Ontology highlight
ABSTRACT:
PROVIDER: PRJNA821839 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18559085_1.fastq.gz | Fastqsanger.gz | |||
| SRR18559085_2.fastq.gz | Fastqsanger.gz | |||
| SRR18559086_1.fastq.gz | Fastqsanger.gz | |||
| SRR18559086_2.fastq.gz | Fastqsanger.gz | |||
| SRR18559087_1.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 12 |
Similar Datasets
Project description:Sumoylation is emerging as a post-translation modification important for chromosome duplication and stability. The origin recognition complex (ORC), which directs DNA replication initiation by loading the MCM replicative helicases onto origins, is sumoylated in both yeast and human cells. However, the biological consequences of ORC sumoylation are largely unclear. Here we report the effects of hyper- and hypo-sumoylation of yeast ORC using multiple approaches. We show that ORC hyper-sumoylation preferentially reduces the activity of a subset of early origins, while Orc2 hypo-sumoylation has an opposing effect. Mechanistically, ORC hyper-sumoylation leads to reduced MCM loading in vitro and diminished MCM chromatin association in vivo. The importance of an appropriate level of ORC sumoylation is suggested by the data that either hyper- or hypo-sumoylation of ORC results in genome instability and a dependence on other genome maintenance factors for cell fitness. Thus, yeast ORC sumoylation status needs to be fine-tuned to achieve optimal origin activity control and genome stability.
2023-03-11 | PXD032977 | Pride
Project description:The evolutionarily conserved Origin Recognition Complex (ORC), plays a key role in origin selection in eukaryotes. However, ORC is strikingly divergent in its DNA binding specificity, ranging from base-specific interactions in Saccharomyces cerevisiae to base-agnostic interactions in humans. The mechanisms underlying this distinct selectivity is unknown. Atomic model of the yeast ORC showed that base-specific interaction with the invariant thymines of the ARS consensus sequence (ACS) is encoded by a 19-amino acid insertion helix (IH) embedded in the winged helix domain (WHD) of Orc4. This IH is absent in the Orc4 of metazoans including humans, suggesting that removal of the IH might give the yeast ORC “human-like” properties. Indeed, yeast strain engineered with IH deficient Orc4 has completely altered ORC-binding sites enriched in poly-dT tracts located in larger nucleosome-depleted and intergenic open chromatin. In vivo and in vitro assays show that the mutant ORC loads MCM efficiently, in spite of its altered specificity in favor of binding patterns more characteristic of those observed in humans/metazoans. This work provides insights for understanding how ORC evolves to adopt a life cycle that requires plasticity in origin selection during development.
2020-10-18 | GSE149163 | GEO
Project description:The Origin Recognition Complex (ORC) is essential for the initiation of eukaryotic chromosome replication as it loads the replicative helicase, the minichromosome maintenance (MCM) complex, at replication origins. Origins display a stereotypic chromatin structure with nucleosome depletion at ORC binding sites and flanking arrays of regularly spaced nucleosomes. Although discovered long ago, it is unknown how this chromatin struture is established and if it matters for replication. By genome-scale biochemical reconstitution we screened ~300 origins and 17 purified budding yeast chromatin factors and found that ORC established nucleosome depletion over origins and flanking nucleosome arrays by orchestrating the chromatin remodelers INO80, ISW1a, ISW2 and Chd1. The functional importance of ORC’s chromatin organizing activity was directly demonstrated by ORC mutations that maintained classical MCM loader activity but lost array generation activity. These mutations impaired replication through chromatin in vitro and were lethal in vivo. Our results establish that ORC, besides its canonical role as MCM loader, has a second essential function as master regulator of the origin chromatin structure that is a crucial prerequisite for efficient chromosome replication.
2023-03-30 | GSE209681 | GEO
Project description:The budding yeast genome is marked by 250-350 origins of DNA replication. These origins are bound by the origin recognition complex (ORC) throughout the cell cycle. ORC has known DNA binding sequence preferences which, though necessary for binding, are not sufficient to fully specify a genomic locus as being bound by ORC, indicating that the cell must use additional chromosomal cues to specify ORC binding sites and origins of replication. Using high-throughput sequencing to precisely locate both ORC binding sites and nucleosome locations genome-wide, we find that a nucleosome depleted region (NDR) and precisely positioned nucleosomes are a ubiquitous feature of yeast replication origins. The ARS consensus sequence (ACS) and adjacent sequences are sufficient to maintain the nucleosome-free properties of the NDR. We use a temperature sensitive ORC1 mutant to demonstrate that ORC is required to maintain precisely positioned nucleosomes at origins of replication. These findings demonstrate the importance of local nucleosome positioning at replication origins, and that chromatin organization is an important determinant of origin selection. Examination of nucleosome positioning in wild-type and orc1-161ts mutant S. cerevisiae at room temperature and heatshock temperatures. Examination of ORC binding locations by ChIP-seq. All reported coordinates are based on the SGD genome build released 12/16/2005.
2010-03-19 | E-GEOD-16926 | biostudies-arrayexpress
Project description:The budding yeast genome is marked by 250-350 origins of DNA replication. These origins are bound by the origin recognition complex (ORC) throughout the cell cycle. ORC has known DNA binding sequence preferences which, though necessary for binding, are not sufficient to fully specify a genomic locus as being bound by ORC, indicating that the cell must use additional chromosomal cues to specify ORC binding sites and origins of replication. Using high-throughput sequencing to precisely locate both ORC binding sites and nucleosome locations genome-wide, we find that a nucleosome depleted region (NDR) and precisely positioned nucleosomes are a ubiquitous feature of yeast replication origins. The ARS consensus sequence (ACS) and adjacent sequences are sufficient to maintain the nucleosome-free properties of the NDR. We use a temperature sensitive ORC1 mutant to demonstrate that ORC is required to maintain precisely positioned nucleosomes at origins of replication. These findings demonstrate the importance of local nucleosome positioning at replication origins, and that chromatin organization is an important determinant of origin selection.
