NCE (Year 6)
The human central nervous system (CNS) has very limited capacity for renewing cells and is not capable of compensating for cells lost after injury or disease. Studies of organisms that can regenerate the CNS have excellent potential to improve our understanding of how adult stem cells can be harnessed for therapeutic purposes. The goal of this work was to establish planarians as models to gain insights into mechanisms regulating stem cell differentiation. We were specifically interested in understanding how the remarkable ability of these animals to regenerate the central nervous system after injury or amputation. We capitalized on the molecular tools available to study planarian regeneration to identify and functionally characterize genes with key roles in nervous system repair, patterning and function.
To advance our knowledge of how regeneration and patterning is achieved in the CNS, we examined in detail the function of the transcription factors COE. COE proteins have been recently implicated in CNS diseases in adult organisms. Using RNA interference, we found that COE is essential for regeneration and to maintain nervous system architecture in planarians. Using genomic techniques, we examined gene expression changes following inhibition of COE expression and uncovered conserved genes required for CNS regeneration. Our studies suggest COE could be explored as a factor for reprogramming somatic cells into neuronal cell types. In addition to completing the analysis of COE, we also completed the generation and characterization of monoclonal antibodies that label specific planarian tissues. These antibodies were deposited to the NIH-funded Developmental Studies Hybridoma Bank and are readily accessible to the scientific community. Finally, during the five-year funding period we completed high-throughput gene expression studies (microarray and RNA-seq) using samples obtained from tissue replaced in the early phases of regeneration of the planarian head after amputation. We performed gene inhibition studies for hundreds of genes chosen from the microarray list based on expression pattern or homology. We identified 25 genes that were necessary for stem cell maintenance, general regenerative capability, or that caused tissue-specific defects upon knockdown. We also found that a homolog of the nuclear transport factor importin- plays a role in stem cell function and tissue patterning, suggesting that controlled nuclear import of proteins is important for regeneration. This information has the potential to contribute to our ability to manipulate pluripotent stem cells to divide and acquire neuronal fates in vivo.