I recently started a postdoctoral position in the Adler lab at Cornell University where we study how stem cells respond to injuries and how animals can regenerate lost organs.
Understanding the basic principles of organ regeneration will allow us to treat ailments such as degenerative diseases, acute injuries, and organ failure. Pluripotent stem cells can undergo cell division to produce a variety of cell types to replace damaged tissues. Therefore, stem cells hold enormous potential for the treatment of human disease, but this requires understanding how stem cells transition into specific tissues and functional organs. Planarian flatworms possess abundant stem cells allowing them to rapidly regenerate virtually any body part after amputation, making them an exceptional system to study these phenomena. While planarian regeneration has typically been studied by amputation of large portions of the animal, the Adler lab has developed a unique method to selectively amputate a single planarian organ, the pharynx. Pharynx regeneration requires pluripotent stems cells to differentiate into distinct cell types and integrate into pre-existing tissues to form a functional organ, making it an excellent model for understanding organ regeneration. However, how stem cells are triggered to initiate regeneration and coordinate the production of only the appropriate tissues types during pharynx regeneration are not fully understood.
The forkhead transcription factor, FoxA, is specifically required for pharynx regeneration but not for other organs. FoxA is expressed in a subset of stem cells in close proximity to the intact pharynx during homeostasis, but is also significantly upregulated in stem cells after amputation of the pharynx. This suggests that pre-existing stems cells must recognize the absence of the pharynx and subsequently initiate expression of organ progenitor genes like FoxA. Therefore, I hypothesize that amputation of the pharynx activates FoxA expression in stem cells to drive pharyngeal regeneration and cell fate specification, a process which is typically inhibited by the presence of the intact organ. Exploring this hypothesis will help develop an understanding of how stem cells are regulated during regeneration and lend insight into how to utilize stems cells for therapeutics for human tissue and organ loss.
During my postdoctoral research position, I plan to address the following aims:
Aim 1: Identify the mechanism by which stem cells are activated to regenerate the pharynx
Our current hypothesis is that the presence of the intact pharynx inhibits expression of FoxA in stem cells in order to retain tissue homeostasis. Thus, amputation of the pharynx allows for increased expression of FoxA within stem cells in order to activate the regeneration program. We will verify this by identifying molecules responsible for the inhibitory and activating effects of the pharynx through mass spectrometry and mining of established pharynx enriched and stem cell transcriptomes. We will then test their role in regulating FoxA expression. Knockdown of a potential inhibitor should lead to increased FoxA expression, even when the pharynx is present. Alternatively, knockdown of potential receptors that activate FoxA expression should block the expected increase in FoxA expression after pharynx amputation.
Aim 2: Characterize the function of FoxA in stem cells during pharynx regeneration
FoxA mRNA is expressed in multiple tissue types as pharynx regeneration progresses, suggesting that FoxA activation in stem cells is an early step toward differentiation of several pharynx-specific cell types. In order to determine how FoxA is specifically involved in pharynx regeneration we will identify potential indirect and direct downstream targets using RNA-Seq and ChIP-seq. We will also screen for further upstream effectors by RNAi knockdown, with the expectation that those genes activated earliest will cause the most profound defects in pharynx regeneration, while those activated later may recapitulate subsets of the FoxA phenotype.
Aim 3: Determine the origin of FoxA+ stem cells
We have shown that amputation of the the pharynx causes an upregulation of FoxA in stem cells surrounding the resulting wound, but the origin of these FoxA expressing cells is unknown. Amputation could stimulate local cell proliferation to produce FoxA expressing cells, migration of FoxA+ cells to the wound or direct transcriptional activation of FoxA in existing stem cells. Using methods to inhibit cell division, transplantation assays and qPCR, will elucidate to what degree each of these events is involved in pharynx regeneration.
Through the above specific aims, we will gain an understanding of how stem cells are triggered to initiate regeneration and coordinate the production of the appropriate tissues types during pharynx regeneration in planaria. This experience will give me the appropriate training needed to develop my skills using planaria as a model to study regeneration, a course of research I plan on continuing in my future career as an independent scientist. Knowledge of how stem cells are regulated to regenerate only the appropriate tissues will lend insight into how to utilize stems cells for therapeutics in treating human disease.