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Sequencing mRNA from cryo-sliced Drosophila embryos to determine genome-wide spatial patterns of gene expression

ABSTRACT: We sequenced mRNA from transverse slices of 3 wild-type and 3 high bcd copy-number embryos at the blastoderm stage to determine genome-wide patterns of gene expression. mRNA from transverse sections of single D. melanogaster embryos was sequenced

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Peter Combs

PROVIDER: E-GEOD-43506 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Sequencing mRNA from cryo-sliced Drosophila embryos to determine genome-wide spatial patterns of gene expression

Project description:We sequenced mRNA from transverse slices of embryos at the blastoderm stage to determine genome-wide patterns of gene expression.

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Genome-wide measurement of spatial expression in mutants of Drosophila Melanogaster

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Impacts of the Ubiquitous Factor Zelda on Bicoid-dependent DNA-binding and Transcription in Drosophila

Project description:In vivo crosslinking studies suggest that the Drosophila transcription factor Bicoid (Bcd) binds to several thousand sites during early embryogenesis, but it is not clear how many of these binding events are functionally important. In contrast, reporter gene studies have identified more than 60 Bcd-dependent enhancers, all of which contain clusters of the consensus binding sequence TAATCC. These studies also identified clusters of TAATCC motifs (inactive fragments) that failed to drive Bcd-dependent activation. In general, active fragments showed higher levels of Bcd-binding in vivo, and were enriched in predicted binding sites for the ubiquitous maternal protein Zelda (Zld). Here we test the role of Zld in Bcd-mediated binding and transcription. Removal of Zld function and mutations in Zld sites caused significant reductions in Bcd-binding to known enhancers, and variable effects on the activation and spatial positioning of Bcd-dependent expression patterns. Genome-wide binding experiments in Zld mutants showed variable effects on Bcd-binding peaks, ranging from strong reductions to significantly enhanced levels of binding. Increases in Bcd binding caused the precocious Bcd-dependent activation of genes that are normally not expressed in early embryos, suggesting that Zld controls the genome-wide binding profile of Bcd at the qualitative level, and is critical for selecting target genes for activation in the early embryo. These results underscore the importance of combinatorial binding in enhancer function, and provide data that will help predict regulatory activities based on DNA sequence. 1-3 hour wild type and zelda mutant embryos were examined for Bicoid binding using Illumina HiSeq 2000. The Bcd antibody is a rabbit polyclonal antiserum raised against full length Drosophila Bcd protein (Ochoa-Espinosa et al. 2009 PNAS). The antiserum was further purified using the Protein A Antibody Purification Kit (Sigma) for ChIP-seq.

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