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Local structures in retrovirus genomes
Sádlová, Adéla ; Kollerová, Silvia (referee) ; Brázda, Václav (advisor)
This bachelor thesis focuses on the study of local structures in the genomes of retroviruses. Retroviruses are a group of viruses with an RNA genome that have the ability to integrate their genome into the host DNA. Local structures in the retroviral genome play an important role in the processes of replication, gene expression regulation, and interactions with host proteins. The aim of this work was to analyze the local structures in selected retroviruses using bioinformatics methods, with G4 Hunter being the key tool for identifying G quadruplexes in the genome. G-quadruplexes are DNA or RNA structures composed of guanine bases arranged into quartets and are connected by stacking guanine tetrads. G quadruplexes are often located in telomeres, gene promoter regions, and can play an important role in regulating gene expression, DNA replication, and other cellular processes. The analysis focused primarily on the presence of potential G-quadruplex-forming sequences (PQS). The results of this work provide important information on the distribution and frequency of PQS in the genomes of selected retroviruses and contribute to a better understanding of their role in viral biology. The results show that G-quadruplexes are significant local structures and are often found in regulatory areas of the genome, indicating their key role in the viral infection process.
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Finding guanine quadruplexes in DNA using decision trees
Kotrys, Kryštof ; Šťastný, Jiří (referee) ; Kaura, Patrik (advisor)
This bachelor’s thesis is focused on creating a functional decision tree model for detection of guanine quadruplexes in the DNA. The first part of the thesis summarizes theoretical knowledge in the fields of local DNA structures, computational prediction methods of guanine quadruplexes and decision trees. The second part describes the creation of a decision tree model, followed by a statistical comparison of the results with the G4Hunter algorithm.
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Sequences forming G-quadruplexes in the amyloid beta precursor human gene and its homologues
Stránská, Anna ; Šedrlová, Zuzana (referee) ; Brázda, Václav (advisor)
The APP gene encodes the transmembrane protein amyloid beta precursor, which is expressed in many cell types, including neurons. Its functions have not yet been fully described, but it is clear that it is being cleaved before being exported to the extracellular space. It is degraded by various degradation pathways also undergoes homodimerization, which can produce particles with protective neuronal function as well as fragments that are toxic and cause nerve cell death. The formation of harmful amyloid beta plaques and their accumulation among neurons in the brain is closely linked to the onset and progression of Alzheimer's disease, a neurodegenerative brain disease manifested by death and loss of neurons, which leads to dementia, i.e. loss of cognitive functions. There is currently a lot of research that deals with the links between neurodegenerative diseases and the occurrence of G-quadruplexes in genes that are involved in disease manifestations. G-quadruplexes are non-canonical DNA and RNA nucleic acid secondary structures that arise in guanine-rich regions. They are important mainly in terms of their connection with biological processes such as the regulation of gene expression in genes and mainly in oncogenes because they occur in important regions of the gene such as the promoter. It is possible to stabilize them with small molecules, and it is this ability that is used in research into the therapeutic treatment of various diseases. A bioinformatics analysis of both the human gene and 346 other gene homologs was performed to determine the importance of G-quadruplexes localization and conservation in the human APP gene. For this purpose, the G4Hunter program was used, which provided information about the found sequences with the potential to form a G-quadruplex, such as their location or G4Hunter score. In vitro analysis was performed using thioflavin T reagent, which tested the ability of the found sequences to form G-quadruplexes under physiological conditions. The results confirmed the presence and evolutionary importance of G-quadruplexes found in the APP gene of Homo sapiens and their ability to assemble into quadruplex structures in the presence of salts such as sodium and potassium.
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Study on the relation between G-quadruplexes and p53-driven regulation
Holotová, Paulína ; Vodička, Juraj (referee) ; Brázda, Václav (advisor)
Táto práca sa zaoberá úlohou štruktúr G-kvadruplexov, ich stabilizáciou pomocou ligandov a úlohou p53 v regulácii transkripcie. G-kvadruplex je typ sekundárnej štruktúry nukleovej kyseliny zloženej z 2 – 4 tetrád. Každá tetráda je tvorená štyrmi guanínmi spojenými prostredníctvom Hoogstenovho párovania báz. Táto práca pozostáva z teoretickej a experimentálnej časti. V teoretickej časti bola opísaná podstata G-kvadruplexových štruktúr a ich potenciál pri liečbe rakoviny, úloha p53 v bunkovom cykle a jeho regulácia a ligandy kurkumín a TMPyP4 použité na stabilizáciu G4. V experimentálnej časti sa študovali interakcie ligandov kurkumínu a TMPyP4 so štruktúrami nukleových kyselín. Reportérové kmene kvasiniek obsahujúce PUMA, KSHV aich kombinácie boli transformované plazmidmi kódujúcimi iba selekčné markery a plazmidmi kódujúcimi divokú formu génu pre proteín p53. V teste životaschopnosti, optimálna koncentrácia ligandov (kurkumínu a TMPyP4) bola stanovená pre reportérový kmeň PUMA, ktorá sa ďalej použila v Luciferázovom teste. Test vytesnenia fluorescenčného indikátora s tioflavínom T ako fluorescenčným farbivom dokázal interakciu medzi oligonukleotidmi tvoriacimi G4 a kurkumínom a TMPyP4. Luciferázový test sa použil na vyhodnotenie interakcie medzi transformovanými reportérovými kmeňmi kvasiniek a oboma ligandmi. Výsledky po 24 hodinách ukázali štatistickú významnosť pre každý kmeň v rovnakom prostredí, pričom kmeň PUMA vykazoval najvyššiu transaktivačnú aktivitu - insert PUMA je cieľovým génom pre p53. Kmene s PUMA aj KSHV vykazovali významne nižšiu transaktivačnú aktivitu, keďže prítomné ligandy mohli stabilizovať štruktúru G4 prítomnú v KSHV, a tým znížiť transkripciu. Prítomnosť G4 v sekvencii KSHV sa potvrdila aj pomocou programu G4Hunter, pričom G4Skóre bolo 3,182. Transaktivácia bola výrazne nižšia aj v kmeňoch PUMA prítomných v prostredí kurkumínu a TMPyP4, aj napriek tomu, že táto sekvencia netvorí G4, čo potvrdil aj program G4Hunter. To naznačuje, že študované ligandy kurkumín a TMPyP4 môžu regulovať metabolizmus buniek iným spôsobom, ako sa doteraz predpokladalo.
