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The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.

The formation of pseudouridine and 5-methyluridine (m5U) in the T-arm of transfer RNAs (tRNAs) is near-universally conserved. These two modifications are formed in Escherichia coli by the pseudouridine synthase TruB and the S-adenosylmethionine-dependent methyltransferase TrmA, respectively. In this thesis, I investigate the function and mechanisms of these two tRNA modifying enzymes. First, in vitro and in vivo analysis of TruB reveals that this enzyme is acting as a tRNA chaperone which proves a long outstanding hypothesis. Secondly, characterization of ligand binding by TrmA shows that binding is cooperative and disruption of tRNA elbow region tertiary interactions by TrmA is essential for efficient tRNA binding and catalysis, leading to future analysis. In conclusion, my studies further our understanding of the mechanism and function of tRNA modifications and modifying enzymes, as well as shed light on why all cells invest substantial resources into fine-tuning the chemical composition of tRNAs.