GSE169227
Global patterns of enhancer activity during sea urchin embryogenesis assessed by eRNA profiling
We used CAGE-seq (Capped Analysis of Gene Expression with Sequencing) to profile eRNA expression and enhancer activity during embryogenesis of the sea urchin, Strongylocentrotus purpuratus. We identified >18,000 enhancers that were active during late oogenesis and early development and documented a burst of enhancer activation during cleavage and early blastula stages. Most enhancers were located near gene bodies and eRNA expression levels were highest for elements near core promoters. Transcriptional signals from enhancers generally paralleled the expression levels of likely target genes. Furthermore, enhancers near lineage-specific genes contained signatures of inputs from developmental gene regulatory networks deployed in those lineages. A large fraction (60%) of sea urchin enhancers previously shown to be active in transgenic reporter assays were associated with eRNA expression. Moreover, a large fraction (50%) of a representative subset of enhancers identified by eRNA profiling drove tissue-specific gene expression in isolation when tested by reporter assays. Our findings provide an atlas of developmental enhancers in a model sea urchin and support the utility of eRNA profiling as a tool for enhancer discovery and regulatory biology. The data generated in this study are publicly available at Echinobase (www.echinobase.org). Overall design: CAGE library preparation, sequencing, mapping, and eRNA identification were performed by DNAFORM (Yokohama, Kanagawa, Japan). Briefly, 30-60 mg total RNA was used for the generation of each CAGE library (9 total) and libraries were subjected to Illumina HiSeq-based sequencing. Sequence reads (>100 million reads/sample) were subjected to quality control (FastQC), filtered to remove small numbers of rRNA reads, and mapped to the S. purpuratus genome (v. 3.1) first with BWA (Li and Durbin, 2010) and then using HISAT2 (Kim et al., 2019) to align reads with BWA MAPQ<20. A total of 60-80 million uniquely mapped reads were obtained per sample and used for all subsequent analysis. De novo eRNA peak-calling was carried out as described by Hirabayashi et al., 2019. Briefly, bidirectional enhancers were identified using the FANTOM5 pipeline (FANTOM CONSRTIUM, 2014). TSSs were identified according to http://fantom.gsc.riken.jp/5/sstar/Protocols:HeliScopeCAGE_read_alignment. and clustered using the decomposition-based peak identification (DPI) software (https://github.com/hkawaji/dpi1/blob/master/identify_tss_peaks.sh). DPI was used with default parameters but without the decomposition parameter. Peaks with at least two supporting CAGE tags were retained and used as input to call bidirectional enhancers using a program available at https://github.com/anderssonrobin/enhancers/blob/master/scripts/bidir_enhancers.