Comparative multi-omic analysis reveals conserved and derived mechanisms of fin and limb regeneration

Comparative studies of vertebrate appendages offer a powerful framework for uncovering shared components of an ancestral regeneration toolkit. Here, we employed a multi-omics comparative approach leveraging the regenerative capacity of the axolotl, zebrafish, and Polypterus senegalus, a fish capable of full fin regeneration. We identified conserved markers of proximal and distal blastema territories, shared activation of DNA damage repair, hif1a-mediated hypoxia response, and sequential activation of pro- and anti-inflammatory program. Apical epithelial ridge markers were expressed in both the wound epidermis and distal mesenchyme during limb and fin regeneration. Notably, hif4a-expressing erythrocytes were uniquely associated with proximal limb and fin amputations but not fin rays, while epidermal myoglobin expression was upregulated only in Polypterus and zebrafish fins. Genome-wide chromatin profiling identified candidate regeneration-responsive elements and a conserved enrichment for AP-1 transcription factor binding. Together, these finding identify shared and derived mechanisms of limb and fin regeneration. Overall design: snRNA-seq profiling of Polypterus senegalus regenerating fins. Single nucleus suspensions were prepared from uninjured and regenerating pectoral fins at 1, 3 and 7 dpa (days post-amputation). Regenerating fins were either from fish treated with DMSO (control) or 2.5 uM rapamycin (mTOR inhibition). Nuclei were fixed and frozen using the Evercode Nuclei Fixation Kit v3 (#NF100, #NF200) from Parse Biosciences following the manufacturer's protocol. Fixed and frozen nuclei were then processed to generate snRNA-seq libraries following the combinatorial barcoding protocol from Parse Biosciences, using the Evercode WT v3 Kit (#WT100A-D, #WT200) as per manufacturer's instructions.