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Preparation of DNA binding domain of transcription factor Fox04 for tryptophan fluorescence measurements
Vácha, Petr
This thesis was worked up as a part of a project that is concerned with the manner of interaction of 14-3-3 proteins with forkhead transcriptional faktor FoxO4. 14-3-3 proteins are proteins interacting with a considerable amount of cell proteins. They affect activity and cell localization through this interaction. Transcriptional factor FoxO4 plays significant role in regulation of a cell cycle, apoptosis, respond to oxidative stress and many other cell proceses. Transcriptional aktivity of FoxO4 is regulated through phophorylation by protein kinases B. This phosphorylation induce inhibition of FoxO4/DNA binding, induce FoxO4/14-3-3 binding and thus relocalization of this complex from a cell core to cytoplasma. One of main goals of this project is to explain molecular mechanisms of these interactions. Main goal of this thesis was to create cDNA of a mutational protein FoxO4 containing just two tryptophan residues by site directed mutagenesis and after gaining this cDNA to express and pyrity this mutational FoxO4. This mutational protein was going to be a part of a model system for conformational changes studies. Besides expression and purification of 14-3-3 protein that is also part of our model system was performed.
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Activity of cytochromes P450 1A1, 1A2 and 3A4 expressed in eukaryotic and prokaryotic systems
Indra, Radek ; Stiborová, Marie (advisor) ; Mizerovská, Jana (referee)
Cytochromes P450 (CYP) are a superfamily of heme proteins distributed widely throughout nature, involved in metabolism of a broad variety of substrates and catalyzing a variety of interesting chemical reactions. They play a central role in metabolism of chemotherapeutic agents. Several prodrug antitumor agents have been found as CYP substrates. Ellipticine, an alkaloid found in Apocynaceae plants, is an example of such type of pro-drug. Here, we investigate the efficiencies of human recombinant CYPs expressed in eukaryotic and prokaryotic expression systems, namely in SupersomesTM , microsomes isolated from insect cells transfected with baculovirus construct containing cDNA of human CYP1A1, 1A2 and 3A4 with NADPH:CYP reductase or in Bactosomes, the membrane fraction of E. coli transfected with cDNA of the same human CYP enzymes and NADPH:CYP reductase to oxidize their marker substrates and ellipticine. Cytochrome b5, an aditional component of the mixed function oxidase system, which metabolize xenobiotics was also expressed in some of the systems. The results found in this work demonstrate that human CYP1A1, 1A2 or 3A4 expressed in both eukaryotic and procaryotic systems oxidize their marker substrates (EROD for CYP1A1/2, MROD for CYP1A2 and testosterone 6β-hydroxylation for CYP3A4). They also oxidize...
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Characterization of mice with constitutively active Wnt/beta-catenin signaling pathway in lens
Antošová, Barbora ; Kozmik, Zbyněk (advisor) ; Pavlínková, Gabriela (referee)
Lens development and differentiation are strictly regulated processes. Various disturbances of these processes can lead to vision-limiting pathologies. The vertebrate lens is composed of epithelial cells and terminally differentiated fiber cells. Differentiation of fiber cells is connected with expression of fiber cell specific proteins (such as crystallins), cell cycle exit, and finally with the degradation of cell nucleus and organelles. Wnt/β-catenin signaling plays important roles during early eye development as well as later during the lens differentiation. To investigate the consequences of constitutive activation of Wnt/β-catenin signaling in lens fiber cells transgenic mouse strain, called CLEF, was created. Constitutive activation of Wnt/β-catenin signaling in fiber cells of CLEF mouse is achieved by transgenic protein CLEF that contains C-terminal activation domain of β-catenin fused to the amino terminus of full-length protein Lef1. The expression of CLEF transgene is under the control of αA-crystallin promoter. As a result of constitutive activation of Wnt/β-catenin signaling in fiber cells, adult CLEF mice develop cataracts and microphthalmia, and the morphology of adult mutant lenses is disrupted. Transgenic CLEF mRNA is expressed starting from E13.5 and by E16.5 transgenic CLEF protein is...
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Design and processing of teaching practices dash navigation
Bandúr, Juraj ; Vosecký, Slavomír (referee) ; Chlebek, Jiří (advisor)
The diploma thesis deals with concepts of various key tasks for flights operated by device navigation, while these tasks are designed under the requirements of the regulation JAR-FCL 1. The work also includes explanation of the principles of operation of selected radio navigation devices, which are demonstrated in various roles, making these tasks serve well as a possible teaching material for navigation subjects. Part of the work also includes the evaluation of the simulator FlitePro for the purposes of its certification as a training device.
