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Vamsi K Mootha - One of the best experts on this subject based on the ideXlab platform.
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structural insights into the ca2 dependent gating of the human mitochondrial calcium Uniporter
eLife, 2020Co-Authors: Yan Wang, Vamsi K Mootha, Yan Han, Ji She, Nam X Nguyen, Xiao Chen Bai, Youxing JiangAbstract:Mitochondrial Ca2+ uptake is mediated by an inner mitochondrial membrane protein called the mitochondrial calcium Uniporter. In humans, the Uniporter functions as a holocomplex consisting of MCU, EMRE, MICU1 and MICU2, among which MCU and EMRE form a subcomplex and function as the conductive channel while MICU1 and MICU2 are EF-hand proteins that regulate the channel activity in a Ca2+-dependent manner. Here, we present the EM structures of the human mitochondrial calcium Uniporter holocomplex (uniplex) in the presence and absence of Ca2+, revealing distinct Ca2+ dependent assembly of the uniplex. Our structural observations suggest that Ca2+ changes the dimerization interaction between MICU1 and MICU2, which in turn determines how the MICU1-MICU2 subcomplex interacts with the MCU-EMRE channel and, consequently, changes the distribution of the uniplex assemblies between the blocked and unblocked states.
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an essential role for cardiolipin in the stability and function of the mitochondrial calcium Uniporter
Proceedings of the National Academy of Sciences of the United States of America, 2020Co-Authors: Sagnika Ghosh, Vamsi K Mootha, Writoban Basu Ball, Travis R Madaris, Subramanya Srikantan, Muniswamy Madesh, Vishal M GohilAbstract:Calcium uptake by the mitochondrial calcium Uniporter coordinates cytosolic signaling events with mitochondrial bioenergetics. During the past decade all protein components of the mitochondrial calcium Uniporter have been identified, including MCU, the pore-forming subunit. However, the specific lipid requirements, if any, for the function and formation of this channel complex are currently not known. Here we utilize yeast, which lacks the mitochondrial calcium Uniporter, as a model system to address this problem. We use heterologous expression to functionally reconstitute human Uniporter machinery both in wild-type yeast as well as in mutants defective in the biosynthesis of phosphatidylethanolamine, phosphatidylcholine, or cardiolipin (CL). We uncover a specific requirement of CL for in vivo reconstituted MCU stability and activity. The CL requirement of MCU is evolutionarily conserved with loss of CL triggering rapid turnover of MCU homologs and impaired calcium transport. Furthermore, we observe reduced abundance and activity of endogenous MCU in mammalian cellular models of Barth syndrome, which is characterized by a partial loss of CL. MCU abundance is also decreased in the cardiac tissue of Barth syndrome patients. Our work raises the hypothesis that impaired mitochondrial calcium transport contributes to the pathogenesis of Barth syndrome, and more generally, showcases the utility of yeast phospholipid mutants in dissecting the phospholipid requirements of ion channel complexes.
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structural insights into the ca2 dependent gating of the human mitochondrial calcium Uniporter
bioRxiv, 2020Co-Authors: Yan Wang, Vamsi K Mootha, Yan Han, Ji She, Nam X Nguyen, Xiao Chen Bai, Youxing JiangAbstract:Abstract Mitochondrial Ca2+ uptake plays an important role in cellular physiology such as modulating ATP production, regulating cytoplasmic Ca2+ dynamics, and triggering cell death, and is mediated by the mitochondrial calcium Uniporter, a highly selective calcium channel localized to the inner mitochondrial membrane. In humans, the Uniporter functions as a holocomplex consisting of MCU, EMRE, MICU1 and MICU2, among which MCU and EMRE form a subcomplex and function as the conductive channel while MICU1 and MICU2 are EF-hand proteins that regulate the channel activity in a Ca2+ dependent manner. Here we present the EM structures of the human mitochondrial calcium Uniporter holocomplex (uniplex) in the presence and absence of Ca2+, revealing distinct Ca2+ dependent assembly of the uniplex. In the presence of Ca2+, MICU1 and MICU2 form a heterotetramer of MICU1-(MICU2)2-MICU1 and bridge the dimeric form of the MCU-EMRE subcomplex through electrostatic interactions between MICU1 and EMRE, leaving the MCU channel pore unblocked. In the absence of Ca2+, multiple uniplex assemblies are observed but is predominantly occupied by the MICU1 subunit from a MICU1-MICU2 heterodimer blocking the MCU channel pore. Our structural observations suggest that Ca2+ changes the dimerization interaction between MICU1 and MICU2, which in turn determines how the MICU1-MICU2 subcomplex interacts with the MCU-EMRE channel and, consequently, changes the distribution of the uniplex assemblies between the blocked and unblocked states.
