 µatm CO2, Omega a=1.59), current deep water or surface conditions during upwelling (760 and 1600-1700 µatm CO2, Omega a=1.17 and 0.56), and future deep water or surface conditions during upwelling (2800-3400 µatm CO2, Omega a=0.28). We measured shell condition using a scoring regime of five shell characteristics that capture different aspects of shell dissolution. We characterized carbon chemistry conditions in statistical analyses with Omega a, and conducted analyses considering Omega a both as a continuous dataset and as discrete treatments. Shell dissolution increased linearly as aragonite saturation state decreased. Discrete treatment comparisons indicate that shell dissolution was greater in undersaturated treatments compared to oversaturated treatments. Survival increased linearly with aragonite saturation state, though discrete treatment comparisons indicated that survival was similar in all but the lowest saturation state treatment. These results indicate that, under starvation conditions, pteropod survival may not be greatly affected by current and expected near-future aragonite saturation state in the NE Pacific, but shell dissolution may. Given that subsurface waters in Puget Sound's main basin are undersaturated with respect to aragonite in the winter and can be undersaturated in the summer, the condition and persistence of the species in this estuary warrants further study.

Effects of pre-hatching pCO2 on development and survival of zoea stages of Arctic spider crab Hyas araneus

Sensitivity of marine crustaceans to anthropogenic CO2 emissions and the associated acidification of the oceans may be less than that of other, especially lower, invertebrates. However, effects on critical transition phases or carry-over effects between life stages have not comprehensively been explored. Here we report the impact of elevated seawater PCO2 values (3100 µatm) on Hyas araneus during the last 2 weeks of their embryonic development (pre-hatching phase) and during development while in the consecutive zoea I and zoea II larval stages (post-hatching phase). We measured oxygen consumption, dry weight, developmental time and mortality in zoea I to assess changes in performance. Feeding rates and survival under starvation were investigated at different temperatures to detect differences in thermal sensitivities of zoea I and zoea II larvae depending on pre-hatch history. When embryos were pre-exposed to elevated PCO2 during maternal care, mortality increased about 60% under continued CO2 exposure during the zoea I phase. The larvae that moulted into zoea II, displayed a developmental delay by about 20 days compared to larvae exposed to control PCO2 during embryonic and zoeal phases. Elevated PCO2 caused a reduction in zoea I dry weight and feeding rates, while survival of the starved larvae was not affected by the seawater CO2 concentration. In conclusion, CO2 effects on egg masses under maternal care carried over to the first larval stages of crustaceans and reduced their survival and development to levels below those previously reported in studies exclusively focussing on acute PCO2 effects on the larval stages.

Energy and power aware scheduling algorithm modeling for battery powered embedded systems

