title: Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.
abstract: The relationship of the inner mitochondrial membrane (IMM) cristae structure and intracristal space (ICS) to oxidative phosphorylation (oxphos) is not well understood. Mitofilin (subunit Mic60) of the mitochondrial contact site and cristae organizing system (MICOS) IMM complex is attached to the outer membrane (OMM) via the sorting and assembly machinery/topogenesis of mitochondrial outer membrane β-barrel proteins (SAM/TOB) complex and controls the shape of the cristae. ATP synthase dimers determine sharp cristae edges, whereas trimeric OPA1 tightens ICS outlets. Metabolism is altered during hypoxia, and we therefore studied cristae morphology in HepG2 cells adapted to 5% oxygen for 72 h. Three dimensional (3D), super-resolution biplane fluorescence photoactivation localization microscopy with Eos-conjugated, ICS-located lactamase-β indicated hypoxic ICS expansion with an unchanged OMM (visualized by Eos-mitochondrial fission protein-1). 3D direct stochastic optical reconstruction microscopy immunocytochemistry revealed foci of clustered mitofilin (but not MICOS subunit Mic19) in contrast to its even normoxic distribution. Mitofilin mRNA and protein decreased by ∼20%. ATP synthase dimers vs monomers and state-3/state-4 respiration ratios were lower during hypoxia. Electron microscopy confirmed ICS expansion (maximum in glycolytic cells), which was absent in reduced or OMM-detached cristae of OPA1- and mitofilin-silenced cells, respectively. Hypoxic adaptation is reported as rounding sharp cristae edges and expanding cristae width (ICS) by partial mitofilin/Mic60 down-regulation. Mitofilin-depleted MICOS detaches from SAM while remaining MICOS with mitofilin redistributes toward higher interdistances. This phenomenon causes partial oxphos dormancy in glycolytic cells via disruption of ATP synthase dimers.-Plecitá-Hlavatá, L., Engstová, H., Alán, L., Špaček, T., Dlasková, A., Smolková, K., Špačková, J., Tauber, J., Strádalová, V., Malínský, J., Lessard, M., Bewersdorf, J., Ježek, P. Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Lydie
authors.initials: L
authors.lastname: Plecitá-Hlavatá
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Hana
authors.initials: H
authors.lastname: Engstová
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Lukáš
authors.initials: L
authors.lastname: Alán
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Tomáš
authors.initials: T
authors.lastname: Špaček
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Andrea
authors.initials: A
authors.lastname: Dlasková
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Katarína
authors.initials: K
authors.lastname: Smolková
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Jitka
authors.initials: J
authors.lastname: Špačková
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and.
authors.forename: Jan
authors.initials: J
authors.lastname: Tauber
authors.affiliation: Microscopy Unit, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Prague, Czech Republic;
authors.forename: Vendula
authors.initials: V
authors.lastname: Strádalová
authors.affiliation: Microscopy Unit, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Prague, Czech Republic;
authors.forename: Jan
authors.initials: J
authors.lastname: Malínský
authors.affiliation: Institute for Molecular Biophysics, The Jackson Laboratory, Bar Harbor, Maine, USA; and.
authors.forename: Mark
authors.initials: M
authors.lastname: Lessard
authors.affiliation: Department of Cell Biology, Yale University, New Haven, Connecticut, USA.
authors.forename: Joerg
authors.initials: J
authors.lastname: Bewersdorf
authors.affiliation: Department of Membrane Transport Biophysics, Department No. 75, Institute of Physiology, and jezek@biomed.cas.cz.
authors.forename: Petr
authors.initials: P
authors.lastname: Ježek
mesh_terms: D025181:ATP Synthetase Complexes / Q000378:metabolism*; D000222:Adaptation, Physiological / Q000502:physiology*; D000255:Adenosine Triphosphate / Q000096:biosynthesis*; D015536:Down-Regulation; D005786:Gene Expression Regulation / Q000502:physiology; D056945:Hep G2 Cells; D006801:Humans; D008928:Mitochondria / Q000502:physiology*; D063154:Mitochondrial Dynamics / Q000502:physiology; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000502:physiology; D010100:Oxygen; D054730:Protein Interaction Domains and Motifs; D021122:Protein Subunits
chemical_list: D024101:Mitochondrial Proteins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D000255:Adenosine Triphosphate; D025181:ATP Synthetase Complexes; D010100:Oxygen
keywords: Mic60; OPA1; dSTORM 3D immunocytochemistry; electron microscopy; mitochondrial cristae morphology; mitofilin