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Mer1 and Nam8 in splicing regulation
Marková, Michaela ; Abrhámová, Kateřina (advisor) ; Stejskalová, Eva (referee)
By the mutual cooperation, Mer1 nad Nam8 proteins activate pre-mRNA splicing of four specific genes: AMA1, MER2, MER3 and SPO22, that are required for meiotic recombination and nuclear division. Expression of these genes does not change during the cell cycle, nevertheless, the efficient splicing of their pre-mRNA occures only during meiosis because Mer1 protein, which facilitates their splicing, is expressed only in meiotic cells. All these pre-mRNAs contain nonconsensual 5'splice site (5'ss) which is less recognizable for the spliceosomal subunit U1 snRNP in comparison with the consensual sequence. There is an enhancer area near to 5'ss on the pre-mRNA of this genes that serves as a binding site for Mer1p which makes recruitment of U1 snRNP more efficient. Mer1p cooperate via other proteins with Nam8p. Protein Nam8, a part of U1 sn RNP, is bound on pre-mRNA downstream from 5'ss close to the enhancer area. Mer1p in cooparation with Nam8p facilitates spliceosome assembly on the nonconsensual 5'ss and subsequently pre-mRNA splicing.
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The role of cyclophilins in splicing
Cit, Zdeněk ; Půta, František (advisor) ; Novotný, Ivan (referee)
Splicing is a process essential for the proper functioning of eukaryotic cells. It is a complicated and highly dynamic process, participated by large numbers of proteins which perform diverse functions, either directly within the splicing complex, or sometimes outside of it. Among the proteins, which play an important role in splicing, are cyclophilins. Cyclophilins probably cause conformational changes in the other components of splicing complex. They can also maintain them in the desire conformation thereby they contribute to the dynamics of the spliceosome. This work provides an overview of cyclophilins, which were confirmed to participate on pre-mRNA splicing, and summarizes suggestions of possible functions that these proteins may perform.
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The role of tyrosine phosphorylation in hnRNA splicing
Koudelková, Lenka ; Brábek, Jan (advisor) ; Kozáková, Eva (referee)
Coding sequences of eukaryotic genes are interrupted by long segments of noncoding intronic DNA, which must be spliced after a transcription into a heterogenous nuclear RNA. Due to an increasing pressure on complexity of proteome eukaryotic organisms evolved alternative splicing. It is enabled through weak consensus sequences of splice sites flanked with accessory regulatory RNA elements, that associate with splicing factors, to create protein products according to current requirements implicated by outer and inner conditions. The net of cooperatively or antagonistic acting factors determines whether splice sites are recognized or not. This molecular system is regulated by enzymatic modifications depending on activity of corresponding signaling pathways. Beside many other enzymes a family of protein tyrosine kinases is involved in the process. Via catalytic activity of their kinase domains, they add phosphate to tyrosines of proteins that participate in RNA metabolism. Phosphorylation affects their affinity for RNA and other interacting partners, localization, enzymatic activity or other properties. The changes result in establishing of new setting of regulatory net and usage of distinct splice sites. Products then may with a different efficiency inhibit or trigger various cell processes or...
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Physical interactions of the splicing factor Prp45
Kratochvílová, Eliška ; Folk, Petr (advisor) ; Doubravská, Lenka (referee)
It is well known that chromatin posttranslational state, transcription and splicing influence each other. Nevertheless, the details of this coupling are not fully understood. In S. cerevisiae, it is possible to induce conditions, in which splicing is uncoupled from transcription. Such situation occurred in cells expressing a mutated splicing factor Prp45, whose human homolog has been proved to participate in transcription regulation and also in splicing reactions. Based on previously indicated interactions in high throughput two-hybrid screens, we have been looking for physical links between Prp45 and proteins involved in chromatin posttranslational modifications. Finding of such a link would provide insight into the relationships of gene expression processes. Using coimmunoprecipitation and affinity purification, we were unable to detect physical interactions between Prp45 and our candidate chromatin regulators. Alternative approaches are discussed. Using the precipitation techniques, we mapped the interaction of Prp46 with truncated variants of Prp45. This observation contributes to our knowledge of protein-protein interactions within the spliceosome.
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