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molecular basis of emre dependence of the human mitochondrial calcium Uniporter
bioRxiv, 2019Co-Authors: Melissa J S Macewen, Olga Goldberger, Vamsi K Mootha, Andrew L Markhard, Mert Bozbeyoglu, Forrest Bradford, Yasemin SancakAbstract:ABSTRACT The mitochondrial Uniporter is calcium-activated calcium channel complex critical for cellular signaling and bioenergetics. MCU, the pore-forming subunit of the Uniporter, contains two transmembrane domains and is found in all major eukaryotic taxa. In amoeba and fungi, MCU homologs are sufficient to form a functional calcium channel, whereas human MCU exhibits a strict requirement for the metazoan-specific, single-pass transmembrane protein EMRE for conductance. Here, we exploit this evolutionary divergence to decipher the molecular basis of the human MCU’s dependence on EMRE. By systematically generating chimeric proteins that consist of EMRE-independent D. discoideum MCU (DdMCU) and H. sapiens MCU (HsMCU), we converged on a stretch of 10 amino acids in DdMCU that can be transplanted to HsMCU to render it EMRE-dependent. We call this region in human MCU the EMRE-dependence domain (EDD). Crosslinking experiments show that HsEMRE directly interacts with MCU at both of its transmembrane domains as well as the EDD. Based on previously published structures of fungal MCU homologs, the EDD segment is located distal to the calcium pore’s selectivity filter and appears flexible. We propose that EMRE stabilizes EDD of MCU, permitting both channel opening and calcium conductance
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exploring the in vivo role of the mitochondrial calcium Uniporter in brown fat bioenergetics
Cell Reports, 2019Co-Authors: Olga Goldberger, Yasemin Sancak, Vamsi K Mootha, Daniel Flicker, Eran MickAbstract:Summary The mitochondrial calcium Uniporter has been proposed to coordinate the organelle’s energetics with calcium signaling. Uniporter current has previously been reported to be extremely high in brown adipose tissue (BAT), yet it remains unknown how the Uniporter contributes to BAT physiology. Here, we report the generation and characterization of a mouse model lacking Mcu, the pore forming subunit of the Uniporter, specifically in BAT (BAT-Mcu-KO). BAT-Mcu-KO mice lack Uniporter-based calcium uptake in BAT mitochondria but exhibit unaffected cold tolerance, diet-induced obesity, and transcriptional response to cold in BAT. Unexpectedly, we found in wild-type animals that cold powerfully activates the ATF4-dependent integrated stress response (ISR) in BAT and upregulates circulating FGF21 and GDF15, raising the hypothesis that the ISR partly underlies the pleiotropic effects of BAT on systemic metabolism. Our study demonstrates that the Uniporter is largely dispensable for BAT thermogenesis and demonstrates activation of the ISR in BAT in response to cold.
Rosario Rizzuto - One of the best experts on this subject based on the ideXlab platform.
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from the identification to the dissection of the physiological role of the mitochondrial calcium Uniporter an ongoing story
Biomolecules, 2021Co-Authors: Giorgia Pallafacchina, Sofia Zanin, Rosario RizzutoAbstract:The notion of mitochondria being involved in the decoding and shaping of intracellular Ca2+ signals has been circulating since the end of the 19th century. Despite that, the molecular identity of the channel that mediates Ca2+ ion transport into mitochondria remained elusive for several years. Only in the last decade, the genes and pathways responsible for the mitochondrial uptake of Ca2+ began to be cloned and characterized. The gene coding for the pore-forming unit of the mitochondrial channel was discovered exactly 10 years ago, and its product was called mitochondrial Ca2+ Uniporter or MCU. Before that, only one of its regulators, the mitochondria Ca2+ uptake regulator 1, MICU1, has been described in 2010. However, in the following years, the scientific interest in mitochondrial Ca2+ signaling regulation and physiological role has increased. This shortly led to the identification of many of its components, to the description of their 3D structure, and the characterization of the Uniporter contribution to tissue physiology and pathology. In this review, we will summarize the most relevant achievements in the history of mitochondrial Ca2+ studies, presenting a chronological overview of the most relevant and landmarking discoveries. Finally, we will explore the impact of mitochondrial Ca2+ signaling in the context of muscle physiology, highlighting the recent advances in understanding the role of the MCU complex in the control of muscle trophism and metabolism.
