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Splicing Factors in the Regulation of Gene Expression - the Relationship Between Splicing and Transcription in Saccharomyces cerevisiae
Hálová, Martina
Transcription and pre-mRNA processing, e.g., splicing, occur at the same place and time in the context of chromatin. A growing amount of evidence supports the hypothesis that these processes are interconnected. Prp45/SKIP is one of the factors which are believed to mediate the interconnection. The human ortholog, SKIP, is known for affecting mRNA formation on the levels of transcription initiation and elongation. Moreover, it interacts with chromatin modifiers and it is a splicing factor, too. The function of the Saccharomyces cerevisiae ortholog, Prp45, has been so far connected only to pre-mRNA splicing. In this work, we characterized the role of Prp45 in splicing and elaborated the results connecting Prp45 to transcription and chromatin modifications. RNA-seq results showed that pre-mRNA is accumulated in prp45(1-169) cells. This accumulation is not caused by the reduced activity of pathways responsible for RNA degradation. The extent of the splicing inefficiency in prp45(1-169) cells did not depend on either the canonicity of the 5' splice site and branch site or the distance between the branch site and the 3' splice site. Using chromatin immunoprecipitation, we found that prp45(1-169) mutation causes delay in U2 snRNP recruitment to assembling spliceosome. This delay transfers to the later...
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Splicing Factors in the Regulation of Gene Expression - the Relationship Between Splicing and Transcription in Saccharomyces cerevisiae
Hálová, Martina ; Folk, Petr (advisor) ; Pospíšek, Martin (referee) ; Staněk, David (referee)
Transcription and pre-mRNA processing, e.g., splicing, occur at the same place and time in the context of chromatin. A growing amount of evidence supports the hypothesis that these processes are interconnected. Prp45/SKIP is one of the factors which are believed to mediate the interconnection. The human ortholog, SKIP, is known for affecting mRNA formation on the levels of transcription initiation and elongation. Moreover, it interacts with chromatin modifiers and it is a splicing factor, too. The function of the Saccharomyces cerevisiae ortholog, Prp45, has been so far connected only to pre-mRNA splicing. In this work, we characterized the role of Prp45 in splicing and elaborated the results connecting Prp45 to transcription and chromatin modifications. RNA-seq results showed that pre-mRNA is accumulated in prp45(1-169) cells. This accumulation is not caused by the reduced activity of pathways responsible for RNA degradation. The extent of the splicing inefficiency in prp45(1-169) cells did not depend on either the canonicity of the 5' splice site and branch site or the distance between the branch site and the 3' splice site. Using chromatin immunoprecipitation, we found that prp45(1-169) mutation causes delay in U2 snRNP recruitment to assembling spliceosome. This delay transfers to the later...
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RWCT methods at the start of the school attendance
Hálová, Martina ; Tomková, Anna (advisor) ; Hejlová, Helena (referee)
The aim of this thesis was to study RWCT methods used at the beginning of a school attendance and to study appropriate conditions for applying RWCT methods. The thesis deals with searching for the best method of the very initial pupils reading and also appropriate teaching materials with regard to RWCT. There is described a three-phase model of E-U-R education and selected method of critical thinking. The practical part of the thesis contains preparation for lessons processed using RWCT methods and summarizes results of pedagogical action research. The outcomes of the thesis confirm that if pupils have suitable conditions at the beginning of a school attendance, they are able to use RWCT methods in practise.
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The purinosome - a multi-protein complex involved in the de novo purine synthesis
Kráčmarová, Markéta ; Zikánová, Marie (advisor) ; Hálová, Martina (referee)
The purinosome is a multiprotein complex involved in the de novo purine synthesis (DNPS). Through a several steps of this metabolic pathway 5-phosphoribosyl-1- pyrophosphate is converted to inosinmonophosphate which is the precursor of purine nucleotides. Purine nucleotides are also synthesized from inosinmonophosphate through a salvage pathway that utilizes hypoxanthine. The purinosome is a dynamic multienzyme complex which is assembled and diassembled by actual need and availability of purines. The purinosome assembly is disrupted particularlly in the inherited disorders of the DNPS enzymes - AICA-ribosiduria and adenylosuccinate lyase deficiency (dADSL). Detailed studies of assembly and dynamics of purinosome and identification of molecular changes associated with the formation of purinosome under physiological and pathological conditions are object of research. Besides better understanding of purine metabolism in the future it could open up new possibilities of drug development especially of chemotherapeutics that block DNPS. Key words: purinosome, de novo purine synthesis, defects of enzymes, metabolism, purinosome interactions, cell control
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Recycling of spliceosomal complexes
Klimešová, Klára ; Staněk, David (advisor) ; Hálová, Martina (referee)
Most human genes are composed of coding sequences (exons) that are interrupted by non-coding sequences (introns). After gene transcription into pre-mRNA, these introns have to be removed in a process called splicing. Splicing is mediated by a very complex and dynamic complex called the spliceosome, which consists of five small nuclear ribonucleoprotein particles (snRNPs) and numerous additional splicing proteins. Each particle contains single small nuclear RNA and a set of specific proteins. SnRNPs are assembled by a stepwise process that takes place both in the nucleus and the cytoplasm and final maturation steps occur in nuclear Cajal bodies. The mature snRNPs interact with pre-mRNA in an ordered pathway and form the spliceosome that catalyzes two trans-esterification reactions leading to intron excision and exons ligation. Subsequently, the spliceosome disassembles again into individual snRNPs that have undergone diverse conformational and compositional transformations during splicing. Thus, before the particles can participate in another round of splicing they have to go through recycling to recover their original form. However, currently the recycling phase of the splicing cycle is surrounded by more questions than answers. The purpose of this work is to discuss latest findings that shed some light on...
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Chromatin modifiers and their relation to transcription regulation in Saccharomyces cerevisiae
Hálová, Martina ; Folk, Petr (advisor) ; Staněk, David (referee)
Relations among transcription, pre-mRNA processing and chromatin modifications are only partially understood. The human protein SNW1/SKIP belongs to factors which couple these processes. The protein plays role in pre-mRNA splicing and transcription on the level of both initiation and elongation. According to the hypothesis of K. Jones laboratory, it physically and functionally interacts with positive transcription elongation factor b during transcription elongation and influences methylation of histone H3 on lysine 4, a modification characteristic for active transcription (Bres et al., Genes Dev. 19:1211-26, 2005, Bres et al., Mol Cell. 36:75-87, 2009). The yeast ortholog of SNW1/SKIP, Prp45, was until now reported only in connection with splicing regulation. However, unpublished results from our Laboratory and others showed that it is employed in transcription elongation as well. The aim of the diploma project was to search for the relations between Prp45 and the factors regulating transcription. It was confirmed that the mutation prp45(1 169) results in the delay of PHO5 and PHO84 expression during transcriptional induction. Next, we discovered new genetic interactions between PRP45 and several genes encoding the effectors of chromatin modifications. How Prp45 influences the expression of PHO5 and PHO84...
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