Transcription profiling by array of Drosophila embryonic chordotonal neurons expressing the cato GFP reporter gene construct
Ontology highlight
ABSTRACT:
ORGANISM(S): Drosophila melanogaster
SUBMITTER: Petra zur Lage
PROVIDER: E-MTAB-7529 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:A challenge in systems biology is to understand the gene regulatory networks that connect early cellular specification to terminal differentiation of specific cell types. In neurogenesis, neural specification has been well studied, but the link between the proneural transcriptional regulators of specification and the genes that must be activated to construct differentiated neurons is obscure. High resolution temporal profiling of gene expression reveals the events downstream of Atonal (Ato) proneural gene function in Drosophila sensory neurons. Unexpectedly, many differentiation genes are activated soon after specification, even before cell cycle exit and overt neuronal differentiation. Prominent among them are genes required for construction of the ciliary dendrite. Ato activates differentiation both directly and indirectly via several intermediate transcriptional regulators, including Rfx and a new Forkhead family factor. Our analysis of these factors and their regulation provides insight into how proneural factors regulate neuronal subtype differentiation. Investigating the molecular mechanisms of peripheral nervous sytem development in Drosophila melanogaster. Affymetrix Drosophila version 2.0 chips used to measure gene expression from GFP+ and GFP- cells from embryos expressing GFP under the control of the atonal gene enhancer in both wild type and mutant embryos. Data generated for three developmental time-points in quadruplicate.
2011-02-18 | E-GEOD-21520 | biostudies-arrayexpress
Project description:A genetic study of the PRF1 gene has shown association of several polymorphisms with multiple sclerosis (MS). Haplotype analysis identified risk haplotypes strongly associated with male patients having the primary-progressive form of MS (PPMS). Gene expression microarrays were performed in 10 male PPMS patients carrying the risk (n=6) and protective haplotypes (n=4) in order to identify pathways associated with the risk haplotypes. Pathway analysis revealed overrepresentation of the cell killing gene ontology category among down-regulated genes in patients carrying risk haplotypes compared with patients carrying protective haplotypes. Number of samples analyzed: 10 Protective haplotype samples: UOM982, EMA1473, MMC-998, CDP1842 Risk haplotype samples: UUS1554, RAU1550, RPS1011, AGS1013, PFB1530, MGA1014
2010-07-28 | E-GEOD-23205 | biostudies-arrayexpress
Project description:The mechanisms by which pulmonary lesions and fibrosis are generated during SARS-CoV infection are not known. Using high-throughput mRNA profiling, we examined the transcriptional response of wild-type (WT), type I interferon receptor knockout (IFNAR1−/−), and STAT1 knockout (STAT1−/−) mice infected with a recombinant mouse-adapted SARS-CoV (rMA15) to better understand the contribution of specific gene expression changes to disease progression. Ten week old 129S6/SvEv wild-type, STAT1−/− (Taconic Farms, Germantown, NY), and IFNAR1−/− mice bred on a 129SvEv background were anesthetized with a ketamine and infected intranasally with either phosphate-buffered saline (PBS) alone (Invitrogen, Carlsbad, CA) or 1 × 10^5 PFU rMA15-PBS. Mice were euthanized and left lungs were harvested from individual mice (a total of 3 infected mice from each strain) at days 2, 5, and 9 postinfection (dpi) for microarray analyses. Lung samples were taken from mock-infected animals from each of the strains at 5 dpi.