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Essay on Financial Innovation, Credit Constraints, and Welfare
Janíčko, Martin ; Chytil, Zdeněk (advisor) ; Tlustý, Adolf (referee) ; Pavelka, Tomáš (referee) ; Frait, Jan (referee)
The submitted thesis is composed of three different articles dealing with issues of financial innovation, credit constraints, and their impact on welfare. The first article treats the contemporary theoretical grasp of the interaction between the financial and real economies, focusing primarily on the role of modern financial innovation in the business cycle. For this purpose, a framework promoted by the Regulation School and Post Keynesians is frequently employed, whilst some other unorthodox streams and mainstream economics are partially discussed as well. All of them aspire -- either per se or under the pressure of the contemporary economic agenda -- to clarify the evolution of financial innovation and credit in the recent era. It is generally found that certain consensus across the schools of economic thought exists, but some of them have done a better job in predicting the consequences of the financial innovation for real economic activity than others. Further, two dynamic macroeconomic models are developed in order to, inter alia, identify the possible effects of extended credit availability presented in the former article on the example of the housing market, and simulate the effects of housing price changes on general welfare. Clearly, this part of the thesis exhibits the indirect consequences of financial innovation as, once again, being rather ambiguous: after having partially unleashed the unprecedented credit granting in the economy, impacting interest rates and loan-to-value ratios, with a subsequent impact on housing prices, it has also influenced credit constrained and unconstrained households in a different manner. Based on an analysis of the situation using partial and general equilibrium analytical frameworks, two somewhat different conclusions are drawn up with respect to the occurrence of various shocks in the models. Under the partial equilibrium framework the effects of relaxation of credit constraints are visible and quite straightforward, indicating relatively simple and intuitive relationship between the price appreciation and general welfare. This is primarily perspicuous for the credit constrained households. In the general equilibrium framework, on the other hand, the transitional dynamics of shock proliferation is more transparent and the impact on credit constrained vs. unconstrained households is more ambiguous and much different from the basic intuition used in the article anchored in the partial equilibrium toolbox.
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Analysis of specific instruments applied in the financial management of TNC with a focus on transfer pricing
Baluchová, Daniela ; Taušer, Josef (advisor) ; Sato, Alexej (referee) ; Baláž, Peter (referee)
Transfer pricing plays a crucial role in the financial management of TNC as it significantly influences revenues and costs allocation among affiliates of TNC that are located in countries with different tax regimes, interest rates, political situation and economic environment. Recently, transfer pricing is scrutinized in respect of tax minimisation strategies adopted by TNC which raises various conflicts of interest with tax authorities in some countries of their presence. The main objective of the dissertation thesis is to provide a comprehensive empirical study on international transfer pricing in the Czech Republic from the perspectives of both taxpayer and the tax authority. With regard to the complexity of transfer pricing, manufacturing afiliates of TNC located in the Czech Republic were selected to be examined in more detail. The thesis is structured into five chapters. The first chapter defines theoretical framework based on which the analytical part of the thesis is elaborated. The strategies applied by TNC in setting transfer prices are strongly affected by the transfer pricing regulation and interpretative experience of the particular countries in which they operate. In this context, the Czech transfer pricing legislation as well as selected case law is analyzed in the second chapter. Given the complexity of the issue of transfer pricing, the third chapter deals with selected aspects that are considered critical when setting transfer prices. The fourth chapter presents key findings regarding transfer pricing strategies applied by TNC located in the Czech Republic to transfer pricing issues. The fifth chapter summarizes the approach of the Czech tax authorities to the transfer pricing audits and at the same time evaluates related risks borne for taxpayers in this respect. The thesis reveals that manufacturing afiliates of TNC located in the Czech Republic generally prefer non market (cost based) transfer pricing methods when setting transfer prices, whereby there are several factors influencing their decision making, out of which internal economic conditions and foreign exchange risk management are deemed the most important factors, while tax optimisation as well as restrictions on profit repatriation are considered relatively less important factors. The study further indicates certain inconsistency between declared functional and risk profiles and decision making competences. In this connection, it was found out that the Czech afiliates in which the foreign parent company is involved in transfer pricing set-up incur tax losses. In response to the increasing importance of transfer pricing and international initiative Action Plan BEPS (Base Erosion and Profit Shifting) it can be stated that the approach of the Czech tax authorities has become more intensified and sophisticated. The Czech tax authorities challenge declared and actual functional and risk profile of taxpayers as well as economic substance of realized intercompany transactions. Furthermore, number of transfer pricing audits has increased and become targeted on risky taxpayers such as companies granting investment incentives or incurring tax losses etc. As a result, additional tax assessment due to incorrect transfer pricing significantly increases over recent years.
