Periodic Reporting for period 2 - FlyGutHomeostasis (Identification of paracrine and systemic signals controlling adult stem cell activity and organ homeostasis) Reporting period: 2020-08-01 to 2022-01-31 Summary of the context and overall objectives of the project Adult tissues with high turnover rates, such the intestine, depend on stem cells (SCs) to provide a continuous source of differentiated cells to maintain tissue homeostasis. To ensure optimal tissue homeostasis and physiology, adult stem cells must coordinate their own maintenance with the generation of differentiated cell types in a temporally and spatially controlled manner. Due to its remarkable self-renewing capacity, the fly gut has recently become a prime paradigm for studying stem-cell function during adult tissue homeostasis. This capacity for self-renewal relays on the proliferative activity of the intestinal stem cells (ISC), which is tightly coupled with cell loss to maintain intestinal homeostasis. Deregulation of ISC proliferation affects gut integrity, which in turn can trigger a number of chronic inflammatory diseases including inflammatory bowel disease and colorectal cancer. ISC proliferation is controlled by multiple local and systemic signals released from the ISC niche (enterocytes (ECs), enteroendocrine (EE) cells, enteroblasts (EBs), and visceral muscles (VMs)) and non-gastrointestinal organs. Identifying niche-derived and systemic signals controlling gut epithelial turnover therefore represents an important step towards treating gut inflammatory diseases. This project takes advantage of the genetic amenability of the fruit fly to identify ISC niche-derived and systemic signals that control ISC activity and gut homeostasis. To identify local and systemic signals controlling intestinal homeostasis in steady-state and challenged conditions, we use RNAis to knock down all genes encoding secreted peptides specifically in ECs, EEs, or VMs and all genes encoding transmembrane and membrane-associated proteins in the VMs. Using this approach, we aim at identifying novel intra- and inter-organ circuitries allowing communication between the gut and other organs to promote gut homeostasis and organismal health. In addition, the systematic knockdown of secreted peptides from the ISC niche could identify gut-derived signals that couple changes in environmental inputs, such as nutrient availability, with systemic changes in feeding behavior, energy balance, and metabolism. Since large-scale approaches are not feasible in vertebrate models, the signals identified in the above screens could potentially reveal novel couplings contributing to mammalian gastrointestinal homeostasis and disease. Finally, the role of TNF signaling in intestinal homeostasis, regenerative growth and disease will be addressed. In particular, the contributions of ligand-independent activation of TNFR signaling in these contexts will be studied. Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far In the first half of the project period, we have completed two large genetic screens aiming at identifying niche-derived signals controlling ISC activity and gut physiology in steady-state and challenged conditions (Aim I and II). These screens have identified a number of exciting candidate genes, of which we have prioritized to work on 9. Several of the projects generated from these screens are well advanced and will be submitted within the next year. I am currently gearing up to start the third and final screen - the “receptor screen” (Aim III). I have hired two postdocs and one PhD student (starting first of January), all of which will participate in this screen. I expect the “receptor screen” to be completed around august 2022. Concerning the role of TNFR signaling in gut homeostasis (aim IV), I have a talented Postdoc working on this part of the proposal. This project is well advanced and I hope to submit a manuscript within 6 months. We recently published a paper related to this part of the project (palmerini et al., Nat. Commun. (2021)). In summary, I am progressing as planned on all four objected. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) The genetic screens required a certain investment of time and man power, but were successful in generating many promising projects that will serve as a foundation for my future work in the coming years. Many of the candidate genes identified in the screens are well-conserved in humans, but with no previous identified role in gut homeostasis. The knowledge generated by this project will therefore be of broad interest to field of stem cell physiology and pathology. As mentioned above, I expect to submit the first manuscripts on the screen projects within the next year and another manuscript on the TNF part (aim IV) within the next 6 months. Overview of projects in the Team including Aim I-IV
