Towards genome-wide discovery of transcription factor binding sites contributing to planarian regeneration

Planarians are flatworms that can regenerate any missing tissue using a population of proliferating cells called neoblasts that includes pluripotent stem cells. Planarian stem cells produce every cell type in the organism. The selection of cellular identity is driven by transcription factors (TF), which activate cell-type specific gene expression programs. Many lineage specifying TFs are known in planarians, yet their direct targets, which are likely essential for cellular differentiation and lineage production, are unknown. The lack of in vivo tools in planarians for analyzing TF binding sites, has further limited progress in TF target discovery. Here, we optimized an in vitro-based DNAseq approach for identifying potential TF binding sites (DAP-seq), previously developed for use in plants (Bartlett et al. 2017), and applied in pilot experiments to several TFs. This in vitro-based approach allows an antibody-independent discovery of potential binding sites, which together with a computational framework that integrates available ATAC-seq and scRNAseq data, facilitates prediction of potentially bona fide direct regulation in planarian stem cell lineage specification.