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Transcription factors CSL and their role in the yeast Schizosaccharomyces pombe
Oravcová, Martina
Proteins of the CSL family (CBF1/RBP-Jκ/Suppressor of Hairless/LAG-1) act as effectors of the Notch signalling pathway in metazoan organisms. They function as repressors or activators of gene transcription in the framework of this pathway and influence many developmental processes. Metazoan CSL proteins can regulate gene expression Notch-independently as well. Notch-independent functions of CSL proteins might be evolutionarily ancestral and in cells and organisms may be important equally as Notch-dependent functions. Presence of CSL proteins was identified in several fungal species, organisms lacking the Notch signalling pathway components and most of known metazoan interacting partners of CSL proteins. CSL paralogs of the fission yeast Schizosaccharomyces pombe, cbf11 and cbf12, are non-essential genes encoding proteins localized in the nucleus of the cell. They exert antagonistic effects on regulation of processes like coordination of nuclear and cellular division and cell cycle progression, ploidy maintenance, cell adhesion and other. In this study, we have proved that both CSL paralogs are able to sequence-specifically bind the CSL-response element DNA in vitro and Cbf11 in vivo as well. Both proteins could activate gene expression in vivo and perform the function of transcription factors....
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Study of the interaction between DNA and transcription factors using mass spectrometry.
Slavata, Lukáš ; Man, Petr (advisor) ; Obšil, Tomáš (referee)
Transcription factors play crucial regulatory role within the cell and the entire multicellular organism. The important factor is its ability to interact with other regulatory proteins and DNA. Despite the fact that a large part of the interaction network is already documented, detailed information on the structure and dynamics of protein-protein and protein-DNA complexes is still scarce. In this thesis we focused on the possibility of studying conformational changes given by the transcription factor-DNA complex formation using the methods of structural mass spectrometry: hydrogen/deuterium exchange and chemical crosslinking. As a model, we chose a transcription factor FOXO4 which DNA binding domain is structurally well characterized both in free form and in the complex with DNA.
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Characterization of promoter regions of HGSNAT and GBA genes, and a contribution to the study of pathogenesis of MPS IIIC and Gaucher disease
Richtrová, Eva ; Hřebíček, Martin (advisor) ; Macek, Milan (referee) ; Adam, Tomáš (referee)
Pathogenesis of mucopolysaccharidosis type IIIC (MPS IIIC) and Gaucher disease has not been yet fully clarified, and the causes of phenotypical variability between the patients with the same genotype in Gaucher disease remain obscure. Because the variants in the regulatory regions of genes can cause phenotypical differences mentioned above, I have studied promoter regions of HGSNAT and GBA genes mutated in these lysosomal disorders. I have shown that there is an alternative promoter of GBA (P2). Additional studies were aimed to elucidate possible physiological functions of P2, and its possible role in the pathogenesis of Gaucher disease. I have found that P2 is not tissue specific, and that its variants do not influence the variability of phenotype in Gaucher patients with the same genotype. P2 is used differentially neither during the differentiation of monocytes to macrophages nor in macrophages from controls and Gaucher patients, in whom there is a prominent storage only in cells of macrophage origin. We have thus not found any changes that would suggest a role for P2 in the pathogenesis of Gaucher disease. I have characterized the promoter region of HGSNAT and shown that the binding of Sp1 transcription factor is important for its expression. Sequence variants found in HGSNAT promoter in...
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Regulation of transcription by proteins of the Early growth response and Myb families
Čermák, Vladimír ; Dvořák, Michal (advisor) ; Vomastek, Tomáš (referee) ; Elleder, Daniel (referee)
The regulation of transcription of tens of thousands of genes in a vertebrate organism is an enormously complex phenomenon which entails the participation of thousands of various regulatory proteins. The largest functional category of these regulators is accounted for by sequence-specific DNA-binding proteins known as transcription factors. Proteins of the EGR and Myb families of transcription factors are long-studied regulators of a variety of physiological processes including cellular proliferation and differentiation. The structural and physical aspects of their function have been well characterized. Their cell-type specific participation in complex gene-regulatory networks, on the other hand, is still incompletely understood and represents a major challenge in the respective research areas. Preliminary analysis of gene expression data from metastasizing PR9692 and non- metastasizing PR9692-E9 chicken sarcoma cell lines revealed that the transcription factor EGR1 is expressed at a higher level in metastasizing cells and can thus take part in the regulatory processes that underlie the differences between the two cell lines. Further investigation demonstrated that the introduction of exogenous EGR1 into PR9692-E9 cells restored their metastatic potential to a level indistinguishable from PR9692...
