Ferroptosis is a form of regulated cell death that generates reactive oxygen species (ROS) in an iron- dependent fashion. The most important of the ROS are lipid peroxides, which self-propogate along the plasma membrane and result in the accumulation of oxidatively damaged lipids. These lipid peroxides lead to a loss of membrane integrity, leakage of cellular and subcellular content and, ultimately, cell death. The main protective mechanism against ferroptosis relies on a gluathione-dependent peroxidase (GPX4); this enzyme converts lipid peroxides to lipid alcohols, thus blocking cell death. Defects in this protective pathway have been implicated in the etiology of a number of neurodegenerative disorders. Conversely, abrogation of GPX4 has sensitized drug-resistant cancer cells to chemotherapeutics. Unfortunately, our understanding of the mechanisms underlying ferroptosis remain limited, but one possibility of ferroptotic regulation may be found in the enzyme acyl-CoA synthetase long-chain 4 (ACSL4). This enzyme converts free long-chain fatty acids into fatty acyl-CoA esters, the building blocks of the phospholipids that are fated to propagate ferroptosis. Knockout of ACSL4 has been shown to stave off ferroptosis in multiple in vivo and in vitro models. Interestingly, the ACSL4 gene can produce two isoenzymes via alternative splicing; these isoforms differ in length and in localization, where the long form co-localizes to lipid droplets (LDs) and the short form co- localizes to the plasma membrane (PM). We hypothesize that this differential localization modulates ferroptotic sensitivity as the sequestration of lipid peroxides to the LDs may prevent death by ferroptosis. In order to address this question, cell lines will be created in which endogenous ACSL4 has been eliminated and replaced with a different expression construct for the two major splice forms of ACSL4. Moreover, these splice forms will be targeted to various subcellular localizations to investigate the role of these compartments in the execution of ferroptotic cell death. Additionally, since ACSL4 is integral to lipid metabolism, we will investigate the composition of fatty acids via Raman spectroscopy and characterize fatty acid flux through lipidomics. This project addresses an important step in the regulation of ferroptosis, namely how sequestration of damaged lipids away from critical membrane regions may negatively affect ferroptotic cell death. The enzyme that governs this sequestration, ACSL4, thus has great pharmacologic potential, not only as a drug target for novel anti-cancer drugs, but also in age-related neurological diseases.

Public Health Relevance

Ferroptosis is a newly discovered form of cell death that is characterized by a self-propagating cycle of toxic lipid compounds. This mechanism is implicated in multiple disease models, notably neurodegenerative diseases, and is also known for sensitizing drug-resistant cancer cells. My proposed research will provide critical information on how ferroptosis is regulated, thereby identifying new factors that can be targeted by therapeutic strategies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Predoctoral Individual National Research Service Award (F31)
Project #
1F31GM134645-01
Application #
9836508
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Brown, Patrick
Project Start
2019-07-09
Project End
2022-07-08
Budget Start
2019-07-09
Budget End
2020-07-08
Support Year
1
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of California Berkeley
Department
Nutrition
Type
Earth Sciences/Resources
DUNS #
124726725
City
Berkeley
State
CA
Country
United States
Zip Code
94704