The genetically encoded tool that increases the NADH/NAD+ ratio of living cells
Impaired reduction/oxidation metabolism (redox) is a major contributor to many diseases including cancer, neurodegeneration and aging. Mechanistic studies on redox balance are difficult due to the limited strategies that can disrupt cellular metabolism and model pathology without creating additional and potentially confusing metabolic perturbations. Most studies on impaired redox metabolic state have concentrated on oxidative stress, and the production of reactive oxygen species (ROS). As a result, little is known about settings with an excess of reducing equivalents. The NADH reductive state has been modelled using pharmacological inhibition of the electron transfer chain (ETC), and ethanol addition. Both methods, however, have some significant disadvantages. We introduce a soluble trhodhydrogenase (Ec STH), a genetically encoded tool, to promote NADH production in living cells. When expressed in mammalian cells, Ec STH, and a mitochondrially-targeted version (mito Ec STH), can elevate the NADH/NAD+ ratio in a compartment-specific manner. We use this tool to determine the metabolic signatures and transcriptomic profiles of NADH-reductive stress within mammalian cell. The cellular response to NADH-reductive stress is dependent on the cell background. We identify metabolic reactions that detect changes in the NADH/NAD+ ratio. Our novel genetically encoded tool is an orthogonal approach to disrupt redox metabolism, characterize its impact on normal physiology, and study disease states.
The authors have not declared any competing interests.
Source:
https://www.biorxiv.org/content/10.1101/2022.09.20.508785v1?ct=
