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Damidseq_pipeline: an automated pipeline for processing DamID sequencing datasets

ABSTRACT: DamID is a powerful technique for identifying regions of the genome bound by a DNA-binding (or DNA-associated) protein. Currently no method exists for automatically processing next-generation sequencing DamID (DamID-seq) data, and the use of DamID-seq datasets with normalisation based on read-counts alone can lead to high background and the loss of bound signal. DamID-seq thus presents novel challenges in terms of normalisation and background minimisation. We describe here damidseq_pipeline, a software pipeline that performs automatic normalisation and background reduction on multiple DamID-seq FASTQ or BAM datasets. Single replicate profiling of pol II occupancy in 3rd instar larval neuroblasts of Drosophila

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Owen Marshall

PROVIDER: E-GEOD-69184 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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damidseq_pipeline: an automated pipeline for processing DamID sequencing datasets

Project description:DamID is a powerful technique for identifying regions of the genome bound by a DNA-binding (or DNA-associated) protein. Currently no method exists for automatically processing next-generation sequencing DamID (DamID-seq) data, and the use of DamID-seq datasets with normalisation based on read-counts alone can lead to high background and the loss of bound signal. DamID-seq thus presents novel challenges in terms of normalisation and background minimisation. We describe here damidseq_pipeline, a software pipeline that performs automatic normalisation and background reduction on multiple DamID-seq FASTQ or BAM datasets.

2015-06-26 | GSE69184 | GEO

A direct role for cohesin in genome-wide gene regulation and Ecdysone-response in Drosophila salivary gland

Project description:Developmental abnormalities observed in Cornelia de Lange Syndrome (CdLS) have been genetically linked to mutations in the cohesin machinery. These findings raise the possibility that cohesin, in addition to its canonical function of mediating sister chromatid cohesion, might also be involved in regulating gene expression. We report that cleavage of cohesin’s kleisin subunit in post-mitotic Drosophila salivary glands induces major changes (both up and down) in the transcript levels of many genes. Kinetic analyses of changes in transcript levels upon cohesin cleavage reveal that a subset of genes responds to cohesin cleavage within a few hours. In addition, cohesin binds to most of these loci, suggesting that cohesin is directly regulating their expression. Amongst these genes are several that are regulated by the steroid hormone Ecdysone. Transcripts at EcR and Eip74EF, which encode an Ecdysone Receptor and an Ecdysone-regulated transcription factor, respectively, decline ten-fold within four hours of cohesin cleavage. Cytological visualization of transcription at selected Ecdysone-responsive genes reveals that puffing at Eip74EF ceases within an hour or two of cohesin cleavage, long before any decline in EcR associated with this locus. We conclude that cohesin regulates expression of a distinct set of genes, including those mediating the Ecdysone response DamID experiments for Rad21 were performed on gDNA from drosophila salivary glands. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. The experiment was done in duplicate in the reverse dye orientation.

2011-03-22 | E-GEOD-21874 | biostudies-arrayexpress

Mapping of Polycomb binding in Drosophila larval brain tissue using DamID

Project description:We made Polycomb (PC) and histone H3 lysine 27 trimethylation (H3K27me3) chromatin binding maps in central brain tissue from 3rd instar larvae, allowing us to make a direct comparison to our 4C data (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE23166). Our results demonstrate that our PC and H3K27me3 maps are highly similar, and fit with previously identified hallmarks of PcG-bound chromatin, namely: PC and H3K27me3 co-occur in the genome as large contiguous domains that largely repress transcription of the underlying genes, which encode important regulators of development. Keywords: Genome binding/occupancy profiling by genome tiling array DamID experiments for Polycomb were performed in Drosophila larval brain tissue. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing.

2011-12-31 | E-GEOD-23298 | biostudies-arrayexpress

Drosophila COP9 Signalosome Subunit 7 interacts with multiple genomic loci

Project description:The COP9 signalosome protein complex has a central role in the regulation of development of multi-cellular organisms. While the function of this complex in ubiquitin-mediated protein degradation is well established, results over the past few years have hinted that the COP9 signalosome may function more broadly in the regulation of gene expression. Here, using DamID technology, we show that COP9 signalosome subunit 7 functionally associates with a large number of genomic loci in the Drosophila genome, and show that the expression of many genes within these loci is COP9 signalosome-dependent. This association is likely direct as we show CSN7 binds DNA in vitro. The genes targeted by CSN7 are preferentially enriched for transcriptionally active regions of the genome, and are involved in the regulation of distinct gene ontology groupings including imaginal disc development and cell-cycle control. DamID experiments were done in Drosophila Kc167 cells. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. The experiment contains 4 samples, three biological replicates and one reverse dye orientation. For each sample Dam-fusion material was hybridized over Dam-only material.

