Chromatin Accessibility Analysis Reveals Distinct Functions for HDAC and EZH2 Activities in Early Regeneration

Xenopus tropicalis tadpoles have the capability to scarlessly regenerate appendages including the limb and tail. Following injury, complex genetic processes must be activated and inactivated with high spatial and temporal resolution to result in a properly patterned appendage. Functional studies have established that histone modifying enzymes that act to close chromatin are required for regeneration, but the genomic regions sensitive to these activities are not established. Here we show that early inhibition of HDAC or Ezh2 activity results in incomplete regeneration. To identify the impact that each of these perturbations has on chromatin accessibility, we applied an assay for transposase accessible chromatin (ATAC-seq) to HDAC or Ezh2 inhibited regenerating tadpoles. We find that neither perturbation results in a global increase in chromatin accessibility, but that both inhibitors have targeted effects on chromatin accessibility and gene expression. Upon HDAC inhibition, promoter regions and regulatory regions neighboring genes associated with neuronal regeneration are precociously accessible, whereas following EZH2 inhibition we find gene bodies and regulatory regions associated with regulation of the immune response and apoptosis are preferentially accessible. Together this suggests distinct roles for EZH2 activity and HDAC activity in appendage regeneration. Overall design: ATAC-seq timecourse analysis of chromatin dynamics of HDAC inhibited or EZH2 inhibited regenerated tadpole tails and their vehicle controls. Samples in duplicate.