Microarray and ChIP-chip of Rrm3-Flag in wild-type and npl3 cells, and ChIP-chip of Npl3-Myc in wild type cells
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
SUBMITTER: AndrM-CM-)s Aguilera
PROVIDER: E-GEOD-50187 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Transcription is a major obstacle for replication fork progression and a cause of genome instability. Such instability increases in mutants with a suboptimal assembly of the nascent messenger ribonucleo-protein particle (mRNP), as THO/TREX and some heterogeneous nuclear ribonucleoproteins (hnRNPs) mutants. Here we show that yeast npl3M-bM-^HM-^F cells show genome-wide replication obstacles as determined by accumulation of the Rrm3 helicase. Such obstacles preferentially occur at long and highly expressed genes, to which Npl3 is preferentially bound in wild-type cells. ChIP-chip studies were perfomed with antibodies against Myc-tagged Npl3 protein in wild-type cells of the yeast S. Cerevisiae, as well as Flag-tagged Rrm3 protein in both wild-type and npl3M-bM-^HM-^F cells.
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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2014-10-08 | E-GEOD-56703 | biostudies-arrayexpress
Project description:Transcription is a major contributor to genome instability.A main cause of transcription-associated instability relies on the capacity of transcription to stall replication. Such genome instability is increased in RNAPII mutants. ChIP-chips performed in asynchronous cultures showed an increase of the Rrm3 binding signal all over the genome in rpb1-1 compared to wild-type. ChIP-chip studies were perfomed with antibody against Flag-tagged Rrm3 protein in both wild-type and rpb1-1 cells.
2014-12-04 | E-GEOD-55184 | biostudies-arrayexpress
Project description:Transcription is a major obstacle for replication fork progression and a cause of genome instability. Yra1 is an essential nuclear factor of the evolutionarily conserved family of hnRNP-like export factors that when overexpressed impairs mRNA export and cell growth. Through this ChIP-chip analysis it is shown that Yra1 binds to active chromatin and is enriched at telomeres when it is overexpressed, in agreement with a possible role of this mRNP factor in the maintenance of telomere integrity. Our data indicate that YRA1 overexpression correlates with replication impairment as inferred by the increase of Rrm3, a helicase involved in the replication fork progression, at transcribed genes and telomeres. ChIP-chip studies were perfomed with antibodies against HA-tagged Yra1 protein in wild-type cells and cells overexpressing YRA1 of the yeast S. Cerevisiae, as well as Flag-tagged Rrm3 protein in both wild-type and cells overexpressing YRA1.
2016-03-11 | E-GEOD-68486 | biostudies-arrayexpress
Project description:Transcription is a major obstacle for replication fork progression and a cause of genome instability. Such instability increases in mutants with a suboptimal assembly of the nascent messenger ribonucleo-protein particle (mRNP), as THO/TREX and the NPC-associated THSC/TREX-2 complex. Here we show that yeast sac3M-bM-^HM-^F and thp1M-bM-^HM-^F cells accumulate genome-wide replication obstacles as determined by the distribution of the Rrm3 helicase. Such obstacles preferentially occur at long and highly expressed genes, to which Sac3 and its interacting partner Thp1 are preferentially bound in wild-type cells. ChIP-chip studies were perfomed with antibodies against Flag-tagged Thp1 and Sac3 proteins in wild-type cells of the yeast S. Cerevisiae, as well as Flag-tagged Rrm3 protein in sac3M-bM-^HM-^F and thp1M-bM-^HM-^F cells that were compared with Rrm3 in wild-type cells from Santos-Pereira et al., 2013 (accession number GSE50185).
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2016-03-11 | E-GEOD-68488 | biostudies-arrayexpress
Project description:Transcription is a major obstacle for replication fork progression and transcription-replication collisions constitute a main cause of genome instability. At a genome-wide scale these obstacles can be detected by the accumulation of the replicative Rrm3 helicase required for RF progression through protein obstacles. Here we show that FACT, a chromatin-reorganizing complex that swaps nucleosomes around the RNA polymerase during transcription elongation and that also has a role in replication, is needed to resolve transcription-replication conflicts in Saccharomyces cerevisiae. Importantly, ChIP-chip analyses of Rrm3 reveal that replication progression impairment in FACT mutants occur genome-wide, but preferentially at highly transcribed regions. ChIP-chip studies were perfomed with antibodies against Rrm3-FLAG in the yeast S. cerevisiae.