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the molecular complexity of the mitochondrial calcium Uniporter
Cell Calcium, 2021Co-Authors: Simona Feno, Anna Raffaello, Rosario Rizzuto, Denis Vecellio ReaneAbstract:The role of mitochondria in regulating cellular Ca2+ homeostasis is crucial for the understanding of different cellular functions in physiological and pathological conditions. Nevertheless, the study of this aspect was severely limited by the lack of the molecular identity of the proteins responsible for mitochondrial Ca2+ uptake. In 2011, the discovery of the gene encoding for the Mitochondrial Calcium Uniporter (MCU), the selective channel responsible for mitochondrial Ca2+ uptake, gave rise to an explosion of studies aimed to characterize the composition, the regulation of the channel and its pathophysiological roles. Here, we summarize the recent discoveries on the molecular structure and composition of the MCU complex by providing new insights into the mechanisms that regulate MCU channel activity.
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overexpression of mitochondrial calcium Uniporter causes neuronal death
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Veronica Granatiero, Anna Raffaello, Diego De Stefani, Marco Pacifici, Rosario RizzutoAbstract:Neurodegenerative diseases are a large and heterogeneous group of disorders characterized by selective and progressive death of specific neuronal subtypes. In most of the cases, the pathophysiology is still poorly understood, although a number of hypotheses have been proposed. Among these, dysregulation of Ca2+ homeostasis and mitochondrial dysfunction represent two broadly recognized early events associated with neurodegeneration. However, a direct link between these two hypotheses can be drawn. Mitochondria actively participate to global Ca2+ signaling, and increases of [Ca2+] inside organelle matrix are known to sustain energy production to modulate apoptosis and remodel cytosolic Ca2+ waves. Most importantly, while mitochondrial Ca2+ overload has been proposed as the no-return signal, triggering apoptotic or necrotic neuronal death, until now direct evidences supporting this hypothesis, especially in vivo, are limited. Here, we took advantage of the identification of the mitochondrial Ca2+ Uniporter (MCU) and tested whether mitochondrial Ca2+ signaling controls neuronal cell fate. We overexpressed MCU both in vitro, in mouse primary cortical neurons, and in vivo, through stereotaxic injection of MCU-coding adenoviral particles in the brain cortex. We first measured mitochondrial Ca2+ uptake using quantitative genetically encoded Ca2+ probes, and we observed that the overexpression of MCU causes a dramatic increase of mitochondrial Ca2+ uptake both at resting and after membrane depolarization. MCU-mediated mitochondrial Ca2+ overload causes alteration of organelle morphology and dysregulation of global Ca2+ homeostasis. Most importantly, MCU overexpression in vivo is sufficient to trigger gliosis and neuronal loss. Overall, we demonstrated that mitochondrial Ca2+ overload is per se sufficient to cause neuronal cell death both in vitro and in vivo, thus highlighting a potential key step in neurodegeneration.
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calcium at the center of cell signaling interplay between endoplasmic reticulum mitochondria and lysosomes
Trends in Biochemical Sciences, 2016Co-Authors: Anna Raffaello, Cristina Mammucari, Rosario Rizzuto, Gaia GherardiAbstract:In recent years, rapid discoveries have been made relating to Ca2+ handling at specific organelles that have important implications for whole-cell Ca2+ homeostasis. In particular, the structures of the endoplasmic reticulum (ER) Ca2+ channels revealed by electron cryomicroscopy (cryo-EM), continuous updates on the structure, regulation, and role of the mitochondrial calcium Uniporter (MCU) complex, and the analysis of lysosomal Ca2+ signaling are milestones on the route towards a deeper comprehension of the complexity of global Ca2+ signaling. In this review we summarize recent discoveries on the regulation of interorganellar Ca2+ homeostasis and its role in pathophysiology.
