Periodic Reporting for period 3 - PLANMod (Regulation of extreme plasticity in planarian stem cells by mRNA modifications)
Período documentado: 2022-11-01 hasta 2024-04-30
Regeneration, the growth of new tissues following injuries, is widespread in the animal kingdom. However, most mammals have a very limited ability to regenerate organs. Planarians, freshwater flatworms, can regrow any missing body part, including an entire brain. Planarians use a large population of stem cells for regeneration. PLANMod studies mechanisms that regulate planarian stem cells, and facilitate their capacity to divide and produce tissues on demand. Similar mechanisms could be widespread in other organisms. Our project is focused on the function of biochemical modifications to planarian RNAs, and their association to production of new cells. The particular biochemical RNA modification that we study, N6-methyladenosine (m6A), is abundant in animals, plants, and unicellular eukaryotes, such as yeast. A similar cellular machinery is found in planarians, and it is required for installing the m6A modification on planarian RNAs. We found that, in planarians, this cellular machinery is highly expressed in stem cells, compared to other cell types. This suggested that m6A is required for stem cell functions. Our analysis of m6A functions demonstrated that m6A is required for production of certain cell types. Moreover, it is essential for the viability of animals and for the formation of certain tissues. The overall objective of this project is to discover the functions of m6A in planarian stem cells, their contribution to stem cell physiology, and how m6A contributes to the ability of planarian stem cells to maintain and regenerate organs.
We systematically analyzed the function of every known gene involved in the m6A pathway and examined its roles in the maintenance of tissues, regeneration, and stem cell regulation. We established new experimental and computational methodology for analyzing m6A in planarians. The work that we performed resulted in the mapping of over 7,000 highly methylated regions in planarian RNAs, and analysis of the effect of inhibition of the m6A pathway on these sites. Our analysis demonstrated that m6A is essential for the production of certain tissues by stem cells, but not for stem cell viability. Therefore, the most dramatic outcome of the inhibition of the m6A pathway – death – is a consequence of defects in tissue production, but not from lack of stem cells. These highlights similarities of early mammalian development and regulation of adult stem cells in planarians. Our analysis further demonstrated that defects in tissue production have two major effects. First, there is lack of formation of intestine tissue. Therefore, the intestine is depleted over time and eventually fails. A second outcome is that abnormal cells populate the animals. These cells are not detectable normally, and they emerge when the m6A pathway is inactive. Analysis of these cells showed that they molecularly resemble progenitors of neurons, although it is unlikely that they function as such. Systematic computational efforts unraveled that similar cells appear following inactivation of cellular complexes that are crucial for regulating the structure and state of DNA packaging, suggesting that there is a direct or indirect association between RNA modifications and changes to the packaging of DNA.
Our results so far have demonstrated that m6A is required particularly for stem cell differentiation, and that inactivation of the m6A pathway produces an imbalance in the formation of certain tissues compared to others. This indicates that a small biochemical change to the RNA molecule can have a dramatic role in the regulation of stem cell biology, and suggests how it precisely tunes the formation of certain tissues. Our results, so far, have characterized an association of m6A with changes to complexes that are involved in packaging and remodeling of the DNA structure. The link, which we do not fully understand, connects two seemingly distinct processes that take place at different times and places.