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  • This SuperSeries is composed of the following subset Series: GSE26895: Drosophila LID RNAi gene expression profiling GSE27078: LID ChIP-Seq in wild type, and H3K4me3 ChIP-Seq in wild type and lid RNAi Drosophila melanogaster GSE40599: POLIISER5 and POLIISER2 ChIP-Seq in mutant RNAi LID Drosophila Melanogaster Refer to individual Series
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  • This data consists of RNA-seq data of whole animal white pre pupa of four Drosophila species: Drosophila melanogaster, Drosophila simulans, Drosophila yakuba, and Drosophila pseudoobscura. The processed RPKM values are calculated following the method in Garber et al 2011 Nature Methods paper. Examination of H3K27me3 in 4 Drosophila species and its correlation with gene expression levels in the same development stage relevant ChIP-seq data can be found in GSE25663, GSE25668
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    • Sequencing Data
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  • We use male gonads isolated from a Drosophila strain that allows us to obtain enough cells at their primitive status as the starting material to study the endogenous chromatin structure of undifferentiated cells using ChIP-seq. We integrate the ChIP-seq data with RNA-seq data that measures the transcriptome in a digital manner. Our genome-wide analyses indicate that the majority of differentiation genes in undifferentiated cells lack an active chromatin mark and paused Pol II; instead, they are associated with either the repressive H3K27me3 mark or no detectable mark. In order to address the possibility that distinct techniques are responsible for such a difference, we also use the Drosophila S2 cells to perform ChIP-seq and RNA-seq and compare the results directly with published work using ChIP-chip and microarray on S2 cells. For the S2 cell ChIP-chip data, we used data from the following paper: Muse GW, Gilchrist DA, Nechaev S, Shah R, Parker JS, Grissom SF, Zeitlinger J, Adelman K: RNA polymerase is poised for activation across the genome. /Nat Genet /2007, 39(12):1507-1511. The accession number for this data is: GSE6714. ChIP-seq: Profiling chromatin modifications using antibodies against 3 histone modifications and RNA Pol II in S2 cells Profiling chromatin structure in bam testis using antibodies against 3 histone modifications and RNA Pol II RNA-seq: Profiling transcriptome of S2 cells using RNA-seq
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    • Sequencing Data
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  • This SuperSeries is composed of the following subset Series: GSE33546: Polycomb repressive complex 2-dependent and –independent functions of Jarid2 in transcriptional regulation in Drosophila [ChIP-Seq] GSE36038: Polycomb repressive complex 2-dependent and –independent functions of Jarid2 in transcriptional regulation in Drosophila [Affymetrix] Refer to individual Series
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  • We report chromatin-associated protein and RNA interactions of HP1a, indentified by BioTAP-XL mass spectrometry and sequencing. We identify an extensive list of both known and novel HP1a-interacting proteins from Drosophila S2 cells and from whole organisms across embryonic, larval and adult stages. BioTAP-XL protocol was used to identify HP1a interacting proteins through mass-spectrometry analysis. ChIP-seq-like pulldown/input samples were generated using BioTAP-XL to validate genomic distribution of the HP1a-BioTAP contructs, and the newly identified interacting proteins (CG8290 and CG3680); To examine the RNAs associated with the HP1a complexes, the RNA fraction of the BioTAP-XL pulldowns was analyzed using Illumina-based RNA-seq protocols, as well as Helicos direct RNA sequencing.
    Data Types:
    • Sequencing Data
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  • Transcription factors, which regulate the spatiotemporal patterns of gene expression during organogenesis, often regulate multiple aspects of tissue morphogenesis, including cell-type specification, cell proliferation, cell death, cell polarity, cell shape, cell arrangement and cell migration. In this work, we describe a distinct role for Ribbon (Rib) in controlling cell shape changes during elongation of the Drosophila salivary gland (SG). Notably, the morphogenetic changes in rib mutants occurred without effects on general SG cell attributes such as specification, proliferation and apoptosis. Moreover, the abnormal cell/organ shape in rib mutants occurred without compromising epithelial-specific morphological attributes such as apicobasal polarity and junctional integrity. To identify the genes regulated by Rib that control cell and organ shape, we performed ChIP-seq analysis in embryos driving rib expression specifically in the SGs. To learn if the Rib binding sites identified in the ChIP-seq analysis were linked to changes in gene expression through transcriptional activation, repression, or both, we performed microarray analysis comparing RNA samples from age-matched wild-type and rib null embryos. From the superposed ChIP-seq and microarray gene expression data, we identified 60 genomic sites of bound Rib most likely to regulate SG-specific gene expression. We confirmed several of the identified Rib targets by qRT-pCR and/or in situ hybridization. Our results indicate that Rib regulates cell shape change in the Drosophila salivary gland via a diverse array of targets through both transcriptional activation and repression. Furthermore, our results suggest that a critical component of the SG morphogenetic gene network involving Rib is its autoregulation. Three independent collections of stage 11 – 16 rib1/ribP7 embryos and three of wild-type embryos were used for hybridization to Drosophila Genome 2.0 Chips. Scanned intensity values were normalized using RMA (Partek software) and statistical analysis analyses were performed using the Spotfire software package (TIBCO). Target genes were identified as those that were upregulated/downregulated (1.5-fold change cutoff, P < 0.05) in rib1/ribP7 embryos when compared with Oregon R controls.