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structure and function of the mitochondrial calcium Uniporter complex
Biochimica et Biophysica Acta, 2015Co-Authors: Diego De Stefani, Maria Patron, Rosario RizzutoAbstract:The mitochondrial calcium Uniporter (MCU) is the critical protein of the inner mitochondrial membrane mediating the electrophoretic Ca²⁺ uptake into the matrix. It plays a fundamental role in the shaping of global calcium signaling and in the control of aerobic metabolism as well as apoptosis. Two features of mitochondrial calcium signaling have been known for a long time: i) mitochondrial Ca²⁺ uptake widely varies among cells and tissues, and ii) channel opening strongly relies on the extramitochondrial Ca²⁺ concentration, with low activity at resting [Ca²⁺] and high capacity as soon as calcium signaling is activated. Such complexity requires a specialized molecular machinery, with several primary components can be variably gathered together in order to match energy demands and protect from toxic stimuli. In line with this, MCU is now recognized to be part of a macromolecular complex known as the MCU complex. Our understanding of the structure and function of the MCU complex is now growing promptly, revealing an unexpected complexity that highlights the pleiotropic role of mitochondrial Ca²⁺ signals. This article is part of a Special Issue entitled: 13th European Symposium on Calcium.
Kimberli J Kamer - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of micu2 and comparison with micu1 reveal insights into the Uniporter gating mechanism
Proceedings of the National Academy of Sciences of the United States of America, 2019Co-Authors: Kimberli J Kamer, Vamsi K Mootha, Zenon Grabarek, Wei Jiang, Virendar K KaushikAbstract:The mitochondrial Uniporter is a Ca2+-channel complex resident within the organelle’s inner membrane. In mammalian cells the Uniporter’s activity is regulated by Ca2+ due to concerted action of MICU1 and MICU2, two paralogous, but functionally distinct, EF-hand Ca2+-binding proteins. Here we present the X-ray structure of the apo form of Mus musculus MICU2 at 2.5-A resolution. The core structure of MICU2 is very similar to that of MICU1. It consists of two lobes, each containing one canonical Ca2+-binding EF-hand (EF1, EF4) and one structural EF-hand (EF2, EF3). Two molecules of MICU2 form a symmetrical dimer stabilized by highly conserved hydrophobic contacts between exposed residues of EF1 of one monomer and EF3 of another. Similar interactions stabilize MICU1 dimers, allowing exchange between homo- and heterodimers. The tight EF1–EF3 interface likely accounts for the structural and functional coupling between the Ca2+-binding sites in MICU1, MICU2, and their complex that leads to the previously reported Ca2+-binding cooperativity and dominant negative effect of mutation of the Ca2+-binding sites in either protein. The N- and C-terminal segments of the two proteins are distinctly different. In MICU2 the C-terminal helix is significantly longer than in MICU1, and it adopts a more rigid structure. MICU2’s C-terminal helix is dispensable in vitro for its interaction with MICU1 but required for MICU2’s function in cells. We propose that in the MICU1–MICU2 oligomeric complex the C-terminal helices of both proteins form a central semiautonomous assembly which contributes to the gating mechanism of the Uniporter.
