Identification of pluripotent stem cells and characterization of glia in the planarian Schmidtea mediterranea

Abstract

Given their regenerative capacity, the planarian Schmidtea mediterranea has emerged as a model system for the study of stem cell biology, tissue specification, and axis formation. Many aspects of the regenerative machinery have yet to be characterized. Although it is known that neoblasts, the population of all proliferative cells in the adult planarian, are the source of new tissue during regeneration, it is unknown whether neoblasts consist of multiple subpopulations of lineage-restricted multipotent stem cells or if there exists a pluripotent stem cell type. We developed two methods for performing clonal analysis to determine the potential of neoblasts: sublethal irradiation and single-cell transplantation. Colonies consisting of both self-renewed neoblasts expressing stem cell markers and differentiated cells expressing specialized tissue markers from single cells that we have termed clonogenic neoblasts. These cells are capable of generating all differentiated cell types in the adult animal and restoring regeneration in hosts where endogenous neoblasts have been ablated. These findings provide insight into the overall process of regeneration and the regulation of pluripotent adult stem cells during regeneration. Regeneration and homeostasis, the gradual turnover and replacement of cells in the adult planarian, both require the formation of new cells and signaling pathways to control their specification and function. The Hedgehog signaling pathway has been implicated in anterior-posterior polarity specification, but no role in planarian nervous system regeneration has been described despite that hedgehog is expressed in neurons in the brain. Although Hedgehog signaling is critical for central nervous system development in vertebrates and Drosophila, our data indicate that it is not involved in patterning the planarian brain. Instead, Hedgehog signaling regulates gene expression in a cell type identified as planarian glia from its localization to the axon-rich neuropil, expression of planarian orthologs of astrocyte genes, and branching morphology in close association with neurons. Evidence of both the existence of planarian glia and their regulation by Hedgehog signaling offers the opportunity to dissect glial cell biology in a highly regenerative model organism and understand the evolution of the cell type.