Tracing Compartmentalized NADPH Metabolism in the Cytosol and Mitochondria of Mammalian Cells

Overview

Journal Mol Cell

Publisher Cell Press

Specialty Cell Biology

Date 2014 Jun 3

PMID 24882210

Citations 334

Authors

Caroline A Lewis

Seth J Parker

Brian P Fiske

Douglas McCloskey

Dan Y Gui

Courtney R Green

Natalie I Vokes

Adam M Feist

Matthew G Vander Heiden

Christian M Metallo

Affiliations

Soon will be listed here.

Abstract

Eukaryotic cells compartmentalize biochemical processes in different organelles, often relying on metabolic cycles to shuttle reducing equivalents across intracellular membranes. NADPH serves as the electron carrier for the maintenance of redox homeostasis and reductive biosynthesis, with separate cytosolic and mitochondrial pools providing reducing power in each respective location. This cellular organization is critical for numerous functions but complicates analysis of metabolic pathways using available methods. Here we develop an approach to resolve NADP(H)-dependent pathways present within both the cytosol and the mitochondria. By tracing hydrogen in compartmentalized reactions that use NADPH as a cofactor, including the production of 2-hydroxyglutarate by mutant isocitrate dehydrogenase enzymes, we can observe metabolic pathway activity in these distinct cellular compartments. Using this system we determine the direction of serine/glycine interconversion within the mitochondria and cytosol, highlighting the ability of this approach to resolve compartmentalized reactions in intact cells.

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