2010-03-19 | GSE16926 | GEO
Project description:The Origin Recognition Complex (ORC) seeds replication-fork formation by binding to DNA replication origins, which in budding yeast contain a 17bp DNA motif. High resolution structure of the ORC-DNA complex revealed two base-interacting elements: a disordered basic patch (Orc1-BP4) and an insertion helix (Orc4-IH). To define the ORC elements guiding its DNA binding in vivo, we mapped genomic locations of 38 designed ORC mutants, revealing that different ORC elements guide binding at different sites. At silencing-associated sites lacking the motif, ORC binding and activity were fully explained by a BAH domain. Within replication origins, we reveal two dominating motif variants showing differential binding modes and symmetry: a non-repetitive motif whose binding requires Orc1-BP4 and Orc4-IH, and a repetitive one where another basic patch, Orc1-BP3, can replace Orc4-IH. Disordered basic patches are therefore key for ORC-motif binding in vivo, and we discuss how these conserved, minor-groove interacting elements can guide specific ORC-DNA recognition.
2024-03-18 | GSE249817 | GEO
Project description:SUMOylation fine-tunes the transcriptional activity of endothelial HEY1 in the regulation of angiogenesis
| PRJNA1006642 | ENA
Project description:Eukaryotic DNA replication origins are selected in G1-phase when the origin recognition complex (ORC) binds chromosomal DNA, triggering a series of molecular events that culminate in the initiation of DNA replication (a.k.a. origin firing) during S-phase. Each chromosome requires multiple origins for its duplication, and each origin fires at a characteristic time during S-phase, creating a cell-type specific genome replication pattern with relevance to differentiation, genome stability and evolution. It is unclear whether ORC-origin interactions are relevant to the regulation of origin activation time. Here we applied a novel genome-wide strategy to classify origins in the model eukaryote Saccharomyces cerevisiae based on the types of molecular interactions used for ORC-origin binding. Specifically, origins were classified as DNA-dependent when the strength of ORC-origin binding in vivo could be explained by the affinity of ORC for origin DNA in vitro, and, conversely, as ‘chromatin-dependent’ when the ORC-DNA interaction in vitro was insufficient to explain the strength of ORC-origin binding in vivo. The two classes of origins were distinct in terms of local nucleosome architecture and dependence on origin-flanking sequences in plasmid replication assays, consistent with local features of chromatin promoting ORC binding at ‘chromatin-dependent’ origins. Importantly, origins that fired in late S-phase, but not those that fired in early S-phase, showed a significant dependence on the ORC-origin DNA interaction for ORC binding in vivo. Therefore we demonstrate for the first time a connection between ORC-origin binding mechanisms and the regulation of origin activation time at a genome-wide level.
2013-09-30 | GSE48440 | GEO
Project description:Post-translational modification with SUMO is known to regulate the activity of transcription factors, but how SUMOylation of individual proteins might influence immunity is mostly unexplored. The NFAT transcription factors play an essential role in antigen receptor-mediated gene regulation. SUMOylation of NFATc1 represses IL-2 in vitro, but its role in T cell-mediated immune responses in vivo is not clear. To this end, we generated a novel Nfatc1 transgenic mouse, which prevents SUMO modification of NFATc1. Avoidance of NFATc1 SUMOylation ameliorated experimental autoimmune encephalomyelitis as well as graft-versus-host disease. An elevated IL-2 production promoted Treg expansion and suppressed autoreactive or alloreactive T cells. Mechanistically, increased IL-2 secretion counteracted IL-17 and IFN-γ expression through STAT5 and Blimp-1 induction. Then, Blimp-1 repressed IL-2 itself and the as well induced, proliferation-associated survival factor Bcl2A1. Collectively, we demonstrate that prevention of NFATc1 SUMOylation fine-tunes T-cell responses towards lasting tolerance. Thus, targeting NFATc1 SUMOylation presents a novel and promising strategy to treat T cell-mediated inflammatory diseases.
2020-07-01 | GSE148087 | GEO
Project description:The origin recognition complex (ORC) marks chromosomal sites as replication origins and is essential for replication initiation. In yeast, ORC also binds to DNA elements called silencers, where its primary function is to recruit silent information regulator (SIR) proteins to establish transcriptional silencing. Indeed, silencers function poorly as chromosomal origins. Several genetic, molecular, and biochemical studies of HMR-E have led to a model proposing that when ORC becomes limiting in the cell, such as in the orc2-1 mutant, only sites that bind ORC tightly, such as HMR-E, remain fully occupied by ORC, while lower affinity sites, including most origins, lose ORC occupancy. Since HMR-E possessed a unique non-replication function, we reasoned that other tight sites might reveal novel functions for ORC on chromosomes. Therefore, we comprehensively determined ORC “affinity” genome-wide by performing an ORC ChIP-on-chip in ORC2 and orc2-1 strains. Here we describe a novel group of orc2-1-resistant ORC-interacting chromosomal sites (ORF-ORC sites) that did not function as replication origins or silencers. Instead, ORF-ORC sites were comprised of protein-coding regions of highly transcribed metabolic genes. In contrast to the ORC-silencer paradigm, transcriptional activation promoted ORC association with these genes. Remarkably, ORF-ORC genes were enriched in proximity to origins of replication, and, in several instances, were transcriptionally regulated by these origins. Taken together, these results suggest a surprising connection between ORC, replication origins and cellular metabolism.
2009-11-07 | GSE18895 | GEO