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  • Throughout Metazoa, developmental processes are controlled by a surprisingly limited number of conserved signaling pathways. Precisely how these signaling cassettes were assembled in early animal evolution remains poorly understood, as do the molecular transitions that potentiated the acquisition of their myriad developmental functions. Here we analyze the molecular evolution of the proto-oncogene YAP/Yorkie, a key effector of the Hippo signaling pathway that controls organ size in both Drosophila and mammals. Based on heterologous functional analysis of evolutionarily distant Yap/Yorkie orthologs, we demonstrate that a structurally distinct interaction interface between Yap/Yorkie and its partner TEAD/Scalloped became fixed in the eumetazoan common ancestor. We then combine transcriptional profiling of tissues expressing phylogenetically diverse forms of Yap/Yorkie with ChIP-seq validation in order to identify a common downstream gene expression program underlying the control of tissue growth in Drosophila. Intriguingly, a subset of the newly-identified Yorkie target genes are also induced by Yap in mammalian tissues, thus revealing a conserved Yap-dependent gene expression signature likely to mediate organ size control throughout bilaterian animals. Combined, these experiments provide new mechanistic insights while revealing the ancient evolutionary history of Hippo signaling. We sought to define the downstream target genes of selected Yap variants by performing RNA sequencing analysis (RNA-seq) on total RNA isolated from GMR-Gal4>Yap eye discs. Transcriptional profiles were generated in triplicate from eye imaginal disks with either endogenous Yki, or GMR-Gal4 over-expressed Yki, Trichoplax Yap, Monosiga Yap, or Monisiga Yap+TEAD domain, using deep sequencing via Illumina Hi Seq.
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  • Steroid hormones act as important developmental switches and their nuclear receptors regulate many genes. However, few hormone-dependent enhancers have been characterized and important aspects of their sequence architecture, cell type-specific activating and repressing functions, or the regulatory roles of their chromatin structure have remained unclear. We used STARR-seq, a recently developed enhancer-screening assay, and ecdysone signaling in two different Drosophila cell types to derive the first genome-wide hormone-dependent enhancer activity maps. We demonstrate that enhancer activation depends on cis-regulatory motif combinations that differ between cell types and can predict cell type-specific ecdysone targeting. Activated enhancers are often not accessible prior to induction. Enhancer repression following hormone treatment is independent of receptor motifs and receptor binding to the enhancer as we show using ChIP-seq, but appears to rely on motifs for other factors, including Eip74. Our strategy is applicable to study signal-dependent enhancers for different pathways and across organisms. STARR-seq was performed in S2 and OSC cells treated with ecdysone in two replicates. DHS-seq before and after treatment was done with single-end sequencing in two replicates. RNA-seq (with and without ecdysone) was performed with a strand-specific protocol using single-end sequencing in two replicates in S2. ChIP-seq (with and without ecdysone) was performed single-end sequencing in two replicates in S2 cells.
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  • The Polycomb group (PcG) and Trithorax group (TrxG) of proteins are required for stable and heritable maintenance of repressed and active gene expression states. Their antagonistic function on gene control, repression for PcG and activity for TrxG, is mediated by binding to chromatin and subsequent epigenetic modification of target loci. Despite our broad knowledge about composition and enzymatic activities of the protein complexes involved, our understanding still lacks important mechanistic detail and a comprehensive view on target genes. In this study, we use an extensive data set of ChIP-seq, RNA-seq, and genome-wide detection of transcription start sites (TSSs) to identify and analyze thousands of binding sites for the PcG proteins and Trithorax from a Drosophila S2 cell line. In addition to finding a preference for stalled promoter regions of annotated genes, we uncover many intergenic PcG-binding sites coinciding with non-annotated transcription start sites. Interestingly, this set includes previously unknown promoters for primary transcripts of microRNA genes, thereby expanding the scope of Polycomb control to non-coding RNAs essential for development, apoptosis and growth. Chromatin from S2 cells was immunoprecipitated using antibodies against Pc, Ph, Psc, Trx-C or H3K4me3. In parallel, we isolated RNA from S2 cells and generated global gene expression profiles by RNA-seq. We also surveyed the Drosophila genome for yet non-annotated transcription start sites (TSSs) using a newly adapted protocol for Illumina sequencing (termed 5’-MACE) with RNA isolated from S2 cells and embryos.
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  • Jarid2 was recently identified as an important component of the mammalian Polycomb Repressive Complex 2 (PRC2), where it has a major effect on PRC2 recruitment in mouse embryonic stem cells. Although Jarid2 is conserved in Drosophila, it has not previously been implicated in Polycomb (Pc) regulation. Therefore, we purified Drosophila Jarid2 and its associated proteins and find that Jarid2 associates with all of the known canonical PRC2 components, demonstrating a conserved physical interaction with PRC2 in flies and mammals. Furthermore, in vivo studies with Jarid2 mutants in flies demonstrate that among several histone modifications tested, only H3K27 methylation, the mark implemented by PRC2, was affected. Genome-wide profiling of Jarid2, Su(z)12 and H3K27me3 occupancy by ChIP-seq indicates that Jarid2 and Su(z)12 have a very similar distribution pattern on chromatin. However, Jarid2 and Su(z)12 occupancy levels at some genes are significantly different with Jarid2 being present at relatively low levels at many Pc response elements (PREs) of certain Homeobox (Hox) genes, providing a rationale for why Jarid2 was never identified in Pc screens. Gene expression analyses show that Jarid2 and E(z) (a canonical PRC2 component) are required not only for transcriptional repression but might also function in active transcription. Identification of Jarid2 as a conserved PRC2 interactor in flies provides an opportunity to begin to probe some of its novel functions in Drosophila development. Expression analyses of Jarid2 mutants in larvae and eye imaginal discs. Expression analyses of E(z) RNAi in larvae.
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