Növényi elektrofiziológiai mérések alkalmazása a NO és az alacsony extracelluláris kálium tartalom sztómaműködésben betöltött szerepének feltárásában = Application of electrophysiological recordings in plant cells in order to reveal the role of NO and low extracellular potassium levels in stomatal function

Ifjúsági OTKA pályázatom kutatási célja a növényi zárósejtek működésének és szabályozásának mélyebb megértése volt. Arra kerestem a választ, hogy a zárósejtek befelé egyenirányító kálium csatornájának (KAT1) aktivitását a drasztikusan csökkenő extracelluláris kálium tartalom hogyan befolyásolja. A német kutatópartnerrel együttműködve KAT1-pEGFP-N2 plazmidot HEK293 heterológ expressziós rendszerbe transzfektáltunk, és a csatornaáramot whole-cell üzemmódban detektáltuk. Megállapítottuk, hogy a KAT1 áram redukciója nem a kapuzás megváltozásával, hanem a működő csatornák számának csökkenésével áll összefüggésben. A folyamat élettani fontossága a szubmillimoláris külső kálium koncentráció esetén a kálium kiszivárgás megakadályozásában rejlik. A kutatásom második felében a nitrogén monoxid (NO) hatását vizsgáltam a zárósejtek membrántranszportja és fotoszintézise tekintetében. Megállapítottam, hogy a homotetramer KAT1 csatorna közvetlenül, még a magas (100 μM) NO koncentráció esetén sem inaktiválódik. A NO közvetített Ca2+-függő szabályozási út a KAT1 homotetramer esetén nem érvényesül. Intakt borsó levelek fotoszintézisének vizsgálatával megállapítottam, hogy a NO a primér töltésszeparációt a QA és QB kötőhely közötti vashoz kapcsolódva in vivo lassítja, gátolja a QA- és a vízbontó komplex S2 állapota közötti töltésrekombinációt. Sejtszintű klorofill fluoreszcencia indukció mérésével zárósejtekben is igazoltam a NO lineáris elektrontranszportlánc gátló hatását. | The scientific goal of my junior OTKA tender was to further understand the function and regulation of guard cells. I examined how the activity of a potassium inward rectifier channel of guard cells (KAT1) is influenced by the drastically decreasing extracellular potassium content. Co-operating with the German research partner, KAT1-pEGFP-N2 plasmids were transfected into HEK293 cells, and the channel currents were recorded in the whole-cell mode. Our results indicate that the reduction of KAT1 current is not related to the gating mechanism, but to the reduction of the number of functional channels. The physiological role of the process in case of submillimolar external potassium resides in the arrest of potassium leaking. In the second part of my research, the influence of nitric oxide (NO) on guard cell membrane transport and photosynthesis was tested. I found that the homotetramer KAT1 channel does not inactivate even at high (100 μM) NO concentrations. The NO mediated Ca2+-dependent pathway does not prevail in the case of KAT1 homotetramer. Investigation of the photosynthesis of intact pea leaves showed that NO slows down the charge separation in vivo by binding to the iron between QA and QB binding site, and impedes charge recombination of QA- with the S2 state of the water-oxidizing complex. Inhibiting effect of NO on electron transport chain was also confirmed in guard cells by the measurement of chlorophyll fluorescence induction at cellular level

pH-dependent regulation of a potassium channel protein encoded by a Chlorella virus PBCV-1

In our project we studied a K+ channel, Kcv, encoded by a large Chlorella virus, PBCV-1. We investigated the pH dependency of the channel. The data reveal a weak dependency of the channel conductance on extracellular and intracellular pH

Inactivation of KAT1 channel of guard cells at submillimolar concentrations of external potassium

In the present study we investigated the effect of depleting K+o on the activity of a plant K+ channel, the inward rectifier KAT1 from Arabidopsis thaliana. Most voltage sensitive K+ channels are able to sense the concentration of extracellular K+ (K+o) and inactivate if K+o decreases to sublillimolar values. The channel was expressed in mammalian HEK293 cells and measured with patch clamp in the whole cell configuration. The effect of K+o depletion on channel activity was monitored from the tail currents before during and after washing K+o from the medium. The data show that a depletion of K+o results in a decrease in channel conductance. This high sensitivity of the channel to K+o could serve as a safety mechanism, which inactivates the channel at low K+o and prevents in this way leakage of K+ from the cells via this type of channel

Deltamethrin rescues run down of K+ outward rectifying channels in Vicia faba guard cells

The dominant outward rectifier K+ currents were examined in protoplasts from Vicia faba guard cells. In whole-cell patch-clamp recordings, we generally observed that the conductance of the K+ inward and the outward rectifier gradually decreases. As a consequence of this run down a new steady state was achieved which was 90±5% lower than that obtained at the beginning of the recording. The run down of the outward rectifier could be greatly reduced by pretreating protoplasts either with the membrane permeable drug deltamethrin or by perfusing protoplasts with a pipette solution containing 5pM cydosporine A. Furthermore, after the rundown the conductance of the outward rectifier could be partially restored upon addition of 5pM deltamethrin to the bath medium. Since deltamethrin and cydosporine A are established inhibitors of the calcium sensitive phosphatas e calcineurin, the data argue for a participation of this type of phosphatase in the control of the activity of K+ outward rectifier channel in guard cells