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micu1 imparts the mitochondrial Uniporter with the ability to discriminate between ca2 and mn2
Proceedings of the National Academy of Sciences of the United States of America, 2018Co-Authors: Yasemin Sancak, Kimberli J Kamer, Dipayan Chaudhuri, Yevgenia Fomina, Joshua D Meisel, Zenon GrabarekAbstract:The mitochondrial Uniporter is a Ca2+-activated Ca2+ channel complex that displays exceptionally high conductance and selectivity. Here, we report cellular metal toxicity screens highlighting the Uniporter’s role in Mn2+ toxicity. Cells lacking the pore-forming Uniporter subunit, MCU, are more resistant to Mn2+ toxicity, while cells lacking the Ca2+-sensing inhibitory subunit, MICU1, are more sensitive than the wild type. Consistent with these findings, Caenorhabditis elegans lacking the Uniporter’s pore have increased resistance to Mn2+ toxicity. The chemical–genetic interaction between Uniporter machinery and Mn2+ toxicity prompted us to hypothesize that Mn2+ can indeed be transported by the Uniporter’s pore, but this transport is prevented by MICU1. To this end, we demonstrate that, in the absence of MICU1, both Mn2+ and Ca2+ can pass through the Uniporter, as evidenced by mitochondrial Mn2+ uptake assays, mitochondrial membrane potential measurements, and mitoplast electrophysiology. We show that Mn2+ does not elicit the conformational change in MICU1 that is physiologically elicited by Ca2+, preventing Mn2+ from inducing the pore opening. Our work showcases a mechanism by which a channel’s auxiliary subunit can contribute to its apparent selectivity and, furthermore, may have implications for understanding how manganese contributes to neurodegenerative disease.
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high affinity cooperative ca2 binding by micu1 micu2 serves as an on off switch for the Uniporter
EMBO Reports, 2017Co-Authors: Kimberli J Kamer, Vamsi K Mootha, Zenon GrabarekAbstract:Abstract The mitochondrial calcium Uniporter is a Ca 2+ ‐activated Ca 2+ channel that is essential for dynamic modulation of mitochondrial function in response to cellular Ca 2+ signals. It is regulated by two paralogous EF‐hand proteins—MICU1 and MICU2, but the mechanism is unknown. Here, we demonstrate that both MICU1 and MICU2 are stabilized by Ca 2+ . We reconstitute the MICU1–MICU2 heterodimer and demonstrate that it binds Ca 2+ cooperatively with high affinity. We discover that both MICU1 and MICU2 exhibit affinity for the mitochondria‐specific lipid cardiolipin. We determine the minimum Ca 2+ concentration required for disinhibition of the Uniporter in permeabilized cells and report a close match with the Ca 2+ ‐binding affinity of MICU1–MICU2. We conclude that cooperative, high‐affinity interaction of the MICU1–MICU2 complex with Ca 2+ serves as an on–off switch, leading to a tightly controlled channel, capable of responding directly to cytosolic Ca 2+ signals.
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the molecular era of the mitochondrial calcium Uniporter
Nature Reviews Molecular Cell Biology, 2015Co-Authors: Kimberli J Kamer, Vamsi K MoothaAbstract:The mitochondrial calcium Uniporter is an evolutionarily conserved calcium channel, and its biophysical properties and relevance to cell death, bioenergetics and signalling have been investigated for decades. However, the genes encoding this channel have only recently been discovered, opening up a new 'molecular era' in the study of its biology. We now know that the Uniporter is not a single protein but rather a macromolecular complex consisting of pore-forming and regulatory subunits. We review recent studies that harnessed the power of molecular biology and genetics to characterize the mechanism of action of the Uniporter, its evolution and its contribution to physiology and human disease.
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in vivo reconstitution of the mitochondrial Uniporter
Biophysical Journal, 2015Co-Authors: Erika Kovacsbogdan, Yasemin Sancak, Molly Plovanich, Kimberli J Kamer, Ashwini Jambhekar, Michael A Myre, Michael D Blower, Robert Huber, Vamsi K MoothaAbstract:The mitochondrial Uniporter is a highly selective calcium channel present broadly in eukaryotes, but absent in Saccharomyces cerevisiae. Therefore, we used yeast as a reconstitution system to identify the minimal components sufficient for in vivo Uniporter activity. First, we considered Dictyostelium discoideum and showed that it has a highly simplified Uniporter machinery: the expression of DdMCU, a single transmembrane component alone is sufficient to reconstitute mitochondrial calcium Uniporter activity. Second, to establish human Uniporter activity, the coexpression of MCU and - the animal specific protein - EMRE is necessary, whereas expression of MCU alone is insufficient. Our work established yeast as a powerful in vivo reconstitution system for the Uniporter to study the evolution and function of this channel.
Anna Raffaello - One of the best experts on this subject based on the ideXlab platform.
