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Determinants of the splice site selection in protein-coding and long non-coding RNAs
Krchňáková, Zuzana ; Staněk, David (advisor) ; Svoboda, Petr (referee) ; Blažek, Dalibor (referee)
In my thesis, I focused on several underexplored areas of RNA splicing regulation. In the first part, I analyzed how chromatin and transcription regulatory elements change pre-mRNA splicing. In the second part, I studied why long non-coding RNAs (lncRNAs) are spliced less efficiently than protein-coding mRNAs. Finally, I was testing the importance of intron for the activating function of lncRNAs. It has been shown that chromatin and promoter identity modulate alternative splicing decisions. Here, I tested whether local chromatin and distant genomic elements that influence transcription can also modulate splicing. Using the chromatin modifying enzymes directly targeted to FOSL1 gene by TALE technology, I showed that changes in histone H3K9 methylation affect constitutive splicing. Furthermore, I provide evidence that deletion of transcription enhancer located several kilobases upstream of an alternative exons changes splicing pattern of the alternative exon. Many nascent lncRNAs undergo the same maturation steps as pre-mRNAs of protein- coding genes (PCGs), but they are often poorly spliced. To identify the underlying mechanisms for this phenomenon, we searched for putative splicing inhibitory sequences. Genome-wide analysis of intergenic lncRNAs (lincRNAs) revealed that, in general, they do not...
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Determinants of the splice site selection in protein-coding and long non-coding RNAs
Krchňáková, Zuzana ; Staněk, David (advisor) ; Svoboda, Petr (referee) ; Blažek, Dalibor (referee)
In my thesis, I focused on several underexplored areas of RNA splicing regulation. In the first part, I analyzed how chromatin and transcription regulatory elements change pre-mRNA splicing. In the second part, I studied why long non-coding RNAs (lncRNAs) are spliced less efficiently than protein-coding mRNAs. Finally, I was testing the importance of intron for the activating function of lncRNAs. It has been shown that chromatin and promoter identity modulate alternative splicing decisions. Here, I tested whether local chromatin and distant genomic elements that influence transcription can also modulate splicing. Using the chromatin modifying enzymes directly targeted to FOSL1 gene by TALE technology, I showed that changes in histone H3K9 methylation affect constitutive splicing. Furthermore, I provide evidence that deletion of transcription enhancer located several kilobases upstream of an alternative exons changes splicing pattern of the alternative exon. Many nascent lncRNAs undergo the same maturation steps as pre-mRNAs of protein- coding genes (PCGs), but they are often poorly spliced. To identify the underlying mechanisms for this phenomenon, we searched for putative splicing inhibitory sequences. Genome-wide analysis of intergenic lncRNAs (lincRNAs) revealed that, in general, they do not...
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Role of the ubiquitin-like protein, Hub1, in the pre-mRNA splicing regulation
Hubáčková, Tereza ; Abrhámová, Kateřina (advisor) ; Krchňáková, Zuzana (referee)
Splicing is a key step of eukaryotic gene expression and as well as other steps of this vital process, splicing has to be tightly regulated. Hub1 protein is a ubiquitin-like protein which noncovalently interacts with spliceosomal proteins Snu66 and Prp5 ATPase. According to the proposed model, low level of Hub1 protein stimulates ATPase activity of Prp5 helicase sufficiently for splicing of optimal splice sites, but not for splicing of suboptimal ones. Nevertheless, high level of Hub1 protein stimulates Prp5 ATPase sufficiently for splicing of both splice-site types. Excessive level of Hub1 protein may be harmful for the cell, because the immoderate splicing of suboptimal splice sites may produce aberrantly-spliced transcripts as a by-product. Hub1-induced negative feedback loop safeguards the cell from Hub1 protein hyperactivity by regulation of Prp5 ATPase level. Additionally, Hub1 protein regulates alternative splicing of Saccharomyces cerevisiae SRC1 gene and ensures appropriate balance of its products.
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