Code coverage measurement framework for android devices

Software testing is a very important activity in the software development life cycle. Numerous general black- and white-box techniques exist to achieve different goals and there are a lot of practices for different kinds of software. The testing of embedded systems, however, raises some very special constraints and requirements in software testing. Special solutions exist in this field, but there is no general testing methodology for embedded systems. One of the goals of the CIRENE project was to fill this gap and define a general testing methodology for embedded systems that could be specialized to different environments. The project included a pilot implementation of this methodology in a specific environment: an Android-based Digital TV receiver (Set-Top-Box). In this pilot, we implemented method level code coverage measurement of Android applications. This was done by instrumenting the applications and creating a framework for the Android device that collected basic information from the instrumented applications and communicated it through the network towards a server where the data was finally processed. The resulting code coverage information was used for many purposes according to the methodology: test case selection and prioritization, traceability computation, dead code detection, etc. The resulting methodology and toolset were reused in another project where we investigated whether the coverage information can be used to determine locations to be instrumented in order to collect relevant information about software usability. In this paper, we introduce the pilot implementation and, as a proof-of-concept, present how the coverage results were used for different purposes

Essays on robust asset pricing

The central concept of this doctoral dissertation is robustness. I analyze how model and parameter uncertainty affect financial decisions of investors and fund managers, and what their equilibrium consequences are. Chapter 1 gives an overview of the most important concepts and methodologies used in the robust asset allocation and robust asset pricing literature, and it also reviews the most recent advances thereof. Chapter 2 provides a resolution to the bond premium puzzle by featuring robust investors, and – as a technical contribution – it develops a novel technique to solve robust dynamic asset allocation problems: the robust version of the martingale method. Chapter 3 contributes to the resolution of the liquidity premium puzzle by demonstrating that parameter uncertainty generates an additional, positive liquidity premium component, the liquidity uncertainty premium. Chapter 4 examines the effects of model uncertainty on optimal Asset Liability Management decisions

Code Coverage Measurement and Fault Localization Approaches

Code coverage measurement plays an important role in white-box testing, both in industrial practice and academic research. Several areas are highly dependent on code coverage as well, including test case generation, test prioritization, fault localization, and others. Out of these areas, this dissertation focuses on two main topics, and the thesis points are divided into two parts accordingly. The first part consists of one thesis point that discusses the differences between methods for measuring code coverage in Java and the effects of these differences. The second part focuses on a fault localization technique called spectrum-based fault localization that utilizes code coverage to estimate the risk of each program element being faulty. More specifically, the corresponding two thesis points are discussing the improvement of the efficiency of spectrum-based approaches by incorporating external information, e.g., users’ knowledge, and context data extracted from call chains

Structural information aided automated test method for magic 4GL

Nowadays testing data intensive, GUI enhanced applications properly on an easily maintainable way has become a more crucial part of the application life cycle. There are many evolving technologies to support automatized GUI testing in various environments. However there are hardly any methods that support 4GLs, especially Magic. Fortunately, the characteristics of the 4th generation of programming languages - like explicit definition of the relations between GUI elements and data - eliminate most of the problems raised during testing the GUI in the case of 3GLs. By utilizing these advantages we were able to develop a generalized testing method that supports 4GLs, and as a proof of concept a system for testing Magic xpa applications was built. In this paper, a generalized testing method for 4GLs, our path and script generator algorithms, and their implementations for Magic xpa applications are presented. In addition, the cooperation of these components with existing solutions is demonstrated, and a test method that has been competed by the application of our tools (and which is an instantiation of the generalized method) is introduced as a possible use of the results

Effect of pyrethroid insecticides on the photosynthetic activity of pea mesophyll protoplasts

Pyrethroids are plant protecting substances whose effects on treated plants are widely monitored with often conflicting results. Field experiments showed no alteration in photosynthetic rates by a synthetic pyrethroid permethrin (Haile et al. 1999), while other studies showed an inhibition in PSII reactions by the halogen side of the molecules, and suggested QA as a target site of interaction, like in the case of urea-type herbicides (Bader and Schuler 1996). In order to clarify these opposing observations, photosynthetic effects of pyrethroids were re-tested

The effect of exogenous NO on PSI photochemistry in intact pea leaves

We investigated the effect of exogenous NO on PSI photochemistry by measuring P700 absorbance changes at 810-870 nm in intact pea leaves treated with the NO donor GSNO. Our results indicate that NO increases PSI quantum efficiency and the pool size of electrons in the intersystem chain