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the molecular complexity of the mitochondrial calcium Uniporter
Cell Calcium, 2021Co-Authors: Simona Feno, Anna Raffaello, Rosario Rizzuto, Denis Vecellio ReaneAbstract:The role of mitochondria in regulating cellular Ca2+ homeostasis is crucial for the understanding of different cellular functions in physiological and pathological conditions. Nevertheless, the study of this aspect was severely limited by the lack of the molecular identity of the proteins responsible for mitochondrial Ca2+ uptake. In 2011, the discovery of the gene encoding for the Mitochondrial Calcium Uniporter (MCU), the selective channel responsible for mitochondrial Ca2+ uptake, gave rise to an explosion of studies aimed to characterize the composition, the regulation of the channel and its pathophysiological roles. Here, we summarize the recent discoveries on the molecular structure and composition of the MCU complex by providing new insights into the mechanisms that regulate MCU channel activity.
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overexpression of mitochondrial calcium Uniporter causes neuronal death
Oxidative Medicine and Cellular Longevity, 2019Co-Authors: Veronica Granatiero, Anna Raffaello, Diego De Stefani, Marco Pacifici, Rosario RizzutoAbstract:Neurodegenerative diseases are a large and heterogeneous group of disorders characterized by selective and progressive death of specific neuronal subtypes. In most of the cases, the pathophysiology is still poorly understood, although a number of hypotheses have been proposed. Among these, dysregulation of Ca2+ homeostasis and mitochondrial dysfunction represent two broadly recognized early events associated with neurodegeneration. However, a direct link between these two hypotheses can be drawn. Mitochondria actively participate to global Ca2+ signaling, and increases of [Ca2+] inside organelle matrix are known to sustain energy production to modulate apoptosis and remodel cytosolic Ca2+ waves. Most importantly, while mitochondrial Ca2+ overload has been proposed as the no-return signal, triggering apoptotic or necrotic neuronal death, until now direct evidences supporting this hypothesis, especially in vivo, are limited. Here, we took advantage of the identification of the mitochondrial Ca2+ Uniporter (MCU) and tested whether mitochondrial Ca2+ signaling controls neuronal cell fate. We overexpressed MCU both in vitro, in mouse primary cortical neurons, and in vivo, through stereotaxic injection of MCU-coding adenoviral particles in the brain cortex. We first measured mitochondrial Ca2+ uptake using quantitative genetically encoded Ca2+ probes, and we observed that the overexpression of MCU causes a dramatic increase of mitochondrial Ca2+ uptake both at resting and after membrane depolarization. MCU-mediated mitochondrial Ca2+ overload causes alteration of organelle morphology and dysregulation of global Ca2+ homeostasis. Most importantly, MCU overexpression in vivo is sufficient to trigger gliosis and neuronal loss. Overall, we demonstrated that mitochondrial Ca2+ overload is per se sufficient to cause neuronal cell death both in vitro and in vivo, thus highlighting a potential key step in neurodegeneration.
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physiological characterization of a plant mitochondrial calcium Uniporter in vitro and in vivo
Plant Physiology, 2017Co-Authors: Enrico Teardo, Stephan Wagner, Sara De Bortoli, Philippe Fuchs, Luca Carraretto, Elide Formentin, Smrutisanjita Behera, Veronique Larosa, Fiorella Lo Schiavo, Anna RaffaelloAbstract:Over the recent years, several proteins that make up the mitochondrial calcium Uniporter complex (MCUC) mediating Ca2+uptake into the mitochondrial matrix have been identified in mammals, including the channel-forming protein MCU. Although six MCU gene homologs are conserved in the model plant Arabidopsis (Arabidopsis thaliana) in which mitochondria can accumulate Ca2+, a functional characterization of plant MCU homologs has been lacking. Using electrophysiology, we show that one isoform, AtMCU1, gives rise to a Ca2+-permeable channel activity that can be observed even in the absence of accessory proteins implicated in the formation of the active mammalian channel. Furthermore, we provide direct evidence that AtMCU1 activity is sensitive to the mitochondrial calcium Uniporter inhibitors Ruthenium Red and Gd3+, as well as to the Arabidopsis protein MICU, a regulatory MCUC component. AtMCU1 is prevalently expressed in roots, localizes to mitochondria, and its absence causes mild changes in Ca2+ dynamics as assessed by in vivo measurements in Arabidopsis root tips. Plants either lacking or overexpressing AtMCU1 display root mitochondria with altered ultrastructure and show shorter primary roots under restrictive growth conditions. In summary, our work adds evolutionary depth to the investigation of mitochondrial Ca2+ transport, indicates that AtMCU1, together with MICU as a regulator, represents a functional configuration of the plant mitochondrial Ca2+ uptake complex with differences to the mammalian MCUC, and identifies a new player of the intracellular Ca2+ regulation network in plants.
