Project description:The submitted data have been utilized in the following two papers: "Drosophila poised enhancers are generated during tissue patterning with the help of repression" (Koenecke et al 2016, Genome Research): Histone modifications are frequently used as markers for enhancer states, but how to interpret enhancer states in the context of embryonic development is not clear. The poised enhancer signature, involving H3K4me1 and low levels of H3K27ac, has been reported to mark inactive enhancers that are poised for future activation. However, future activation is not always observed and alternative reasons for the widespread occurrence of this enhancer signature have not been investigated. By analyzing enhancers during dorsal-ventral (DV) axis formation in the Drosophila embryo, we find that the poised enhancer signature is specifically generated during patterning in the tissue where the enhancers are not induced, including at enhancers that are known to be repressed by a transcriptional repressor. These results suggest that, rather than serving simply as an intermediate step before future activation, the poised enhancer state may mark enhancers for spatial activation during tissue patterning. We discuss the possibility that the poised enhancer state is more generally the result of repression by transcriptional repressors. "Genome-wide identification of Drosophila dorso-ventral enhancers by differential histone acetylation analysis" (Koenecke and Johnston et al 2016, Genome Biol): Background: Drosophila dorso-ventral (DV) patterning is one of the best-understood regulatory networks to date, and illustrates the fundamental role of enhancers in controlling patterning, cell fate specification and morphogenesis during development. Histone acetylation such as H3K27ac is an excellent marker for active enhancers, but it is challenging to obtain precise locations for enhancers as the highest levels of this modification flank the enhancer regions. How to best identify tissue-specific enhancers in a developmental system de novo with a minimal set of data is still unclear. Results: Using DV patterning as a test system, we develop a simple and effective method to identify tissue-specific enhancers de novo. We sample a broad set of candidate enhancer regions using data on CBP co-factor binding or ATAC-seq chromatin accessibility, and then identify those regions with significant differences in histone acetylation between tissues. This method identifies hundreds of novel DV enhancers and outperforms ChIP-seq data of relevant transcription factors when benchmarked with mRNA expression data and transgenic reporter assays. These DV enhancers allow the de novo discovery of the relevant transcription factor motifs involved in DV patterning and contain additional motifs that are evolutionarily conserved and for which the corresponding transcription factors are expressed in a DV-biased fashion. Finally, we identify novel target genes of the regulatory network, implicating morphogenesis genes as early targets of DV patterning. Conclusions: Taken together, our approach has expanded our knowledge of the DV patterning network even further and is a general method to identify enhancers in any developmental system, including mammalian development.

Project description:Spatiotemporal gene regulation during embryonic development is driven by cis-regulatory DNA sequences called enhancers. Enhancers are activated through a combination of transcription factors (TFs) that bind to short sequence motifs within these sequences, but the order of events by which TFs read out motifs is not clear. Some TFs can only bind chromatin that is already accessible, while other TFs called pioneers can open chromatin themselves. Identifying motifs and the order by which they drive chromatin accessibility is very challenging. The recent implementation of convolutional neural networks, which learn complex cis-regulatory sequence rules that are predictive for genomics data, provides an unprecedented opportunity to dissect this problem. Here, we trained base-resolution deep learning models and applied them to high-resolution TF binding and chromatin accessibility data from the well-studied early Drosophila embryo. We uncover a clear hierarchical relationship between the pioneer Zelda and the TFs involved in the spatiotemporal patterning of the embryo, consistent with Zelda being a pioneer. However, the models predict that patterning TFs can also augment chromatin accessibility in a context-specific manner. Using a series of Drosophila mutant strains, we find that the two types of TFs increase chromatin accessibility by distinct mechanisms. Zelda’s pioneering is proportional to motif affinity, while the patterning TFs specifically increase chromatin accessibility when they mediate enhancer activation. This was conclusive because Dorsal can function both as activator and repressor, and the effect on chromatin accessibility depended on Dorsal’s transactivation effect and not on its binding per se. In conclusion, chromatin accessibility occurs in two phases: one through pioneering, which makes regions first accessible but not necessarily active, and a second when the correct combination of transcription factors lead to enhancer activation.

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Genome-wide profiles of chromatin accessibility in spatially-restricted domains along the antero-posterior axis of Drosophila blastoderm

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