Injury-induced Neuregulin-ErbB signaling from muscle mobilizes stem cells for whole-body regeneration in Acoels

The activation of progenitor cells near wound sites is a common feature of regeneration across species, but the conserved signaling mechanisms responsible for this step in whole-body regeneration are still incompletely understood. The acoel Hofstenia miamia undergoes whole-body regeneration using Piwi+ pluripotent adult stem cells (neoblasts) that accumulate at amputation sites early in the regeneration process. The EGFR signaling pathway has broad roles in controlling proliferation, migration, differentiation, and cell survival across metazoans. Using a candidate RNAi screening approach, we identify the Hofstenia EGFR erbB4-2 and Neuregulin nrg-1 genes as essential for blastema formation. Structure prediction of NRG-1 and ERBB4-2 proteins supports the likelihood of these factors interacting directly. After amputation injuries, nrg-1 expression is induced in body-wall muscle cells at the wound site by 6 hours and localizes to the tip of the outgrowing blastema over the next several days, while erbB4-2 is broadly expressed, including in muscle and neoblasts. Under nrg-1(RNAi) and erbB4-2(RNAi) conditions that impair blastema formation, animals still undergo the earliest responses to injury to activate expression of the Early Growth Response transcription factor egr, indicating a crucial role for EGFR signaling downstream of initial wound activation. nrg-1(RNAi) and erbB4-2(RNAi) animals possess Piwi+ and H3P+ mitotic neoblasts which hyperproliferate normally after amputation, but these cells fail to accumulate at the wound site. Therefore, muscle provides a source for Neuregulin-ErbB signaling necessary for the mobilization of proliferative progenitors to enable blastema outgrowth for whole-body regeneration in Hofstenia. These results indicate a shared functional requirement for muscle signaling to enable regeneration between planarians and acoels across 550 million years of evolution.