2014-03-17 | E-GEOD-51653 | biostudies-arrayexpress
Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis and prevents transcription-associated recombination. Whether or not it has a ubiquitous role in the genome is an open question. ChIP-chip studies reveal that the Hpr1 component of THO and the Sub2 RNA-dependent ATPase have genome wide-distributions at active ORFs in yeast. In contrast to RNAPII, evenly distributed from promoter to termination regions, THO and Sub2 are absent at promoters and distributed in a sharp 5M-bM-^@M-^YM-bM-^FM-^R3M-bM-^@M-^Y gradient. Importantly, ChIP-chips reveal an over-recruitment of Rrm3 in active genes in THO mutants that is reduced by overexpression of RNase H1. Our work establishes a genome-wide function for THO-Sub2 in transcription elongation and mRNP biogenesis that function to prevent the accumulation of transcription-mediated replication obstacles, including R-loops. ChIP-chip studies were perfomed with tagged forms of the Hpr1 component of THO (Hpr1-FLAG), the Sub2 RNA-dependent ATPase of TREX (Sub2-FLAG), the Rpb3 subunit of RNA polymerase II (Rpb3-PK) and the Rrm3 protein (Rrm3-FLAG) in the yeast S. cerevisiae.
2011-06-28 | E-GEOD-24960 | biostudies-arrayexpress
Project description:Replication stress activates the Mec1ATR and Rad53 kinases. Rad53 phosphorylates nuclear pores to counteract gene gating, thus preventing aberrant transitions at forks approaching transcribed genes. Here, we show that Rrm3 and Pif1, DNA helicases assisting fork progression across pausing sites, are detrimental in rad53 mutants experiencing replication stress. Rrm3 and Pif1 ablations rescue cell lethality, chromosome fragmentation, replisome-fork dissociation, fork reversal, and processing in rad53 cells. Through phosphorylation, Rad53 regulates Rrm3 and Pif1; phospho-mimicking rrm3 mutants ameliorate rad53 phenotypes following replication stress without affecting replication across pausing elements under normal conditions. Hence, the Mec1-Rad53 axis protects fork stability by regulating nuclear pores and DNA helicases. We propose that following replication stress, forks stall in an asymmetric conformation by inhibiting Rrm3 and Pif1, thus impeding lagging strand extension and preventing fork reversal; conversely, under unperturbed conditions, the peculiar conformation of forks encountering pausing sites would depend on active Rrm3 and Pif1. BrdU incorporation profiles by ssDNA-BrdU IP on chip have been generated as described (Katou et al., 2003). Protein binding profiles by ChIP-chip analysis were generated as described (Bermejo et al., 2009). Labeled probes were hybridized to Affymetrix S.cerevisiae Tiling 1.0 (P/N 900645) arrays and processed with TAS software.
2015-09-24 | E-GEOD-68214 | biostudies-arrayexpress
Project description:R-loops are transcription by-products that may constitute a threat to genome integrity. In addition to specific enzymes to remove them, eukaryotes rely on a number of mRNP biogenesis factors such as the THO complex, to prevent co-transcriptional R-loop formation. We show in Saccharomyces cerevisiae that R-loops are tightly and specifically linked with histone H3-Ser10 phosphorylation (H3S10P), a mark of chromatin condensation. Importantly, ChIP-chip analyses reveal a clear H3S10P accumulation at the pericentromeric chromatin during the G1-phase of the cell cycle only in R loop-accumulating yeast strains but not in those non-accumulating R-loops, and a significantly higher accumulation during S-phase. Such a difference can also be detected in a number of genes along the genome. ChIP-chip studies were perfomed with antibodies against Histone H3 and the phosphorylated Histone H3 at Serine10 in the yeast S. cerevisiae.
2013-11-13 | E-GEOD-46627 | biostudies-arrayexpress