A common set of DNA regulatory elements shapes Drosophila appendages
Ontology highlight
ABSTRACT:
ORGANISM(S): Drosophila melanogaster
PROVIDER: GSE38727 | GEO | 2013/11/07
SECONDARY ACCESSION(S): PRJNA168643
REPOSITORIES: GEO
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Project description:We mapped open chromatin by FAIRE-seq and measured gene expression by RNA-seq in 3 types of Drosophila samples: staged whole embryos, imaginal discs, pharate appendages. We first demonstrate that regions of open chromatin precisely define regions of enhancer activity in developing embryos. In contrast to the dynamic changes in open chromatin observed between different stages of embryogenesis, we found that the open chromatin profiles in wing, leg, and haltere imaginal discs are nearly identical. This was also true again later in development, where the adult appendages also share nearly identical open chromatin profiles. Therefore, at a given developmental time point, different appendages are specified using a shared set of DNA regulatory elements. However, from one time point to the next, the set of accessible regulatory elements changes. Open chromatin profiles in appendage imaginal discs are almost entirely different than those of the adult appendages. We propose that master regulator transcription factors create morphologically distinct structures by differentially influencing the function of the same set of DNA regulatory modules. Open chromatin profiling during Drosophila development: 3 stages of embryogenesis (2-replicates each); wing, leg, and haltere 3rd instar imaginal discs (3-replicates each); 3rd larval central nervous system (2-replicates); eye-antennal imaginal discs (2-replicates); wing, leg, and haltere pharate appendages (2-replicates each); Genomic DNA Inputs. Sequencing performed on Illumina GAII and HiSeq.
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Project description:The Hox genes are responsible for generating morphological diversity along the anterior-posterior axis during animal development. The Drosophila Hox gene Ultrabithorax (Ubx), for example, is required for specifying the identity of the third thoracic (T3) segment of the adult, which includes the dorsal haltere, an appendage required for flight, and the ventral T3 leg. Ubx mutants show homeotic transformations of the T3 leg towards the identity of the T2 leg and the haltere towards the wing. All Hox genes, including Ubx, encode homeodomain containing transcription factors, raising the question of what target genes Ubx regulates to generate these adult structures. To address this question, we carried out whole genome ChIP-chip studies to identify all of the Ubx bound regions in the haltere and T3 leg imaginal discs, which are the precursors to these adult structures. In addition, we used ChIP-chip to identify the sites bound by the Hox cofactor, Homothorax (Hth). This is a dataset generated by the Drosophila Regulatory Elements modENCODE Project led by Kevin P. White at the University of Chicago. This dataset was generated in collaboration with Richard S. Mann at Columbia University. It contains ChIP-chip data on Affymetrix Drosophila Tiling 2.0R arrays for multiple transcription factor antibodies in multiple Drosophila tissues. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf Haltere or leg imaginal discs ChIPped for Ubx or Hth vs. input DNA from corresponding imaginal discs. For each combination of tissue and antibody, ChIP experiments have been performed and hybridized on Affymetrix Drosophila Tiling 2.0R arrays. At least 3 biological replicates for the ChIP sample have been hybridized.
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Project description:Hox proteins are transcription factors and key regulators of segmental identity along the anterior posterior axis across all bilateral animals. Despite decades of research, mechanism by which Hox proteins select their targets and specify segmental identity remains elusive. To address this question we carried out whole genome ChIP-chip experiments to identify direct targets of Hox protein Ultrabithorax (Ubx) during haltere development in Drosophila. When mis-expressed in wing segment (T2) Ubx converts its identity to that of haltere segment (T3). We used CbxHm/+ wing discs ectopically expressing Ubx in the pouch region of discs to obtain chromatin. This helped us focus on targets of Ubx involved in pouch development without mixing with the targets involved in notum development. Polyclonal Ubx antibodies against N-terminal region (excluding homeodomain) were generated in our lab and used to pull down Ubx bound regions from CbxHm/+ wing discs. Mock DNA (No antibody) was used as control. Test Vs. Mock experiment. CbxHm/+ discs for Chromatin source. Biological replicates: 3 [Agilent two-color ChIP-on-Chip experiment,Organism: Drosophila melanogaster ,Drosophila Whole Genome ChIP-on-Chip Set 244K Microarray 1 of 2 (AMADID: 014816 and 014817)]
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Project description:Hox proteins have been proposed to act at multiple levels within regulatory hierarchies and to directly control the expression of a plethora of target genes. However, for any specific Hox protein or tissue, very few direct in vivo-regulated target genes have been identified. Here, we have identified target genes of the Hox protein Ultrabithorax (UBX), which modifies the genetic regulatory network of the wing to generate the haltere, a modified hindwing. We used whole-genome microarrays and custom arrays including all predicted transcription factors and signaling molecules in the D. melanogaster genome to identify differentially expressed genes in wing and haltere imaginal discs. To elucidate the regulation of selected genes in more detail, we isolated cis-regulatory elements (CREs) for genes that were specifically expressed in either the wing disc or haltere disc. We demonstrate that UBX binds directly to sites in one element, and these sites are critical for activation in the haltere disc. These results indicate that haltere and metathoracic segment morphology is not achieved merely by turning off the wing and mesothoracic development programs, but rather specific genes must also be activated to form these structures. The evolution of haltere morphology involved changes in UBX-regulated target genes, both positive and negative, throughout the wing genetic regulatory network. Keywords: tissue comparison
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Project description:We profiled the chromatin accessibility, transcription factor binding, and gene expression in the homologous wing and haltere imaginal disc of Drosophila melanogaster, which are distinguished by the expression of the Hox protein Ultrabithorax (Ubx). Through the analysis of chromatin changes, we show Ubx has widespread effects on the chromatin landscape to specify cell identity.
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