Global mapping of ribonucleotides embedded in DNA
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE61464 | GEO | 2015/01/15
SECONDARY ACCESSION(S): PRJNA261234
REPOSITORIES: GEO
Similar Datasets
Project description:Capture and massively parallel DNA sequencing of ribonucleotides embedded in S. cerevisiae genomic DNA We developed a new method to map the positions of ribonucleotides embedded in DNA using the unique specificity of A. thaliana tRNA ligase. Ribonucleotides were generated in budding yeasts of different genetic backgrounds and mapped to single nucleotide resolution using the new method.
2015-01-15 | E-GEOD-61464 | biostudies-arrayexpress
Project description:Yeast Saccharomyces cerevisiae has been widely used as a model system for studying genome instability. Here, heterozygous S. cerevisiae zygotes were generated to determine the genomic alterations induced by sudden introduction of active RNase H2. In combination of a custom SNP microarray, the patterns of chromosomal instability could be explored at a whole genome level. Ribonucleotides can be incorporated into DNA during replication by the replicative DNA polymerases. These aberrant DNA subunits are efficiently recognized and removed by Ribonucleotide Excision Repair, which is initiated by the heterotrimeric enzyme RNase H2. While RNase H2 is essential in higher eukaryotes, the yeast Saccharomyces cerevisiae can survive without RNase H2 enzyme, although the genome undergoes mutation, recombination and other genome instability events at an increased rate. Although RNase H2 can be considered as a protector of the genome from the deleterious events that can ensue from recognition and removal of embedded ribonucleotides, under conditions of high ribonucleotide incorporation and retention in the genome in a RNase H2-negative strain, sudden introduction of active RNase H2 causes massive DNA breaks and genome instability in a condition which we term “ribodysgenesis”. The DNA breaks and genome instability arise solely from RNase H2 cleavage directed to the ribonucleotide-containing genome. Survivors of ribodysgenesis have massive loss of heterozygosity events stemming from recombinogenic lesions on the ribonucleotide-containing DNA, with increases of over 1000X from wild-type. DNA breaks are produced over one to two divisions and subsequently cells adapt to RNase H2 and ribonucleotides in the genome and grow with normal levels of genome instability.
2022-03-26 | GSE199262 | GEO
Project description:We report the application of high through-put tag sequencing to measure the location and strand of DNA embedded ribonucleotides in the yeast genome. Mutations in the catalytic subunits of the polymerases (pol1-L868M, pol2-M644G and pol3-L612M) lead to the increased incorporation of ribonucleotides during DNA replication, providing an in vivo label with which to track the contribution of each polymerase to the fully replicated genome. Yeast strains used in this study are deleted for rnh201, encoding the catalytic subunit of the RNase H2 gene so that embedded ribonucleotides are not rapidly removed by ribonucleotide excision repair following DNA replication. Analysis of this data demonstrates that polymerase alpha contributes to the fully replicated genome. Sequencing of DNA embedded ribonucleotides in S. cerevisiae strains to map the contribution of replicative polymerases to the fully replicated genome.
2015-01-26 | E-GEOD-64521 | biostudies-arrayexpress
Project description:Misincorporation of ribonucleotides into DNA during genome replication has recently become recognized as a significant source of genomic instability. The frequency of ribonucleotides in the genome is determined by dNTP/rNTP ratios, the ability of the DNA polymerases to discriminate against ribonucleotides, and by the capacity of repair mechanisms to remove misincorporated ribonucleotides. To simultaneously compare how the nuclear and mitochondrial genomes incorporate and remove ribonucleotides, we challenged these processes by imbalancing cellular dNTP pools. Using a collection of yeast strains with altered dNTP pools, we discovered an inverse relationship between the concentration of individual dNTPs and the amount of the corresponding ribonucleotides incorporated in mitochondrial DNA, while in nuclear DNA the ribonucleotide pattern was only altered in the absence of ribonucleotide excision repair. Our analysis uncovers major differences in ribonucleotide repair between the two genomes and provides concrete evidence that yeast mitochondria lack mechanisms for repair of misincorporated ribonucleotides.