Energy management has always been recognized as a challenge in mobile systems, especially in modern OS-based mobile systems where multi-functioning are widely supported. Nowadays, it is common for a mobile system user to run multiple applications simultaneously while having a target battery lifetime in mind for a specific application. Traditional OS-level power management (PM) policies make their best effort to save energy under performance constraint, but fail to guarantee a target lifetime, leaving the painful trading off between the total performance of applications and the target lifetime to the user itself. This thesis provides a new way to deal with the problem. It is advocated that a strong energy-aware PM scheme should first guarantee a user-specified battery lifetime to a target application by restricting the average power of those less important applications, and in addition to that, maximize the total performance of applications without harming the lifetime guarantee. As a support, energy, instead of CPU or transmission bandwidth, should be globally managed as the first-class resource by the OS. As the first-stage work of a complete PM scheme, this thesis presents the energy-based fair queuing scheduling, a novel class of energy-aware scheduling algorithms which, in combination with a mechanism of battery discharge rate restricting, systematically manage energy as the first-class resource with the objective of guaranteeing a user-specified battery lifetime for a target application in OS-based mobile systems. Energy-based fair queuing is a cross-application of the traditional fair queuing in the energy management domain. It assigns a power share to each task, and manages energy by proportionally serving energy to tasks according to their assigned power shares. The proportional energy use establishes proportional share of the system power among tasks, which guarantees a minimum power for each task and thus, avoids energy starvation on any task. Energy-based fair queuing treats all tasks equally as one type and supports periodical time-sensitive tasks by allocating each of them a share of system power that is adequate to meet the highest energy demand in all periods. However, an overly conservative power share is usually required to guarantee the meeting of all time constraints. To provide more effective and flexible support for various types of time-sensitive tasks in general purpose operating systems, an extra real-time friendly mechanism is introduced to combine priority-based scheduling into the energy-based fair queuing. Since a method is available to control the maximum time one time-sensitive task can run with priority, the power control and time-constraint meeting can be flexibly traded off. A SystemC-based test-bench is designed to assess the algorithms. Simulation results show the success of the energy-based fair queuing in achieving proportional energy use, time-constraint meeting, and a proper trading off between them. La gestión de energía en los sistema móviles está considerada hoy en día como un reto fundamental, notándose, especialmente, en aquellos terminales que utilizando un sistema operativo implementan múltiples funciones. Es común en los sistemas móviles actuales ejecutar simultaneamente diferentes aplicaciones y tener, para una de ellas, un objetivo de tiempo de uso de la batería. Tradicionalmente, las políticas de gestión de consumo de potencia de los sistemas operativos hacen lo que está en sus manos para ahorrar energía y satisfacer sus requisitos de prestaciones, pero no son capaces de proporcionar un objetivo de tiempo de utilización del sistema, dejando al usuario la difícil tarea de buscar un compromiso entre prestaciones y tiempo de utilización del sistema. Esta tesis, como contribución, proporciona una nueva manera de afrontar el problema. En ella se establece que un esquema de gestión de consumo de energía debería, en primer lugar, garantizar, para una aplicación dada, un tiempo mínimo de utilización de la batería que estuviera especificado por el usuario, restringiendo la potencia media consumida por las aplicaciones que se puedan considerar menos importantes y, en segundo lugar, maximizar las prestaciones globales sin comprometer la garantía de utilización de la batería. Como soporte de lo anterior, la energía, en lugar del tiempo de CPU o el ancho de banda, debería gestionarse globalmente por el sistema operativo como recurso de primera clase. Como primera fase en el desarrollo completo de un esquema de gestión de consumo, esta tesis presenta un algoritmo de planificación de encolado equitativo (fair queueing) basado en el consumo de energía, es decir, una nueva clase de algoritmos de planificación que, en combinación con mecanismos que restrinjan la tasa de descarga de una batería, gestionen de forma sistemática la energía como recurso de primera clase, con el objetivo de garantizar, para una aplicación dada, un tiempo de uso de la batería, definido por el usuario, en sistemas móviles empotrados. El encolado equitativo de energía es una extensión al dominio de la energía del encolado equitativo tradicional. Esta clase de algoritmos asigna una reserva de potencia a cada tarea y gestiona la energía sirviéndola de manera proporcional a su reserva. Este uso proporcional de la energía garantiza que cada tarea reciba una porción de potencia y evita que haya tareas que se vean privadas de recibir energía por otras con un comportamiento más ambicioso. Esta clase de algoritmos trata a todas las tareas por igual y puede planificar tareas periódicas en tiempo real asignando a cada una de ellas una reserva de potencia que es adecuada para proporcionar la mayor de las cantidades de energía demandadas por período. Sin embargo, es posible demostrar que sólo se consigue cumplir con los requisitos impuestos por todos los plazos temporales con reservas de potencia extremadamente conservadoras. En esta tesis, para proporcionar un soporte más flexible y eficiente para diferentes tipos de tareas de tiempo real junto con el resto de tareas, se combina un mecanismo de planificación basado en prioridades con el encolado equitativo basado en energía. En esta clase de algoritmos, gracias al método introducido, que controla el tiempo que se ejecuta con prioridad una tarea de tiempo real, se puede establecer un compromiso entre el cumplimiento de los requisitos de tiempo real y el consumo de potencia. Para evaluar los algoritmos, se ha diseñado en SystemC un banco de pruebas. Los resultados muestran que el algoritmo de encolado equitativo basado en el consumo de energía consigue el balance entre el uso proporcional a la energía reservada y el cumplimiento de los requisitos de tiempo real.

The type II secretion system (Xcp) of Pseudomonas putida is active and involved in the secretion of phosphatases.

The genome of the Gram-negative bacterium Pseudomonas putida harbours a complete set of xcp genes for a type II protein secretion system (T2SS). This study shows that expression of these genes is induced under inorganic phosphate (Pi ) limitation and that the system enables the utilization of various organic phosphate sources. A phosphatase of the PhoX family, previously designated UxpB, was identified, which was produced under low Pi conditions and transported across the cell envelope in an Xcp-dependent manner demonstrating that the xcp genes encode an active T2SS. The signal sequence of UxpB contains a twin-arginine translocation (Tat) motif as well as a lipobox, and both processing by leader peptidase II and Tat dependency were experimentally confirmed. Two different tat gene clusters were detected in the P.?putida genome, of which one, named tat-1, is located adjacent to the uxpB and xcp genes. Both Tat systems appeared to be capable of transporting the UxpB protein. However, expression of the tat-1 genes was strongly induced by low Pi levels, indicating a function of this system in survival during Pi starvation.