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Regulation of financial markets in international context
Nistorenco, Taisia ; Žamberský, Pavel (advisor) ; Jiránková, Martina (referee)
This diploma thesis comprises a theoretical, descriptive, comparative and econometric analysis of financial markets regulation in international context. In the first chapter I describe and analyse characteristics of modern financial markets, reasons for their regulation and institutional models of financial regulation and supervision. The second chapter offers an overview of the historical development of financial regulation and evaluates the role of regulatory factor in the outburst of the world financial crisis of 2008. In the third chapter I carry out a comparison of regulatory response to financial crisis in the USA and the EU. Forth chapter deals with regression analysis of the relation between Financial Soundness Indicators of three selected countries. The conclusions driven from this diploma thesis demonstrate that regulation is generally effective as a remedy for market failures, but in other aspects its effects are ambiguous. Intensity of regulation is a secondary impact factor in the formation of financial crises, in fact it is more reasonable to state that crises occur because of the failure of supervision rather than regulation. Due to the international attempts to harmonise the process of remediation of the consequences of the financial crisis, the regulatory response in the USA and the EU was very similar. Statistical analysis did not confirm the common idea that regulation represents a significant barrier for increasing the profitability of credit institutions.
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Commission and financial market
Macura, Marek ; Smrčka, Luboš (advisor) ; Zámečník, Petr (referee)
This thesis deals with the financial market intermediation and distribution on the financial market. By using situational analysis reveals connections and principles of the functioning of financial intermediation and focuses on the role of financial market regulation and commission fees. Thesis is analyzing the effects of changes in commission system and impact to different distribution models.
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The influence of estrogens on mouse sperm capacitation and acrosome reaction in vitro.
Tejnická, Magda ; Komrsková, Kateřina (advisor) ; Maňásková, Pavla (referee)
- 7 - Abstract There are an increasing amount of compounds in the environment that can have a negative effect on reproductive parameters in both male and female organism. There has been a worldwide decline of sperm quality during past decades and this fact lead to an increase of unnatural ways of conception through assisted reproduction techniques in the specialised centres. Natural estrogens are one of these compounds and they get into waste water after being excluded from the body by the urine. They get back into the human body from drinking water or from the food, and they can interfere with function of endogenous hormones in very low concentrations. For these reasons it is up to date to deal with the influence of these compounds on mammalian sperm. For many years, estrogens have been considered typically female sex hormones. It is now certain that they are also very important in the regulation of male reproduction. Endogenous estrogens in mammalian males are an important part of the endocrine system. Estrogens play an important role in the development of germ cells, spermatogenesis and processes leading to successful egg fertilization such as a capacitation or acrosomal reaction. Tyrosine phosphorylation is one of the essential steps for the properly ongoing process of capacitation in sperm followed by...
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Assessment of expression markers of HLA class II genes.
Zajacová, Marta ; Černá, Marie (advisor) ; Černý, Jan (referee)
BACKGROUND: HLA molecules play a central role in the immune response. HLA class II are involved in the selection of the T-cell repertoire in the thymus, and in presentation of antigenic peptides to antigen reactive CD4-positive T cells. The HLA class II act as restriction determinants in the presentation of antigens to T lymphocytes and their expression on the cell surface is necessary for triggering the immune responses. Regulated transcription of HLA class II genes is a complex system involving cis-acting sequence elements and trans-acting protein factors. It has been reported that allelic polymorphism exists in the regulatory regions of HLA class II DQA1, DQB1 and DRB1 genes. Most of the polymorphisms appear to be conserved within a haplotype. The hierarchy of sequence homology which exists among the structural genes is not paralleled among their promoter sequences. It is of interest that the most striking discrepancy was found for the DR4 linked alleles, DQB1*0301 and DQB1*0302, which were described as frequent risk factors for a variety of autoimmune diseases. The structural genes of these two alleles are the most closely related, but their regulatory sequences are the most heterogeneous among DQB1 variants. These sequence differences correspond to functional variation: The promoter strength...
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