title: NCLX protein, but not LETM1, mediates mitochondrial Ca2+ extrusion, thereby limiting Ca2+-induced NAD(P)H production and modulating matrix redox state.
abstract: Mitochondria capture and subsequently release Ca(2+) ions, thereby sensing and shaping cellular Ca(2+) signals. The Ca(2+) uniporter MCU mediates Ca(2+) uptake, whereas NCLX (mitochondrial Na/Ca exchanger) and LETM1 (leucine zipper-EF-hand-containing transmembrane protein 1) were proposed to exchange Ca(2+) against Na(+) or H(+), respectively. Here we study the role of these ion exchangers in mitochondrial Ca(2+) extrusion and in Ca(2+)-metabolic coupling. Both NCLX and LETM1 proteins were expressed in HeLa cells mitochondria. The rate of mitochondrial Ca(2+) efflux, measured with a genetically encoded indicator during agonist stimulations, increased with the amplitude of mitochondrial Ca(2+) ([Ca(2+)]mt) elevations. NCLX overexpression enhanced the rates of Ca(2+) efflux, whereas increasing LETM1 levels had no impact on Ca(2+) extrusion. The fluorescence of the redox-sensitive probe roGFP increased during [Ca(2+)]mt elevations, indicating a net reduction of the matrix. This redox response was abolished by NCLX overexpression and restored by the Na(+)/Ca(2+) exchanger inhibitor CGP37157. The [Ca(2+)]mt elevations were associated with increases in the autofluorescence of NAD(P)H, whose amplitude was strongly reduced by NCLX overexpression, an effect reverted by Na(+)/Ca(2+) exchange inhibition. We conclude that NCLX, but not LETM1, mediates Ca(2+) extrusion from mitochondria. By controlling the duration of matrix Ca(2+) elevations, NCLX contributes to the regulation of NAD(P)H production and to the conversion of Ca(2+) signals into redox changes.
authors.affiliation: From the Mitochondrial Function, Nestlé Institute of Health Sciences, EPFL Innovation Park, Building G, CH-1015 Lausanne, Switzerland, the Department of Cell Physiology and Metabolism, University of Geneva, Rue Michel-Servet, 1, CH-1211 Genève, Switzerland, and Umberto.DeMarchi@rd.nestle.com.
authors.forename: Umberto
authors.initials: U
authors.lastname: De Marchi
authors.affiliation: From the Mitochondrial Function, Nestlé Institute of Health Sciences, EPFL Innovation Park, Building G, CH-1015 Lausanne, Switzerland, the Department of Cell Physiology and Metabolism, University of Geneva, Rue Michel-Servet, 1, CH-1211 Genève, Switzerland, and.
authors.forename: Jaime
authors.initials: J
authors.lastname: Santo-Domingo
authors.affiliation: the Department of Cell Physiology and Metabolism, University of Geneva, Rue Michel-Servet, 1, CH-1211 Genève, Switzerland, and.
authors.forename: Cyril
authors.initials: C
authors.lastname: Castelbou
authors.affiliation: the Department of Physiology, Ben-Gurion University of Negev, Beer-Sheva 84105, Israel.
authors.forename: Israel
authors.initials: I
authors.lastname: Sekler
authors.affiliation: From the Mitochondrial Function, Nestlé Institute of Health Sciences, EPFL Innovation Park, Building G, CH-1015 Lausanne, Switzerland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Wiederkehr
authors.affiliation: the Department of Cell Physiology and Metabolism, University of Geneva, Rue Michel-Servet, 1, CH-1211 Genève, Switzerland, and Nicolas.Demaurex@unige.ch.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Demaurex
mesh_terms: D020013:Calcium Signaling / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002998:Clonazepam / Q000031:analogs & derivatives / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D009243:NAD / Q000096:biosynthesis; D009249:NADP / Q000096:biosynthesis; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013841:Thiazepines / Q000494:pharmacology
chemical_list: D002135:Calcium-Binding Proteins; C488528:LETM1 protein, human; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D011994:Recombinant Proteins; C483046:SLC8B1 protein, human; D019831:Sodium-Calcium Exchanger; D013841:Thiazepines; D009243:NAD; D009249:NADP; D002998:Clonazepam; C079032:CGP 37157
keywords: Calcium Imaging; Calcium Signaling; Mitochondria; Mitochondrial Transport; Redox Signaling