Hypoxic cell death in the form of stroke and myocardial infarction is the largest single cause of death in the United States. The fundamental molecular mechanisms of hypoxic cell death are incompletely understood. Studies in genetically tractable model organisms such as Caenorhabditis elegans and Drosophila have made major advances in the fields of cell death, hypoxia sensing, and adaptation. The proposed work will utilize the powerful genetic tools in C. elegans to explore manipulation of protein synthesis, trafficking, and degradation as a means to control hypoxic cell death.
Our specific aims are: (1) Define the mechanisms whereby aminoacyl-tRNA synthetases (aaRS's) and other translational machinery proteins control hypoxic injury. In a screen in C. elegans for mutations that produce hypoxia resistance, we isolated a missense mutation in the rrt- 1 gene, which encodes an arginyl tRNA synthetase, a fundamental protein required for translation.
This specific aim proposes a systematic study of translation factors as regulators of hypoxic sensitivity. Through these experiments, we should learn which translation factors are the best candidates to target for hypoxic protection. We will also learn what level of translational suppression is required for hypoxia resistance. Finally, we will learn whether the widely held assumption that translational suppression produces hypoxia resistance by reducing energy consumption is correct or if the mechanism of protection is more complex. (2) Examine the role of unfolded proteins and the unfolded protein response (UPR) in hypoxic injury. Multiple studies primarily with cancer cells have found that hypoxia results in an increase in unfolded proteins that may promote cell death. We have evidence that translational suppression protects from hypoxia by reducing the level or toxicity of unfolded proteins. Using C. elegans genetic tools, we will perform a more extensive examination of how translational suppression, unfolded proteins, and the cellular response to unfolded proteins interact to control acute hypoxic injury. (3) Determine whether the orthologs of the implicated C. elegans genes similarly control hypoxic death of mouse primary neurons. In collaboration with Jeff Milbrandt in the Department of Pathology at Washington University, we will determine whether reducing the expression of genes implicated in specific aims 1 and 2 protect mouse neurons from traumatic and hypoxic injury. These experiments will accomplish two goals: First, determine whether the mouse orthologs of the C. elegans genes play similarly important roles in hypoxic neuronal injury and second, whether these genes can be targeted without baseline neural toxicity. Completion of these aims will provide a more complete understanding of the mechanism whereby translational suppression and the unfolded protein response control hypoxic sensitivity and will determine the feasibility of genetic manipulation of these pathways for hypoxic protection of mammalian neurons.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
2R01NS045905-05A2
Application #
7730548
Study Section
Neural Oxidative Metabolism and Death Study Section (NOMD)
Program Officer
Sutherland, Margaret L
Project Start
2009-08-01
Project End
2011-08-31
Budget Start
2009-09-15
Budget End
2010-08-31
Support Year
5
Fiscal Year
2009
Total Cost
$380,000
Indirect Cost
Name
Washington University
Department
Anesthesiology
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Sun, Chun-Ling; Zhang, Huiliang; Liu, Meng et al. (2017) A screen for protective drugs against delayed hypoxic injury. PLoS One 12:e0176061
Kaufman, Daniel M; Wu, Xia; Scott, Barbara A et al. (2017) Ageing and hypoxia cause protein aggregation in mitochondria. Cell Death Differ 24:1730-1738
Mao, X R; Kaufman, D M; Crowder, C M (2016) Nicotinamide mononucleotide adenylyltransferase promotes hypoxic survival by activating the mitochondrial unfolded protein response. Cell Death Dis 7:e2113
Kaufman, Daniel M; Crowder, C Michael (2015) Mitochondrial Proteostatic Collapse Leads to Hypoxic Injury. Curr Biol 25:2171-6
Sun, C-L; Kim, E; Crowder, C M (2014) Delayed innocent bystander cell death following hypoxia in Caenorhabditis elegans. Cell Death Differ 21:557-67
Scott, Barbara; Sun, Chun-Ling; Mao, Xianrong et al. (2013) Role of oxygen consumption in hypoxia protection by translation factor depletion. J Exp Biol 216:2283-92
Mao, Xianrong R; Crowder, C Michael (2010) Protein misfolding induces hypoxic preconditioning via a subset of the unfolded protein response machinery. Mol Cell Biol 30:5033-42
Mabon, Meghann E; Mao, Xianrong; Jiao, York et al. (2009) Systematic identification of gene activities promoting hypoxic death. Genetics 181:483-96
Anderson, Lori L; Mao, Xianrong; Scott, Barbara A et al. (2009) Survival from hypoxia in C. elegans by inactivation of aminoacyl-tRNA synthetases. Science 323:630-3
Crowder, C Michael (2009) Cell biology. Ceramides--friend or foe in hypoxia? Science 324:343-4

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