2017-11-07 | GSE100352 | GEO
Project description:Ribodysgenesis: sudden genome instability in the yeast Saccharomyces cerevisiae arising from RNase H2 cleavage at genomic-embedded ribonucleotides
| PRJNA819077 | ENA
Project description:We report here the consequences of loss of Saccharomyces cerevisiae RNase H2 by comparing mRNA expression profiles in wild type yeast versus a strain lacking the RNH201 gene, encoding the catalytic subunit. Deleting RHN201 alters mRNA expression for hundreds of genes. Expression changes were based on seven biological replicates (3 from one batch and 4 from another batch) and are seen for many genes involved in stress responses and genome maintenance, consistent with a role of RNase H2 in removing ribonucleotides incorporated into DNA during replication. Differentially expressed genes include, those involved in ribosomal RNA processing and biogenesis and in tRNA modification, supports a role for RNase H2 in resolving R-loops formed during transcription of rRNA and tRNA. Other differentially expresses genes include putative or known helicases and nucleases maintenance of dsRNA viruses, which could be relevant to how mammalian RNase H2 dysfunction elicits an innate immune response leading to neurodegeneration. We examine the consequences of loss of Saccharomyces cerevisiae RNase H2 by comparing mRNA expression profiles in wild type yeast versus a strain lacking the RNH201 gene encoding the catalytic subunit. Expression changes were based on seven biological replicates (3 from one batch and 4 from another batch). For strain reference please see S.A. Nick McElhinny, et al, Genome instability due to ribonucleotide incorporation into DNA, Nat Chem Bio 6, 774-781 (2010).
2012-12-20 | E-GEOD-34668 | biostudies-arrayexpress
Project description:We report here the consequences of loss of Saccharomyces cerevisiae RNase H2 by comparing mRNA expression profiles in wild type yeast versus a strain lacking the RNH201 gene, encoding the catalytic subunit. Deleting RHN201 alters mRNA expression for hundreds of genes. Expression changes were based on seven biological replicates (3 from one batch and 4 from another batch) and are seen for many genes involved in stress responses and genome maintenance, consistent with a role of RNase H2 in removing ribonucleotides incorporated into DNA during replication. Differentially expressed genes include, those involved in ribosomal RNA processing and biogenesis and in tRNA modification, supports a role for RNase H2 in resolving R-loops formed during transcription of rRNA and tRNA. Other differentially expresses genes include putative or known helicases and nucleases maintenance of dsRNA viruses, which could be relevant to how mammalian RNase H2 dysfunction elicits an innate immune response leading to neurodegeneration.
2012-12-20 | GSE34668 | GEO
Project description:We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing.
2012-04-25 | GSE37003 | GEO
Project description:Multiple DNA polymerases are needed to replicate genetic information. Here we describe the use of ribonucleotide incorporation as a biomarker of replication enzymology in vivo. We find that ribonucleotides are incorporated into the yeast nuclear genome in replicase specific and strand-specific patterns that identify replication origins and where polymerase switching occurs. Ribonucleotide density varies across the genome as a function of the replicase, base, local sequence and proximity to nucleosomes and transcription start sites. Ribonucleotides are present in one strand at high densityat mitochondrial replication origins, implying unidirectional replication of a circular genome. The evolutionary conservation of the enzymes that incorporate and process ribonucleotides in DNA suggests that the use of ribonucleotides as biomarkers of DNA synthesis in cells will have widespread applicability. Mapping genomic ribonucleotides in 14 Saccharomyces cerevisiae strains (seven DNA polymerase backgrounds, with or without RNH201), via HydEn-seq (end sequencing of genomic fragments generated by alkaline hydrolysis).
2015-01-26 | E-GEOD-62181 | biostudies-arrayexpress
Project description:We developed a novel approach, m6A-seq, for high-resolution mapping of the transcriptome-wide m6A landscape, based on antibody-mediated capture followed by massively parallel sequencing.
2012-04-25 | GSE37002 | GEO