br Conflict of interest br
Conflict of interest
Acknowledgments The research was funded by the Major Programs of Pharmaceutical research project of Jiangsu Province (Q2018044).
Introduction Lipids are important regulators of cell death. In mammalian cells, both apoptotic and non-apoptotic cell death pathways can be induced, modulated, or suppressed by different lipids, including free fatty acids and more complex lipids (Das, 1991, Green et al., 2014, Magtanong et al., 2016). For example, apoptosis can be triggered by increasing the ratio of intracellular saturated fatty acids (SFAs) to monounsaturated fatty acids (MUFAs), a process termed lipotoxicity (Garbarino and Sturley, 2009, Listenberger et al., 2003, Young et al., 2013). Necroptosis, a form of non-apoptotic cell death, is associated with increased very long chain fatty eletriptan receptor and ceramide levels that contribute to the initiation of cell death (Parisi et al., 2017). Ferroptosis, a distinct form of non-apoptotic cell death, is characterized by the iron-dependent accumulation of toxic lipid reactive oxygen species (ROS) (Dixon et al., 2012). The initiation and execution of cell death is therefore intimately linked to lipid metabolism. Ferroptosis is a non-apoptotic form of cell death that can be selectively activated in certain cancer cells, and that mediates a number of pathological cell death events in vivo (Stockwell et al., 2017). Unlike other forms of cell death, ferroptosis is not thought to require a specific protein effector, like a pore-forming protein. Rather, lipid oxidation and the resultant membrane damage are thought to be sufficient for the execution of this process (Stockwell et al., 2017). During ferroptosis, toxic lipid ROS arise from the reaction between iron and lipid peroxides, which are themselves generated by the oxidation of polyunsaturated fatty acid (PUFA)-containing phospholipids (PUFA-PLs) (Stockwell et al., 2017). Normally, lipid peroxides are reduced to non-reactive (i.e., non-toxic) lipid alcohols by the reduced glutathione (GSH)-dependent lipid hydroperoxidase glutathione peroxidase 4 (GPX4). However, GPX4 is inactivated under ferroptosis-inducing conditions, leading to lipid peroxide accumulation (Stockwell et al., 2017). Oxidizable PUFA-PLs are found in membranes throughout the cell, and mitochondria, lysosomes, and endoplasmic reticulum are all sites of lipid ROS accumulation during ferroptosis (Gaschler et al., 2018, Kagan et al., 2017, Krainz et al., 2016, Torii et al., 2016). Whether lipid oxidation at one or more of these sites is necessary for ferroptosis is presently unclear. The execution of ferroptosis can be prevented by deletion of the acyl-coenzyme A (CoA) synthetase long-chain family member 4 (ACSL4), one of five human ACSL enzymes (ACSL1, 3–6; ACSL2 does not exist) (Dixon et al., 2015, Doll et al., 2017, Kagan et al., 2017, Mashek et al., 2004). ACSL enzymes activate free fatty acids to fatty acyl-CoAs, which can then be incorporated into glycerophospholipids (Grevengoed et al., 2014). ACSL4 has a marked preference for activating PUFAs (Kang et al., 1997); therefore, deletion of ACSL4 prevents PUFAs from being incorporated into membrane PLs where they would become oxidized following GPX4 inactivation. Endogenous mechanisms that may act to inhibit ferroptosis by opposing PUFA activation and incorporation into membrane PLs have not been described. Exogenous metabolites including lipids are potent modulators of cell function and fate (Cantor et al., 2017, Yao et al., 2016). Given the links between lipid metabolism and cell death, we examined how exogenous fatty acids impacted cell sensitivity to both ferroptotic and non-ferroptotic lethal stimuli. Here we show that exogenous MUFAs potently suppress ferroptosis. Mechanistically, MUFAs inhibit the accumulation of lipid ROS specifically at the plasma membrane and displace PUFAs from this location in the cell. We find that ACSL3 is required for exogenous MUFAs to protect cells against ferroptosis, but not lipotoxicity induced by exogenous SFAs. ACSL3-dependent MUFA metabolism therefore emerges as key a regulator of ferroptotic cell death.