| Summary: | Whole-body regeneration requires an organism to produce all missing cell types. The planarian flatworm Schmidtea mediterranea contains an estimated 150 distinct cell types, which can all be regenerated after injury. Cell-type production in planarians is regulated by the expression of fate-specific transcriptions factors (FSTFs) in dividing cells called neoblasts. However, it remains unclear whether all ~150 fate choices are made in neoblasts, or whether additional mechanisms for generating cell type diversity are used. We used single-cell RNA-sequencing of S/G2/M neoblasts and early post-mitotic cells to identify new neoblasts states corresponding to mature cell types, along with evidence that some cell type diversity is generated in post-mitotic neoblast progeny. We find that strategies for generating cell type diversity differ across tissue types. Furthermore, by annotating a complete set of predicted planarian transcription factor-encoding genes (the planarian TFome), we identify novel FSTFs and additional neoblast states. These data indicate that different strategies for generating cell type diversity exist across tissues, including fate choice outside of neoblasts.
Understanding how cells choose their fates is an important challenge in development and regeneration. In regenerative planarians, fate choices are primarily made in neoblasts through the expression of FSTFs. But how individual neoblasts within the intact animal choose which fate to adopt and what role their spatial position plays remains poorly understood. Using fluorescent in situ hybridizations, we find that neoblast fate choice is spatially heterogenous and not tightly regulated by position. We find that specialized neoblasts of different classes are commonly found neighboring specialized neoblasts of other classes, creating an intermingled, salt-and-pepper distribution in the animal. Furthermore, we develop the in situ RNA-sequencing technique, STARmap, for use in planarians, and utilize it to study the in vivo spatial distribution of specialized neoblasts through spatial cell-type mapping. We identify the gene nlg-7 as a candidate regulator of the migratory targeting that spatially heterogenous neoblasts progenitors must undergo to maintain and regenerate tissues, given their heterogenous fate specification pattern. These results indicate that fate specification in neoblasts is not precisely regulated by position, and therefore, migratory targeting of progenitors is a major driver of tissue formation.
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