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The relationship between splicing and posttranslational modifications of chromatin in Saccharomyces cerevisiae
Kovaľová, Libuša ; Folk, Petr (advisor) ; Čáp, Michal (referee)
Protein Prp45, the yeast ortholog of the human transcription coregulator SNW1/SKIP, has been previously associated only with the regulation of pre-mRNA splicing. However, our laboratory found that protein Prp45 genetically interacts not only with the proteins involved in pre-mRNA splicing, but also with factors important for transcription elongation and with chromatin modifying enzymes. Our data and the information about the human ortholog SNW1/SKIP suggest that Prp45 could serve as a regulator coupling splicing, transcription and chromatin state in S. cerevisiae. The main aim of this diploma thesis was to find out whether the protein Prp45, which is essential for cotranscriptional assembly of the spliceosome, affects posttranslational modifications of chromatin on transcribed genes. Using chromatin immunoprecipitation, the influence of prp45(1-169) mutation on trimethylation of histone H3 at lysine 4 and acetylation of histone H3 at lysines 9, 14 and 18 on transcriptionally active genes was not confirmed. The other aim was to analyse the behavior of cells synchronized by α-factor by using flow cytometry. According to our results, prp45(1-169) mutation leads to the prolongation of the cell cycle. For the purpose of monitoring the dynamics of nucleosomes in S. cerevisiae strains, the system of...
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The relationship between splicing and posttranslational modifications of chromatin in Saccharomyces cerevisiae
Kovaľová, Libuša ; Folk, Petr (advisor) ; Čáp, Michal (referee)
Protein Prp45, the yeast ortholog of the human transcription coregulator SNW1/SKIP, has been previously associated only with the regulation of pre-mRNA splicing. However, our laboratory found that protein Prp45 genetically interacts not only with the proteins involved in pre-mRNA splicing, but also with factors important for transcription elongation and with chromatin modifying enzymes. Our data and the information about the human ortholog SNW1/SKIP suggest that Prp45 could serve as a regulator coupling splicing, transcription and chromatin state in S. cerevisiae. The main aim of this diploma thesis was to find out whether the protein Prp45, which is essential for cotranscriptional assembly of the spliceosome, affects posttranslational modifications of chromatin on transcribed genes. Using chromatin immunoprecipitation, the influence of prp45(1-169) mutation on trimethylation of histone H3 at lysine 4 and acetylation of histone H3 at lysines 9, 14 and 18 on transcriptionally active genes was not confirmed. The other aim was to analyse the behavior of cells synchronized by α-factor by using flow cytometry. According to our results, prp45(1-169) mutation leads to the prolongation of the cell cycle. For the purpose of monitoring the dynamics of nucleosomes in S. cerevisiae strains, the system of...
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Use of antibodies for the characterization of chromatin modifications in Saccharomyces cerevisiae
Kovaľová, Libuša ; Hodek, Petr (advisor) ; Malík, Radek (referee)
Transcription of precursor mRNA (pre-mRNA) and its splicing were originally conceived as two separate processes. Using Saccharomyces cerevisiae as a model, it was shown that the assembly of the complex catalyzing pre-mRNA splicing (spliceosome) can occur cotranscriptionally, i. e., during the time before the termination of transcription by RNA polymerase II. Research on cotranscriptional splicing revealed that proteins involved in transcription and specific chromatin modifications may affect pre-mRNA splicing and its regulation. It is also possible that spliceosome assembly and chromatin modifications can affect each other. Prp45, the yeast ortholog of the human transcription coregulator SKIP/SNW1, has been previously associated only with splicing. The results obtained in our laboratory suggest that Prp45 could be used as a regulator coupling the processes of transcription and splicing. We have shown that PRP45 has genetic interactions with factors important for transcription elongation, as well as chromatin modifications, and that it affects early stage of spliceosome assembly. The aim of this bachelor project was document the relationship between the physiological role of Prp45 and H3K4 trimethylation using chromatin immunoprecipitation. It was found that prp45(1-169) mutation does not markedly...
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