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The presence and localization of local DNA structures in the genome of Schizosaccharomyces pombe
Kubínová, Michaela ; Šedrlová, Zuzana (referee) ; Brázda, Václav (advisor)
The thesis focuses on the study of local DNA structures (cruciforms and G quadruplexforming sequence) in the genome of Schizosaccharomyces pombe, a yeast used in the food industry. The analysed local structures are non-randomly distributed within the genome. Based on previous studies, it has been found that they often colocalize with regulatory regions of genes and that the role of these secondary structures in the regulation of basic cellular processes (e.g. replication or transcription) is significant. This analysis was performed using specialized bioinformatics tools (G4Hunter and Palindrome Analyser) that allowed me to identify and analyze these structures in terms of their presence and localization. Many times less IR was found in mtDNA compared to the occurrence of IR in chromosomes. The number and frequency of PQS in mtDNA was also found to be very low. It is very different from the yeast Saccharomyces cerevisiae in this respect. It was also found that the number of IRs found decreases with increasing IR length and about 17% of IRs do not have a loop. A large enrichment of IRs was observed in the repeat_region and rRNA, and in the case of PQS in the rRNA and mRNA regions, i.e. sequences important for cellular processes.
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Sequences forming G-quadruplexes in the amyloid beta precursor human gene and its homologues
Stránská, Anna ; Šedrlová, Zuzana (referee) ; Brázda, Václav (advisor)
The APP gene encodes the transmembrane protein amyloid beta precursor, which is expressed in many cell types, including neurons. Its functions have not yet been fully described, but it is clear that it is being cleaved before being exported to the extracellular space. It is degraded by various degradation pathways also undergoes homodimerization, which can produce particles with protective neuronal function as well as fragments that are toxic and cause nerve cell death. The formation of harmful amyloid beta plaques and their accumulation among neurons in the brain is closely linked to the onset and progression of Alzheimer's disease, a neurodegenerative brain disease manifested by death and loss of neurons, which leads to dementia, i.e. loss of cognitive functions. There is currently a lot of research that deals with the links between neurodegenerative diseases and the occurrence of G-quadruplexes in genes that are involved in disease manifestations. G-quadruplexes are non-canonical DNA and RNA nucleic acid secondary structures that arise in guanine-rich regions. They are important mainly in terms of their connection with biological processes such as the regulation of gene expression in genes and mainly in oncogenes because they occur in important regions of the gene such as the promoter. It is possible to stabilize them with small molecules, and it is this ability that is used in research into the therapeutic treatment of various diseases. A bioinformatics analysis of both the human gene and 346 other gene homologs was performed to determine the importance of G-quadruplexes localization and conservation in the human APP gene. For this purpose, the G4Hunter program was used, which provided information about the found sequences with the potential to form a G-quadruplex, such as their location or G4Hunter score. In vitro analysis was performed using thioflavin T reagent, which tested the ability of the found sequences to form G-quadruplexes under physiological conditions. The results confirmed the presence and evolutionary importance of G-quadruplexes found in the APP gene of Homo sapiens and their ability to assemble into quadruplex structures in the presence of salts such as sodium and potassium.
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Finding guanine quadruplexes in DNA using decision trees
Kotrys, Kryštof ; Šťastný, Jiří (referee) ; Kaura, Patrik (advisor)
This bachelor’s thesis is focused on creating a functional decision tree model for detection of guanine quadruplexes in the DNA. The first part of the thesis summarizes theoretical knowledge in the fields of local DNA structures, computational prediction methods of guanine quadruplexes and decision trees. The second part describes the creation of a decision tree model, followed by a statistical comparison of the results with the G4Hunter algorithm.
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