2012-02-22 | E-GEOD-36016 | biostudies-arrayexpress
Project description:Background: Differential gene expression specifies the highly diverse cell types that constitute the nervous system. With its sequenced genome and simple, well-defined neuroanatomy, the nematode C. elegans is a useful model system in which to correlate gene expression with neuron identity. The UNC-4 transcription factor is expressed in thirteen embryonic motor neurons where it specifies axonal morphology and synaptic function. These cells can be marked with an unc-4::GFP reporter transgene. Here we describe a powerful strategy, Micro-Array Profiling of C. elegans cells (MAPCeL), and confirm that this approach provides a comprehensive gene expression profile of unc-4::GFP motor neurons in vivo. Results: Fluorescence Activated Cell Sorting (FACS) was used to isolate unc-4::GFP neurons from primary cultures of C. elegans embryonic cells. Microarray experiments detected 6,217 unique transcripts of which ~1,000 are enriched in unc-4::GFP neurons relative to the average nematode embryonic cell. The reliability of these data was validated by the detection of known cell-specific transcripts and by expression in UNC-4 motor neurons of GFP reporters derived from the enriched data set. In addition to genes involved in neurotransmitter packaging and release, the microarray data include transcripts for receptors to a remarkably wide variety of signaling molecules. The added presence of a robust array of G-protein pathway components is indicative of complex and highly integrated mechanisms for modulating motor neuron activity. Over half of the enriched genes (537) have human homologs, a finding that could reflect substantial overlap with the gene expression repertoire of mammalian motor neurons. Conclusion: We have described a microarray-based method, MAPCeL, for profiling gene expression in specific C. elegans motor neurons and provide evidence that this approach can reveal candidate genes for key roles in the differentiation and function of these cells. These methods can now be applied to generate a gene expression map of the C. elegans nervous system. Experiment Overall Design: Our goal is to profile gene expression throughout the nervous system of the model organism Caenorhabditis elegans. As a first goal, we profiled a single class of embryonic motor neurons. To isolate transcripts from thesec neurons we developed the MAPCeL (Microarray Profiling C. elegans Cells) technique in which unc-4::GFP+ cells are captured by FACS for RNA isolation. We verified these data by bioinformatic means and by in vivo validation by creating GFP reporters for a random set of genes in our enriched gene list.
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Project description:Stable zebrafish cell lines were created that expressed GFP under the control of a previously characterized mouse Smarcd3 enhancer (10.7554/eLife.03848). Specifically, the 2.7 kb Mouse Smarcd3-F6 sequence (chr5: 24113559 -24116342 from mm9 assembly) was sub-cloned from a gateway entry vector into the Zebrafish Enhancer Detection (ZED). Tol2-mediated transgenesis was performed and stable lines were created. The Smarcd3-F6:EGFPhsc70 allele was used for all genomics experiments. Smarcd3-F6:EGFPhsc70 embryos were dissociated at 10 hours post fertilization for fluorescent activated cell sorting (FACS) to collect GFP+ cells. Single-cell cDNA libraries were prepared using Fluidgim C1 system ((Fluidgim, PN 100-7168 Rev. B1)). 96 single-cell libraries were collected from three batches of experiments and sequenced on the Illumina HiSeq 2500 platform. Data from 92 cells (number of genes detected > 2000) were kept for clustering analysis. For the tube control experiments we performed mRNA-seq on ~4000 cells from the same embryo batches as the scRNA-seq experiments. GFP negative cells were also included to enable a GFP+ versus GFP- comparison. Libraries were made following the Fluidgim C1 mRNA-seq tube control protocol (Fluidgim, PN 100-7168 Rev. B1). Polyadenylated mRNA was captured by Oligo-dT primers and PCR amplified after reverse transcription. Final sequencing libraries were made using Nextera XT DNA Sample Preparation Kit and pair-end sequenced on the Illumina HiSeq 2500 platform.
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Project description:We, being members of modENCODE consortium, have established an experimetal pipeline in C.elegans that allows global identification of the binding sites for transcription factors using chromatin immunoprecipitation followed by illumina high-throughput sequencing (ChIP-seq). In current study, we identified the binding sites for CEH-14 at L2 stage, which is required for specification of the AFD thermosensory neurons and for normal thermotactic behavior. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf CEH-14 was tagged with a dual GFP:3xFLAG tag at the carboxy terminus, whose expression pattern was confirmed through both fluorescence imaging and immunoblot analysis. The binding sites were determined using ChIP-seq. Two biological replicates were used in this study, but replicate1 is at a later L2 stage. Only one Input sample (non-immunoprecipitated chromatin (input) from replicate2 was used as a control. The concordance between two replicates is just above 50% when p-value is 0.05.
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