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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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Does the pre-mRNA splicing occur in S. cerevisiae co- or post-transcriptionally?
Cihlářová, Zuzana ; Půta, František (advisor) ; Kozáková, Eva (referee)
Until recently, the splicing and transcription were seen as almost independent processes. However, today a lot of studies provide plenty of evidence about their connection, even in the yeast Saccharomyces cerevisiae. The connection of these processes is particularly mediated by C-terminal domain of RNA polymerase II, which is consisted of tandemly repeated heptapeptide sequence - YSPTSPS. Amino acid residues of this heptapeptide sequence are specifically phosphorylated during transcription, which regulates transcription process and also the binding of specific factors. These factors are necessary for processing of the nascent transcript. Modifications of the primary transcript occur especially cotranscriptionally in higher eukaryotes, thus before the transcription is terminated and also before the functional mRNA is released. Opinion on cotranscriptional splicing in S. cerevisiae were significantly changed in the last years. However, nowadays the splicing of pre-mRNA of most genes in S. cerevisiae is seen as cotranscriptional process. RNA polymerase II pauses within the terminal exons and this pausing event provides sufficient time for each spliceosomal component to assemble on the pre-mRNA and also for catalysis of splicing before the transcription termination. Keywords: cotranscriptional...
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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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