Ontology highlight
ABSTRACT:
PROVIDER: PRJNA615974 | ENA |
REPOSITORIES: ENA
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Project description:Fused in sarcoma regulates DNA replication timing and progression
| PRJNA616354 | ENA
Project description:Fused-in-sarcoma (FUS) encodes an RNA-binding protein with diverse roles in transcriptional activation and RNA splicing. While oncogenic fusions of FUS and transcription factor DNA-binding domains are associated with soft tissue sarcomas, dominant mutations in FUS cause amyotrophic lateral sclerosis (ALS). FUS has also been implicated in genome maintenance. However, the underlying mechanisms are unknown.
Here, we applied gene editing, functional reconstitution and integrated proteomic and transcriptomics to illuminate roles for FUS in DNA replication and repair. Consistent with a supportive role in DNA double-strand break (DSB) repair FUS deficient cells exhibited subtle alterations in the recruitment and retention of DSB-associated factors, including 53BP1 and BRCA1. FUS-/- cells also exhibited reduced proliferative potential that correlated with reduced replication fork speed, diminished loading of pre-replication complexes, enhanced micronucleus formation, and attenuated expression and splicing of S-phase associated genes. Finally, FUS-deficient cells exhibited genome-wide alterations in DNA replication timing that were reversed upon reeexpression of FUS cDNA. FUS-dependent replication domains were enriched in transcriptionally active chromatin and FUS was required for the timely replication of transcriptionally active DNA. These findings suggest that alterations DNA replication kinetics and programming contribute to genome instability and functional defects in FUS deficient cells.
2021-06-24 | MSV000087698 | MassIVE
Project description:SAMHD1 is a triphosphohydrolase and a 3'-5' exonuclease that restricts HIV-1 infection in non-cycling cells. It is also mutated in the Aicardi-Goutières Syndrome (AGS) and in different cancers, including chronic lymphotic leukemia. We report herre that SAMHD1 localizes to DNA replication foci during S phase suggesting that it is involved in DNA synthesis. Using microarray analysis, we examine the replication timing program an we show that SAMHD1 regulates the fork progression but also the replication timing program.
2017-07-29 | GSE77392 | GEO
Project description:Control of DNA copy number is essential to maintain genome stability and ensure proper cell and tissue function. In Drosophila, the SNF2-domain-containing SUUR protein inhibits replication fork progression within specific regions of the genome to promote DNA underreplication. While dissecting the function of SUUR’s SNF2 domain, we identified a physical interaction between SUUR and Rif1. Rif1 has many roles in DNA metabolism and regulates the replication timing program. We demonstrate that repression of DNA replication is dependent on Rif1. Rif1 localizes to active replication forks in an SUUR-dependent manner and directly regulates replication fork progression. Importantly, SUUR associates with replication forks in the absence of Rif1, indicating that Rif1 acts downstream of SUUR to inhibit fork progression. Our findings uncover an unrecognized function of the Rif1 protein as a direct regulator of replication fork progression suggesting developmental regulation of Rif1 activity.
2018-10-15 | GSE114370 | GEO
Project description:Fused in sarcoma (FUS) encodes a low complexity RNA-binding protein with diverse roles in transcriptional activation and RNA processing. While oncogenic fusions of FUS and transcription factor DNA-binding domains are associated with soft tissue sarcomas, dominant mutations in FUS cause amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). FUS has also been implicated in DNA double-strand break repair (DSBR) and genome maintenance. However, the underlying mechanisms are unknown. Here we employed quantitative proteomics, transcriptomics, and DNA copy number analysis (Sort-Seq), in conjunction with FUS-/- cells to ascertain roles of FUS in genome protection. FUS-/- cells exhibited alterations in the recruitment and retention of DSBR factors BRCA1 and 53BP1 but were not overtly sensitive to genotoxins. By contrast, FUS-deficient cells had reduced proliferative potential that correlated with reduced replication fork speed, diminished loading of pre-replication complexes, and attenuated expression of S-phase associated genes. FUS interacted with lagging strand DNA synthesis factors and other replisome components, but did not translocate with active replication forks. Using a Sort-Seq workflow, we show that FUS contributes to genome-wide control of DNA replication timing and is essential for the early replication of transcriptionally active DNA. These findings illuminate new roles for FUS in DNA replication initiation and timing that may contribute to genome instability and functional defects in cells harboring disease-associated FUS fusions.
2020-05-13 | GSE147784 | GEO
Project description:DNA replication is spatially and temporally regulated during S-phase. DNA replication timing is established in early G1-phase at a point referred to as TDP (timing decision point). We show that Rif1 (Rap1-interacting-factor1), originally identified as a telomere binding factor in yeast, is a critical determinant of the replication timing program in human cells. Depletion of Rif1 results in specific loss of mid-S replication foci profiles, stimulation of initiation events in early S-phase and changes in long-range replication timing domain structures. Overall replication timing is shifted toward mid-S in both directions, suggesting that replication timing regulation is abrogated in the absence of Rif1. Rif1 tightly binds to nuclear insoluble structures at late-M to early-G1 and regulates the chromatin-loop sizes. Furthermore, Rif1 colocalizes specifically with the mid-S replication foci. Thus, Rif1 establishes the mid-S replication domains that are restrained from being activated at early S-phase. Our results indicate that Rif1 plays crucial roles in determining the replication timing domain structures through regulating higher-order chromatin architecture.
