Disruption of protein folding in the endoplasmic reticulum??ER stress??is associated with many different metabolic diseases, particularly those associated with obesity that affect between 22 and 30 percent of adults in the U.S. Because of this exceptional disease burden, it is important to understand the factors that cause ER stress during metabolic dysregulation. Yet the pathways by which metabolic activity and ER homeostasis are coupled are poorly understood. Mitochondria are central to metabolism, and the TCA cycle is the hub of this activity, accepting substrates from glycolysis and fatty acid oxidation for catabolism, generating reducing equivalents for electron transport and for the maintenance of cellular redox homeostasis, and providing building materials for the reductive biosynthesis of lipids, glucose, and amino acids. Because of its centrality to so many processes, flux through the TCA cycle is likely to affect many diverse cellular pathways, even those with no obvious direct connection. This includes ER protein processing, which is sensitive to changes in redox state, amino acid availability, and cellular lipid content. In this proposal, we provide evidence for a previously unknown functional relationship between TCA cycle activity and ER homeostasis in metabolically active cells, including hepatocytes, myocytes, and adipocytes, that depends on production of NADPH by the TCA cycle and redox regulation of glutathione. This proposal is designed to identify the basic mechanisms linking TCA-dependent NADPH production in the mitochondria to homeostasis in the ER. Toward that end, we propose three specific aims: (1) Determine how NADPH production and compartmentalization link nutrient flow to ER stress; (2) Determine how changes to mitochondrial and cytosolic glutathione redox promote ER oxidation; and (3) Determine how TCA activity and glutathione redox alter ER function. We will achieve these aims using a combination of genetic and pharmacological tools to manipulate TCA cycle activity; cutting-edge biosensors to monitor changes in cellular redox status; manipulation and analysis of ER-mitochondrial contacts; and molecular biology approaches to manipulate and assess ER functionality. The outcome of this work will be a mechanistic understanding of how metabolic activity alters ER function to contribute to disease.
Endoplasmic reticulum (ER) stress is associated with many different human metabolic diseases including diabetes, atherosclerosis, steatohepatitis, and others, but the pathways connecting metabolic activity to ER stress are poorly understood. We have uncovered a novel means of mitochondria-to-ER communication by which TCA cycle activity regulates ER homeostasis, and this work is directed toward identifying the basic mechanisms by which these processes are linked. The fundamental insights arising from our work will be relevant to understanding how ER stress arises during metabolic dysregulation, and how it might be prevented to mitigate disease.
| Rutkowski, D Thomas (2018) Liver function and dysfunction - a unique window into the physiological reach of ER stress and the unfolded protein response. FEBS J : |
| Diedrichs, Danilo R; Gomez, Javier A; Huang, Chun-Sing et al. (2018) A data-entrained computational model for testing the regulatory logic of the vertebrate unfolded protein response. Mol Biol Cell 29:1502-1517 |
| DeZwaan-McCabe, Diane; Sheldon, Ryan D; Gorecki, Michelle C et al. (2017) ER Stress Inhibits Liver Fatty Acid Oxidation while Unmitigated Stress Leads to Anorexia-Induced Lipolysis and Both Liver and Kidney Steatosis. Cell Rep 19:1794-1806 |
| Staber, J M; Pollpeter, M J; Anderson, C-G et al. (2017) Long-term correction of hemophilia A mice following lentiviral mediated delivery of an optimized canine factor VIII gene. Gene Ther 24:742-748 |
| Gomez, Javier A; Rutkowski, D Thomas (2016) Experimental reconstitution of chronic ER stress in the liver reveals feedback suppression of BiP mRNA expression. Elife 5: |
