Turning off autophagy in a multicellular organism

Cells are survivors. In a living organism, cells need to be able to respond to changes in food supply. When organisms starve, certain organs respond by activating a cellular recycling pathway called autophagy. Autophagy is a process where cells “eat” their own damaged, unused or superfluous parts. Activation of autophagy provides energy and building blocks to sustain essential cellular functions and protect organisms against diseases like cancer and neurodegeneration. Importantly, these cells must also turn off autophagy when nutrient supplies are replenished, because unrestricted autophagy is harmful to cellular fitness. A lot is known about how autophagy is turned on, but surprisingly little is known about how autophagy is turned off, especially in the context of multicellular organisms. This project aims at understanding the mechanisms of how autophagy is turned off, using fruit flies. To reach our goal of understanding how autophagy is turned off, we will use gene silencing and monitor the effect when fly cells in culture are turning off autophagy, to find regulators of this process. Top hits from this will be further investigated in fly larvae. These larvae will have some cells where a candidate gene is silenced, and these cells will be surrounded by normal cells. These larvae will be monitored as they go through periods of starvation and feasting on fresh food. Particularly, the effect on fruit fly physiology will be investigated when autophagy is correctly or incorrectly turned off. Two postdoctoral fellows have now worked three years on the project. One of them has optimized the set-up to silence gene expression in fly cells in culture and study the effect when the cells are turning off autophagy. The other has established the methods to study the same genes in adipose tissue of fly larvae. In both systems we have established how quickly autophagy is normally terminated upon refeeding. We have also optimized the image analysis methods to extract information on the autophagy process from images of fly cells in culture and from images of different fly tissues upon starvation and refeeding. We have identified potential regulators of how autophagy is turned off and have partially elucidated the mechanisms of how these work. We are also investigating how these regulators affect physiology and a fly cancer model. In 2021 we published an article on how autophagy could be considered as a target for treatment in a form of brain cancer using a fruit fly model of this cancer type. In 2023 we published an article presenting a fly model for MLL-rearranged leukemia. This will be used to further study the effect of regulators of autophagy termination in this cancer model. In 2023 we also published an article presenting a data set of annotated images of Drosophila S2 cells in different stages of autophagy. This can be used by the field to further develop analysis methods, such as by machine learning. Findings from the project have been presented at international conferences in 2021 (Nordic Autophagy Society meeting in Tromsø, Norway), 2022 (EMBO Autophagy conference in Eger, Hungary) and 2023 (European Drosophila Research Conference in Lyon, France). Based on findings from this project, the project leader successfully applied for an ERC Starting Grant which started in 2023. This is a prestigious grant from the European Research Council. The implications of their findings in the years to come will be relevant not just for understanding this important process in fruit flies, but for this survival process in all living organisms.

Cells in a living organism need to be able to readily respond to changes in nutritional status. Upon starvation, certain tissues respond by activating catabolic processes, such as autophagy, to provide energy and building blocks to sustain essential cellular functions. Importantly, these cells must also turn off autophagy when nutrient supplies are replenished, because unrestricted autophagy is harmful to cellular fitness. Surprisingly little is known about how autophagy is terminated, especially in the context of a multicellular organism. This project aims at uncovering novel mechanisms of autophagy termination, using fruit flies as a model. To reach this goal, my team will first identify novel regulators of autophagy termination among transcription factors and signaling pathway components through a RNAi-mediated high-content imaging-based screen in fly cells in culture. Top candidates from this screen will be further evaluated in vivo in fly larvae using clonal experiments where wild-type and mutant cells are juxtaposed within the same tissue. To obtain a detailed mechanistic understanding of one or two identified novel regulators of autophagy termination, my team will use epistasis analysis, evaluate their relationship to conserved signaling pathways, study morphological changes in autophagy-related membranes and investigate their subcellular localization. Finally, we will analyze the impact of defects in autophagy termination on developmental timing of fly larvae, fitness of adult flies and a cancer cachexia phenotype. I envisage that the proposed screening strategy and follow-up experiments will lead to fundamental breakthroughs in our understanding of autophagy termination. In addition, this “Young Research Talents” project will allow my transition to a project group leader and propel my career as an independent researcher.