Mechanisms and consequences of lipid accumulations in necroptosis

Project Summary Lipids contribute to numerous cellular processes. The overarching goal of my laboratory is to study lipid regula- tion and function in pathophysiological processes and identify targetable lipid-related pathways to better human health. Necroptosis is a form of regulated cell death that contributes to the inflammatory components of many diseases, including neurodegenerative and cardiovascular diseases and chronic inflammation. A hallmark of necroptotic activity is the permeabilization of cellular membranes and the release of inflammatory molecules. My laboratory has pioneered the efforts to study the involvement of lipids in necroptosis, leveraging untargeted lip- idomics integrated with functional characterizations using pharmacological and genetic perturbations. We have demonstrated that de novo lipid production increases via activated sterol regulatory binding element proteins during necroptosis and results the accumulation of lipids that exacerbate membrane permeabilization. We have thus far focused on very-long-chain fatty acid (VLCFA) accumulations in necroptosis and demonstrated that VLCFA accumulation regulates the function of a key signaling protein of necroptosis, Mixed Lineage Kinase Like Protein (MLKL), responsible for the permeabilization of the plasma membrane. Specifically, we showed that MLKL undergoes S-acylation by VLCFAs, which increases its membrane binding and consequent plasma mem- brane permeabilization in necroptosis. In the next five years, we will pursue three main research areas to answer key questions toward better understanding the mechanism of activated lipid production in necroptosis and to characterize the acylation of MLKL by VLCFAs in necroptosis. What are lipid modulators of necroptosis? In Research Area 1, we will identify lipids that contribute to mem- brane permeabilization in necroptosis. We will integrate state-of-the-art lipidomics approaches developed in my lab with perturbation studies for functional investigations. This will reveal lipid groups linked to membrane per- meabilization in necroptosis and lipid-related enzymes to reduce the inflammatory activity of necroptosis. What causes lipid accumulation in necroptosis? In Research Area 2, we will investigate cellular and molec- ular mechanisms that activate lipid production in necroptosis, integrating proteomics and imaging-based ap- proaches. These results will advance our fundamental knowledge of lipid regulation in necroptosis. How does S-acylation regulate MLKL functioning in necroptosis? In Research Area 3, we will focus on the acylation of MLKL by VLCFAs, study the effect of acylation on MLKL fate during necroptosis and identify S- acyltransferases that can incorporate these unusual fatty acids into MLKL. Impact. Answering these questions will provide a fundamental understanding of the regulation of necroptosis by lipids and pave the way for developing lipid-centric therapeutic strategies targeting necroptosis-mediated enter- opathies.