title: The mitochondrial calcium uniporter is a multimer that can include a dominant-negative pore-forming subunit.
abstract: Mitochondrial calcium uniporter (MCU) channel is responsible for Ruthenium Red-sensitive mitochondrial calcium uptake. Here, we demonstrate MCU oligomerization by immunoprecipitation and Förster resonance energy transfer (FRET) and characterize a novel protein (MCUb) with two predicted transmembrane domains, 50% sequence similarity and a different expression profile from MCU. Based on computational modelling, MCUb includes critical amino-acid substitutions in the pore region and indeed MCUb does not form a calcium-permeable channel in planar lipid bilayers. In HeLa cells, MCUb is inserted into the oligomer and exerts a dominant-negative effect, reducing the [Ca(2+)]mt increases evoked by agonist stimulation. Accordingly, in vitro co-expression of MCUb with MCU drastically reduces the probability of observing channel activity in planar lipid bilayer experiments. These data unveil the structural complexity of MCU and demonstrate a novel regulatory mechanism, based on the inclusion of dominant-negative subunits in a multimeric channel, that underlies the fine control of the physiologically and pathologically relevant process of mitochondrial calcium homeostasis.
authors.affiliation: Department of Biomedical Sciences, University of Padua and CNR Neuroscience Institute, Padua, Italy.
authors.forename: Anna
authors.initials: A
authors.lastname: Raffaello
authors.forename: Diego
authors.initials: D
authors.lastname: De Stefani
authors.forename: Davide
authors.initials: D
authors.lastname: Sabbadin
authors.forename: Enrico
authors.initials: E
authors.lastname: Teardo
authors.forename: Giulia
authors.initials: G
authors.lastname: Merli
authors.forename: Anne
authors.initials: A
authors.lastname: Picard
authors.forename: Vanessa
authors.initials: V
authors.lastname: Checchetto
authors.forename: Stefano
authors.initials: S
authors.lastname: Moro
authors.forename: Ildikò
authors.initials: I
authors.lastname: Szabò
authors.forename: Rosario
authors.initials: R
authors.lastname: Rizzuto
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D006367:HeLa Cells; D006706:Homeostasis; D006801:Humans; D008051:Lipid Bilayers; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D021122:Protein Subunits
chemical_list: D015220:Calcium Channels; D008051:Lipid Bilayers; D021122:Protein Subunits; C034703:mitochondrial calcium uniporter; D002118:Calcium
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