Opa is a late-acting, pioneer factor that coordinates with Zelda to broadly regulate gene expression in early embryos (ATAC-seq, RNA-seq)
Pioneer factors such as Zelda (Zld) help initiate zygotic transcription in Drosophila early embryos, but whether other factors support this dynamic process is unclear. Odd-paired (Opa), a zinc-finger transcription factor expressed at cellularization, controls the transition of genes from pair-rule to segmental patterns along the anterior-posterior axis. Finding that Opa also regulates expression through enhancer sog_Distal along the dorso-ventral axis, we hypothesized Opa's role is more general. Chromatin-immunoprecipitation (ChIP-seq) confirmed its in vivo binding to sog_Distal but also identified widespread binding throughout the genome, comparable to Zld. Furthermore, chromatin assays (ATAC-seq) demonstrate that Opa, like Zld, influences chromatin accessibility genome-wide at cellularization, suggesting both are pioneer factors with common as well as distinct targets. Lastly, embryos lacking opa exhibit widespread, late patterning defects spanning both axes. Collectively, these data suggest Opa is a general timing factor and likely late-acting pioneer factor that drives a secondary wave of zygotic gene expression contributor: Theodora Koromila (ATAC-seq-RNA-seq) contributor: Fan Gao (Bioinformatic analysis) contributor: Angelike Stathopoulos (PI) contributor: Peng He (Bioinformatic analysis) Overall design: 1. Genome-wide data from mid nc14B and nc14D Drosophila single-embryo Assay for Transposase-Accessible Chromatin followed by sequencing (ATAC-seq) provide insight into wt versus Opa (sh opa and UAS-opa). ATAC-seq libraries were generated from the Illumina HiSeq2500 platform. HOMER (version 4.7, parameters: -localSize 50000 -minDist 50 -size 150 -fragLength 0) (Heinz et al. 2010) was used to call ATAC peaks. 2. Following total RNA isolation from control and sh_opa single embryos at nc14D, RNA was quality controlled and quantified using a Bioanalyzer. Differential expression analysis was performed using Cuffdiff module of Cufflinks with default parameters, and FPKM.