Identifying and characterizing a planarian ortholog of FOXO transcription factors in the regulation of stem cell function and tissue regeneration

Abstract

Tissue/organ regenerative capacity varies among different species. The more complex organisms (such as mammals) tend to have a weaker regenerating ability while less complex organisms tend to have a better regenerating ability (such as Hydra and planarians). Tissue/organ regenerative capacity is believed to be governed by the potency of stem cells that are responsible for regeneration. The potencies of stem cells in regenerative species are thought to be maintained during development. The pluripotent adult stem cells can differentiate into any cell type in regenerative organisms while the multipotent adult stem cells in less regenerative organisms can only differentiate into several cell types, resulting in limited regenerative capacity. The detailed mechanisms of regulation and maintenance of stem cells remain unclear. Planarians have a superb regenerative ability and a high population of pluripotent stem cells and they were used as the animal model for my study. From the result of a transcriptomic analysis, forkhead box (FOX) transcription factors were found to be highly expressed in human embryonic stem cells, mouse embryonic stem cells and planarian adult stem cells. FOXO family members were recently shown to regulate the pluripotency genes (OCT4 and SOX2) and they are known to be important in stem cell maintenance required for longevity. Investigating the role of FOXO ortholog(s) in the regulation of stem cell function during planarian regeneration would be very interesting. One FOXO ortholog was identified from the planarian EST databases based on phylogenetic analysis. Using in situ hybridisation analysis, I demonstrated the ubiquitous expression of Smed FOXO mRNA which is in accordance with the expression pattern of FOXO in humans. Suppression of FOXO expression by RNA interference (RNAi) did not show effect on stem cell migration and regeneration when planarians were amputated but a forked tail phenotype was observed, accompanied by increases in the rate of apoptosis shown by TUNEL assay. The forked tail phenotype suggests an involvement of FOXO in the patterning of tail regeneration. Upon amputation, a ROS burst generated at the wound area is believed to be the signal that induces tissue regeneration. ROS is FOXO is an upstream regulator for ROS metabolism in humans and apoptosis can be induced by ROS (oxidative stress). The knockdown of FOXO can result in an inhibition of ROS metabolism and therefore increases in apoptotic rate. This study has provided interesting insight in role of FOXO during planarian regeneration. Further functional analysis of FOXO in other processes involved in regeneration is required to define clearly the role of FOXO in stem cell maintenance and regulation during planarian regeneration.