2014-09-22 | E-GEOD-61605 | biostudies-arrayexpress

Examination of three chromatin proteins in slivary glands in presence or absence of SuUR mutation using DamID technique

Project description:Heterochromatin contains repressively modified histones and replicates late in S phase of the cell cycle. Besides the shortage in replication origins, little is known about replication timing regulation in silenced regions. In Drosophila polytene cells, late replication results in under-replication and decreased DNA copy number in heterochromatic regions of the genome. The Suppressor of Under-replication (SUUR) protein controls this feature â?? in its absence the DNA polytenization level in most silenced regions is restored, however the repressive histone marks are lost. We hypothesized that SUUR regulates the re-establishment of repressive histone pattern during replication which results in delayed replication completion of heterochromatin. Measuring DNA copy number in mutants with disrupted repressive pathways, we found that under-replication is directly linked to repressive histone marks supply. DamID-seq and ChIP-seq experiments revealed that SuUR mutation does not affect the establishment of heterochromatin domains. Here, we identified a novel SUUR protein interaction (CG12018) that supports SUUR association with replication complex. SUUR loads onto replication forks shortly after the origin firing and participates in chromatin maintenance rather than its establishment. Thus, our findings provide comprehensive evidence that late replication in Drosophila is caused by the time-consuming process of replication-coupled repressive chromatin renewal. Examination of three chromatin proteins in slivary glands in presence or absence of SuUR mutation using DamID technique.

2016-01-10 | E-GEOD-74907 | biostudies-arrayexpress

DamID profiling of proneural transcription factor binding in Drosophila melanogaster wing imaginal discs

Project description:Genome-wide identification of the binding sites of the Drosophila transcription factors Achaete, Asense, E(spl)m3-HLH and Senseless in wing imaginal cells using DamID profiling. Each Dam-fusion-derived sample is compared to a control Dam-only sample. Two biological replicates were performed for sca-Asense, neur-Asense, sca-Achaete, neur-Achaete, neur-Sens and sca-E(spl)m3-HLH.

2014-01-01 | E-GEOD-53009 | biostudies-arrayexpress

DamID of Dam::LMN-1 and Dam::EMR-1 in Caenorhabditis elegans adult worms

Project description:Laminopathies are caused by mutations in components of the nuclear envelope (NE). While most NE components are widely expressed, laminopathies affect only a subset of tissues. However, the understanding of the molecular mechanisms that explain this phenomenon is still elusive. Here we have performed a genome wide DamID analysis in adult C. elegans nematodes comparing the DNA association profile of two components of the NE, Lamin/LMN-1 and Emerin/EMR-1. Although both proteins were associated to silent DNA, EMR-1 showed a predominant role in the anchoring of muscle and neuronal promoters to the nuclear periphery. Deletion of either EMR-1 or LEM-2, another integral NE protein, caused local changes in nuclear architecture with both increased and decreased LMN-1 association. Comparison of Dam::LMN-1 and Dam::EMR-1 DNA assotiation in wild type strains and Dam::LMN-1 DNA association in wild type, lem-2(tm1582) and emr-1(gk119) mutant backgrounds.

2013-12-02 | E-GEOD-44188 | biostudies-arrayexpress

DamID data for Comr binding in drosophila testes

Project description:Comr protein was found to be a major regulator of gene activity in drosophila spermatocytes. We obtained Comr binding profile to determine targets of Comr. Comr binding in drosophila male germ line cells was determined using DamID technique. Comparison of Dam-Comr binding to Dam-alone signal in duplicate for each sample type.

2013-08-13 | E-GEOD-49799 | biostudies-arrayexpress

Identifying targets of DSX with DamID-seq

Project description:We have adapted the DamID protocol for use with high throughput sequencing. We have used DamID to identify the positions within the Drosophila genome where the transcription factor DSX is bound. We sequenced DpnI-digested genomic DNA from fly tissues containing UAS-Dam (control) or UAS-Dam-DsxF or UAS-Dam-DsxM. We have performed DamID-seq on adult male and female fatbody and on ovary. We used two biological replicates for each tissue and sex.

2014-10-30 | E-GEOD-27269 | biostudies-arrayexpress

DamID data for SUUR binding in 12-14h Embryo, larval salivary glands and larval brain

Project description:SUUR protein was found to be a major component of three repressive chromatin states named BLACK, BLUE and GREEN chromatins. Here we addressed a question of developmental dynamics of these chromatin states using SUUR as a marker. SUUR binding in larval salivary glands, brains and 12-14 hours embryos was determined using DamID technique. Comparison of Dam-SUUR binding to Dam-alone signal in duplicate for each sample type.

2011-11-23 | E-GEOD-33873 | biostudies-arrayexpress

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