title: Analysis of mitochondrial metabolism in situ: Combining stable isotope labeling with selective permeabilization.
abstract: To date, it is well-established that mitochondrial dysfunction does not only play a vital role in cancer but also in other pathological conditions such as neurodegenerative diseases and inflammation. An important tool for the analysis of cellular metabolism is the application of stable isotope labeled substrates, which allow for the tracing of atoms throughout metabolic networks. While such analyses yield very detailed information about intracellular fluxes, the determination of compartment specific fluxes is far more challenging. Most approaches for the deconvolution of compartmented metabolism use computational models whereas experimental methods are rare. Here, we developed an experimental setup based on selective permeabilization of the cytosolic membrane that allows for the administration of stable isotope labeled substrates directly to mitochondria. We demonstrate how this approach can be used to infer metabolic changes in mitochondria induced by either chemical or genetic perturbations and give an outlook on its potential applications.
authors.affiliation: Technische Universiät Braunschweig, Department of Bioinfomatics and Biochemistry and Braunschweig Integrated Center of Systems Biology (BRICS), Rebenring 56, 38106 Braunschweig, Germany; Luxemburg Centre for Systems Biomedicine, University of Luxemburg, House of Biomedicine II, L-4367 Belvaux, Luxemburg.
authors.forename: Yannic
authors.initials: Y
authors.lastname: Nonnenmacher
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126 Milan, Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Palorini
authors.affiliation: Luxemburg Centre for Systems Biomedicine, University of Luxemburg, House of Biomedicine II, L-4367 Belvaux, Luxemburg.
authors.forename: Aymeric Fouquier
authors.initials: AF
authors.lastname: d'Herouël
authors.affiliation: Technische Universiät Braunschweig, Department of Bioinfomatics and Biochemistry and Braunschweig Integrated Center of Systems Biology (BRICS), Rebenring 56, 38106 Braunschweig, Germany; Luxemburg Centre for Systems Biomedicine, University of Luxemburg, House of Biomedicine II, L-4367 Belvaux, Luxemburg.
authors.forename: Lisa
authors.initials: L
authors.lastname: Krämer
authors.affiliation: Luxemburg Centre for Systems Biomedicine, University of Luxemburg, House of Biomedicine II, L-4367 Belvaux, Luxemburg.
authors.forename: Meina
authors.initials: M
authors.lastname: Neumann-Schaal
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126 Milan, Italy.
authors.forename: Ferdinando
authors.initials: F
authors.lastname: Chiaradonna
authors.affiliation: Technische Universiät Braunschweig, Department of Bioinfomatics and Biochemistry and Braunschweig Integrated Center of Systems Biology (BRICS), Rebenring 56, 38106 Braunschweig, Germany; National Centre for Microscopy and Imaging Research, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0608, USA.
authors.forename: Alexander
authors.initials: A
authors.lastname: Skupin
authors.affiliation: Technische Universiät Braunschweig, Department of Bioinfomatics and Biochemistry and Braunschweig Integrated Center of Systems Biology (BRICS), Rebenring 56, 38106 Braunschweig, Germany.
authors.forename: Andre
authors.initials: A
authors.lastname: Wegner
authors.affiliation: Technische Universiät Braunschweig, Department of Bioinfomatics and Biochemistry and Braunschweig Integrated Center of Systems Biology (BRICS), Rebenring 56, 38106 Braunschweig, Germany; Luxemburg Centre for Systems Biomedicine, University of Luxemburg, House of Biomedicine II, L-4367 Belvaux, Luxemburg; Computational Biology of Infection Research, Helmholtz Centre for Infection Research, Inhoffenstraße 7, 38124 Braunschweig, Germany. Electronic address: karsten.hiller@tu-braunschweig.de.
authors.forename: Karsten
authors.initials: K
authors.lastname: Hiller
mesh_terms: D000072283:A549 Cells; D000230:Adenocarcinoma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D007553:Isotope Labeling; D008928:Mitochondria / Q000378:metabolism* / Q000473:pathology; D051336:Mitochondrial Membranes / Q000378:metabolism*; D010539:Permeability
keywords: Cancer; Metabolism; Mitochondria; Permeabilization; Stable isotopes