Assaying chromatin accessibility using ATAC-seq in invertebrate chordate embryos

Cis-regulatory elements (CREs) are non-coding DNA regions involved in the spatio-temporal regulation of gene expression. Gene regulatory changes drive animal development and play major roles during evolution of animal body plans. Therefore, we believe that determining CREs at different developmental stages and across animal lineages is critical to understand how evolution operates through development. The Assay for Transposase-Accessible Chromatin followed by high-throughput sequencing (ATAC-seq) is a powerful technique for the study of CREs that takes advantage of Tn5 transposase activity. Starting from fewer than 105 cells, in a 1-day procedure, it is possible to detect, at a genome-wide level, CREs located in open chromatin regions with high resolution. Here, we describe a detailed step-by-step ATAC-seq protocol for invertebrate chordate marine embryos. We have successfully applied this technique to amphioxus and two species of tunicate embryos. We also show an easy workflow to analyze data generated with this technique. Moreover, we point out that this method and our bioinformatic pipeline are efficient to detect CREs associated with Wnt signaling pathway by simply using embryos treated with a drug that perturbs this pathway. This approach can be extended to other signaling pathways and also to embryo mutants for critical genes. Our results therefore demonstrate the power of ATAC-seq for the identification of CREs that play essential functions during animal development in a wide range of invertebrate or vertebrate animals.This project has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 Research and Innovation Program (Grant Agreement No. 740041), the Spanish Ministerio de Economía y Competitividad (Grant No. BFU2016-74961-P), and the Andalusian Government (Grant No. BIO-396). This work was also supported by the institutional grant Unidad de Excelencia María de Maeztu (MDM-2016-0687 to the Department of Gene regulation and morphogenesis of Centro Andaluz de Biología del Desarrollo) and by the EMBO short-term fellowship. The laboratory of HE and SB was supported by the Centre National de la Recherche Scientifique (CNRS) and the Agence Nationale de la Recherche (Grant No. ANR-16-CE12-0008-01).Peer reviewe