Identification of rare, transient post-mitotic cell states that are induced by injury and required for whole-body regeneration in Schmidtea mediterranea

Regeneration requires the coordination of stem cells, their progeny and distant differentiated tissues. Here, we present a comprehensive atlas of whole-body regeneration in Schmidtea mediterranea and identify wound-induced cell states. An analysis of 299,998 single-cell transcriptomes captured from regeneration-competent and regeneration-incompetent fragments identified transient regeneration-activated cell states (TRACS) in the muscle, epidermis and intestine. TRACS were independent of stem cell division with distinct spatiotemporal distributions, and RNAi depletion of TRACS-enriched genes produced regeneration defects. Muscle expression of notum, follistatin, evi/wls, glypican-1 and junctophilin-1 was required for tissue polarity. Epidermal expression of agat-1/2/3, cyp3142a1, zfhx3 and atp1a1 was important for stem cell proliferation. Finally, expression of spectrinß and atp12a in intestinal basal cells, and lrrk2, cathepsinB, myosin1e, polybromo-1 and talin-1 in intestinal enterocytes regulated stem cell proliferation and tissue remodelling, respectively. Our results identify cell types and molecules that are important for regeneration, indicating that regenerative ability can emerge from coordinated transcriptional plasticity across all three germ layers. Overall design: For each time course, 100-150 biopsies were dissociated and fixed from each sample, then ~500,000 cells from each sample were sorted, barcoded and pooled into 8 sub-libraries for sequencing. Each sub-library contained 15,000-20,000 individual cells pooled from all timepoint within the timecourse. Each timecourse was split into 8 sub-libraries, resulting in a total of 24 sub-libraries across the three timecourses. The read barcode structure is as follows for the demultiplexed fastq files: UMI: 1-10bp CellBarcode: 11-34bp. Note: SRA fixed an internal database issue about the samples on September 22, 2022. Please redownload the data if needed.