Abstract LB186: Rapid photoinhibition of the autophagy pathway

Abstract

Abstract Autophagy is a highly regulated and conserved cellular recycling process that is activated under stress conditions. Although cancer cells are known to rely on autophagy for survival, recent studies indicate that when the nutrient recycling process is blocked, a few rare cancer cells still can circumvent autophagy inhibition and maintain their aggressive growth rate. To understand the autophagy inhibition adaptation mechanism, we have developed a novel optogenetic autophagy inhibition tool (ASAP) that can rapidly and reversibly block the autophagy pathway in response to blue light. Our preliminary data shows ASAP releases a mutant autophagy inhibitory protein from the plasma membrane within 5 minutes of blue light exposure, blocks autophagosome-lysosome fusion, and induces LC3-II protein accumulation. In comparison, bafilomycin, a known autophagosome-lysosome fusion inhibitor, takes 4-hours to reproduce similar inhibitory effects as ASAP. Additionally, unlike bafilomycin, ASAP-induced autophagy inhibition is reversible within 2-6 hours. As light-induced autophagy inhibition has not been previously tested, our system brings a novelty to the conventional genetic and pharmacological approaches of autophagy inhibition. We will utilize the novel ASAP tool to understand the rapid changes in cellular proteomics and signaling events that dictate autophagy inhibition adaptation in cancer cells. We will perform tandem mass tag (TMT) based total proteomics and phospho-proteomics screen to visualize the temporal kinetics of protein turnover rates and changes in cell-signaling pathways following rapid autophagy inhibition. Additionally, our optogenetic tool will be applied in vivo in C. elegans to rapidly block autophagy and answer key questions of the spatial and temporal roles of autophagy in aging. Citation Format: Payel Mondal, Amanda Cyril, Christina Towers. Rapid photoinhibition of the autophagy pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB186.