title: Superresolution microscopy reveals spatial separation of UCP4 and F0F1-ATP synthase in neuronal mitochondria.
abstract: Because different proteins compete for the proton gradient across the inner mitochondrial membrane, an efficient mechanism is required for allocation of associated chemical potential to the distinct demands, such as ATP production, thermogenesis, regulation of reactive oxygen species (ROS), etc. Here, we used the superresolution technique dSTORM (direct stochastic optical reconstruction microscopy) to visualize several mitochondrial proteins in primary mouse neurons and test the hypothesis that uncoupling protein 4 (UCP4) and F0F1-ATP synthase are spatially separated to eliminate competition for the proton motive force. We found that UCP4, F0F1-ATP synthase, and the mitochondrial marker voltage-dependent anion channel (VDAC) have various expression levels in different mitochondria, supporting the hypothesis of mitochondrial heterogeneity. Our experimental results further revealed that UCP4 is preferentially localized in close vicinity to VDAC, presumably at the inner boundary membrane, whereas F0F1-ATP synthase is more centrally located at the cristae membrane. The data suggest that UCP4 cannot compete for protons because of its spatial separation from both the proton pumps and the ATP synthase. Thus, mitochondrial morphology precludes UCP4 from acting as an uncoupler of oxidative phosphorylation but is consistent with the view that UCP4 may dissipate the excessive proton gradient, which is usually associated with ROS production.
authors.affiliation: Institute of Applied Physics, Vienna University of Technology, A-1040 Vienna, Austria; and enricoklotzsch@gmail.com schuetz@iap.tuwien.ac.at elena.pohl@vetmeduni.ac.at.
authors.forename: Enrico
authors.initials: E
authors.lastname: Klotzsch
authors.affiliation: Institutes of Physiology, Pathophysiology and Biophysics and.
authors.forename: Alina
authors.initials: A
authors.lastname: Smorodchenko
authors.affiliation: Institute of Applied Physics, Vienna University of Technology, A-1040 Vienna, Austria; and.
authors.forename: Lukas
authors.initials: L
authors.lastname: Löfler
authors.affiliation: Medical Biochemistry, University of Veterinary Medicine, A-1210 Vienna, Austria.
authors.forename: Rudolf
authors.initials: R
authors.lastname: Moldzio
authors.affiliation: Institute of Applied Physics, Vienna University of Technology, A-1040 Vienna, Austria; and.
authors.forename: Elena
authors.initials: E
authors.lastname: Parkinson
authors.affiliation: Institute of Applied Physics, Vienna University of Technology, A-1040 Vienna, Austria; and enricoklotzsch@gmail.com schuetz@iap.tuwien.ac.at elena.pohl@vetmeduni.ac.at.
authors.forename: Gerhard J
authors.initials: GJ
authors.lastname: Schütz
authors.affiliation: Institutes of Physiology, Pathophysiology and Biophysics and enricoklotzsch@gmail.com schuetz@iap.tuwien.ac.at elena.pohl@vetmeduni.ac.at.
authors.forename: Elena E
authors.initials: EE
authors.lastname: Pohl
mesh_terms: D000818:Animals; D053078:Membrane Potential, Mitochondrial; D026901:Membrane Transport Proteins / Q000378:metabolism*; D051379:Mice; D008853:Microscopy / Q000379:methods*; D008928:Mitochondria / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000378:metabolism; D000071245:Mitochondrial Uncoupling Proteins; D009474:Neurons / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D011522:Protons; D050994:Voltage-Dependent Anion Channels / Q000378:metabolism
chemical_list: D026901:Membrane Transport Proteins; D000071245:Mitochondrial Uncoupling Proteins; D011522:Protons; C547516:Slc25a27 protein, mouse; D050994:Voltage-Dependent Anion Channels; D006180:Proton-Translocating ATPases
keywords: direct stochastic optical reconstruction microscopy; mitochondrial membrane proteins; proton diffusion; reactive oxygen species; uncoupling