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Study of the mechanism of DNA recognition by transcription factor FOXO4
Zusková, Iva ; Obšil, Tomáš (advisor) ; Bouřa, Evžen (referee)
Forkhead transcription factor FOXO4 is involved in a wide range of processes including metabolism control, cell cycle regulation, apoptosis, DNA repair or oxidative stress resistance. The crystal structure of FOXO4 DNA-binding domain bound to the target DNA revealed that FOXO4 interacts with DNA in the same manner as other forkhead proteins. The DNA-binding interface consists of helix H3, which binds to the major groove of DNA, the N- terminal segment and flexible loops located at the C-terminus of forkhead domain. However, many questions concerning the regulation of DNA-binding specificity and the role of different parts of forkhead domain in this process remain elusive. In this diploma thesis, the interactions between the DNA-binding domain of FOXO4 and DNA with various sequences were studied. The DNA-binding domain of FOXO4 and its mutants were expressed and purified and their DNA-binding properties were studied using fluorescence spectroscopy techniques and surface plasmon resonance. Results of these experiments revealed that the nucleotide sequence at the 5′ end of the consensus binding motif affects the stability of the complex between the DNA-binding domain of FOXO4 and DNA. In addition, the importance of individual amino acid residues of FOXO4 that are involved in DNA binding for the...
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Cis and trans elements in plant response to drought (with a particular focus on photosynthetic genes)
Panchártek, Daniel ; Holá, Dana (advisor) ; Feketová, Zuzana (referee)
Abiotic stresses, such as drought, high salinity and cold can strongly affect plant development and growth. Cis and trans elements are one of the options how plants regulate their own metabolism in those cases. That's the way how the expression of many target genes is induced. The products of these genes function not only in stress tolerance but also in general stress response. Many transcription factors and regulatory proteins (trans elements) are involved in these adaptions; those often interact with specific sequences in gene promoters (cis elements). Recenly, a progress has been made in analyzing the signal paths and complex cascades of gene expression regulation, although a little is still known about this regulation during drought conditions. Some of these target genes code products participating in photosyntesis and the regulation of their expression can significantly affect this process. This essay briefly describes main cis and trans elements of plant response to drought (with a particular focus on the regulation of photosyntetic genes). Keywords: drought, gene expression, cis-element, transcription factor, abcisic acid, photosyntesis, promoter.
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Detection of Transcription Factor Binding Sites
Hlávka, Ondřej ; Vogel, Ivan (referee) ; Martínek, Tomáš (advisor)
Nowadays, it is very important to study gene expression mechanism in molecular biology. Gene expression is also regulated by sequence specific transcription factors which binds to regulatory regions of the genes. Searching for this specific sequences can be very problematic because transcription factor binding sites can be very degenerative. There are several possible methods that can be aplied to this problem. First part of this paper describes few algorithms for transcription binding sites search. Second part contains design and implementation of algorithm for searching binding sites of transcription factor p53.
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Analysis of DNA Structure Elements
Knytl, Marek ; Burgetová, Ivana (referee) ; Martínek, Tomáš (advisor)
The aim of this master's thesis is the design and implementation of tool trackAnalysis for statistical analysis of DNA structure elements. The positions of individual elements in genome are obtained in the form of the track, and with these positions the tool performs a set of analyzes, including randomness test of track, test examining distances between track and genes, detection of clusters and overlaps. The indivudual tests results can be linked together. The results will be displayed in the form of a list, a graph or a new annotation track. An important part of this thesis is also testing the resulting tool on a set of real data.
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