2012-05-15 | GSE37971 | GEO
Project description:DNA replication timing and 3D chromatin organisation are associated with epigenomic changes across large domains during human differentiation and cancer progression. However, it is unclear if epigenome changes, in particular cancer-associated DNA hypomethylation, is a consequence or cause of changes observed in higher order genome architecture. Here, we compare replication timing profiles and three dimensional (3D) genome organisation, using Hi-C and single cell Repli-Seq in the DNMT1 and DNMT3B DNA methyltransferases double knockout hypomethylated DKO1 colorectal cancer cell line and its parental HCT116 cell line. We find that the hypomethylated cells show a profound loss of replication timing precision, gain of single cell replication timing heterogeneity and loss of chromatin conformation integrity. Discrete regions, that undergo a large change in replication timing in the hypomethylated cells, are associated with a loss of allelic replication timing and shrinking of late replicating Partially Methylated Domain (PMD) boundaries. In contrast, conservation of replication timing after DNA methylation depletion at PMDs is associated with the formation of new H3K9me3/H3K4me3 bivalent domains which may serve to prevent ectopic transcription and maintain cell viability. Together our results show that a loss of global methylation, a common hallmark of cancer, directly impacts on the precision of replication timing and contribute to deregulation of the 3D chromatin architecture.
2021-09-21 | GSE159663 | GEO
Project description:Chromosomal DNA replication involves the coordinated activity of hundreds to thousands of replication origins. Individual replication origins are subject to epigenetic regulation of their activity during S-phase, resulting in differential efficiencies and timings of replication initiation during S-phase. This regulation is thought to involve chromatin structure and organization into timing domains with differential ability to recruit limiting replication factors. Rif1 has recently been identified as a genome-wide regulator of replication timing in fission yeast and in mammalian cells. However, previous studies in budding yeast have suggested that Rif1’s role in controlling replication timing may be limited to subtelomeric domains and derives from its established role in telomere length regulation. We have analyzed replication timing by analyzing BrdU incorporation genome-wide, and report that Rif1 regulates the timing of late/dormant replication origins throughout the S. cerevisiae genome. Analysis of pfa4∆ cells, which are defective in palmitoylation and membrane association of Rif1, suggests that replication timing regulation by Rif1 is independent of its role in localizing telomeres to the nuclear periphery. Intra-S checkpoint signaling is intact in rif1∆ cells, and checkpoint-defective mec1∆ cells do not comparably deregulate replication timing, together indicating that Rif1 regulates replication timing through a mechanism independent of this checkpoint. Our results indicate that the Rif1 mechanism regulates origin timing irrespective of proximity to a chromosome end, and suggest instead that telomere sequences merely provide abundant binding sites for proteins that recruit Rif1. Still, the abundance of Rif1 binding in telomeric domains may facilitate Rif1-mediated repression of non-telomeric origins that are more distal from centromeres. 4 samples BrdU-IP-seq in HU, 2 strains with 2-replicates each (strains:WT and rif1 delta)
2014-06-01 | E-GEOD-55155 | biostudies-arrayexpress
Project description:DNA replication is spatially and temporally regulated during S-phase. DNA replication timing is established in early G1-phase at a point referred to as TDP (timing decision point). We show that Rif1 (Rap1-interacting-factor1), originally identified as a telomere binding factor in yeast, is a critical determinant of the replication timing program in human cells. Depletion of Rif1 results in specific loss of mid-S replication foci profiles, stimulation of initiation events in early S-phase and changes in long-range replication timing domain structures. Overall replication timing is shifted toward mid-S in both directions, suggesting that replication timing regulation is abrogated in the absence of Rif1. Rif1 tightly binds to nuclear insoluble structures at late-M to early-G1 and regulates the chromatin-loop sizes. Furthermore, Rif1 colocalizes specifically with the mid-S replication foci. Thus, Rif1 establishes the mid-S replication domains that are restrained from being activated at early S-phase. Our results indicate that Rif1 plays crucial roles in determining the replication timing domain structures through regulating higher-order chromatin architecture. HeLa cells (ATCC), with a total of 4 individual replicates
2012-05-14 | E-GEOD-37971 | biostudies-arrayexpress
Project description:In multicellular organisms, developmental changes to replication timing occur in 400- 800 kb domains across half the genome. While clear examples of epigenetic control of replication timing have been described, a role for DNA sequence in mammalian replication timing has not been substantiated. To assess the role of DNA sequences in directing these changes, we profiled replication timing in mice carrying a genetically rearranged Human Chromosome 21 [Hsa21]. In two distinct mouse cell types, Hsa21 sequences maintained human-specific replication timing, except at points of Hsa21 rearrangement. Changes in replication timing at rearrangements extended up to 900 kb and consistently reconciled with the wild-type replication pattern at developmental boundaries of replication-timing domains. Our results demonstrate DNA sequencedriven regulation of Hsa21 replication timing during development and provide evidence that mammalian chromosomes consist of multiple independent units of replication timing regulation.
2012-06-05 | GSE38472 | GEO