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calcium at the center of cell signaling interplay between endoplasmic reticulum mitochondria and lysosomes
Trends in Biochemical Sciences, 2016Co-Authors: Anna Raffaello, Cristina Mammucari, Rosario Rizzuto, Gaia GherardiAbstract:In recent years, rapid discoveries have been made relating to Ca2+ handling at specific organelles that have important implications for whole-cell Ca2+ homeostasis. In particular, the structures of the endoplasmic reticulum (ER) Ca2+ channels revealed by electron cryomicroscopy (cryo-EM), continuous updates on the structure, regulation, and role of the mitochondrial calcium Uniporter (MCU) complex, and the analysis of lysosomal Ca2+ signaling are milestones on the route towards a deeper comprehension of the complexity of global Ca2+ signaling. In this review we summarize recent discoveries on the regulation of interorganellar Ca2+ homeostasis and its role in pathophysiology.
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The mitochondrial calcium Uniporter (MCU): Molecular identity and physiological roles
Journal of Biological Chemistry, 2013Co-Authors: Maria Patron, Veronica Granatiero, Anna Tosatto, Diego De Stefani, Anna Terrin, Anna Raffaello, Giulia Merli, Giorgia Pallafacchina, Lauren Wright, Cristina MammucariAbstract:The direct measurement of mitochondrial [Ca(2+)] with highly specific probes demonstrated that major swings in organellar [Ca(2+)] parallel the changes occurring in the cytosol and regulate processes as diverse as aerobic metabolism and cell death by necrosis and apoptosis. Despite great biological relevance, insight was limited by the complete lack of molecular understanding. The situation has changed, and new perspectives have emerged following the very recent identification of the mitochondrial Ca(2+) Uniporter, the channel allowing rapid Ca(2+) accumulation across the inner mitochondrial membrane.
Ming-feng Tsai - One of the best experts on this subject based on the ideXlab platform.
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structure and mechanism of the mitochondrial ca 2 Uniporter holocomplex
Nature, 2020Co-Authors: Minrui Fan, Jinru Zhang, Benjamin J. Orlando, Maofu Liao, Chen-wei Tsai, Ming-feng Tsai, Madison X Rodriguez, Liang FengAbstract:Mitochondria take up Ca2+ through the mitochondrial calcium Uniporter complex to regulate energy production, cytosolic Ca2+ signalling and cell death1,2. In mammals, the Uniporter complex (uniplex) contains four core components: the pore-forming MCU protein, the gatekeepers MICU1 and MICU2, and an auxiliary subunit, EMRE, essential for Ca2+ transport3-8. To prevent detrimental Ca2+ overload, the activity of MCU must be tightly regulated by MICUs, which sense changes in cytosolic Ca2+ concentrations to switch MCU on and off9,10. Here we report cryo-electron microscopic structures of the human mitochondrial calcium Uniporter holocomplex in inhibited and Ca2+-activated states. These structures define the architecture of this multicomponent Ca2+-uptake machinery and reveal the gating mechanism by which MICUs control Uniporter activity. Our work provides a framework for understanding regulated Ca2+ uptake in mitochondria, and could suggest ways of modulating Uniporter activity to treat diseases related to mitochondrial Ca2+ overload.
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the conserved aspartate ring of mcu mediates micu1 binding and regulation in the mitochondrial calcium Uniporter complex
eLife, 2019Co-Authors: Charles B Phillips, Chen-wei Tsai, Ming-feng TsaiAbstract:The mitochondrial calcium Uniporter is a Ca2+ channel that regulates intracellular Ca2+ signaling, oxidative phosphorylation, and apoptosis. It contains the pore-forming MCU protein, which possesses a DIME sequence thought to form a Ca2+ selectivity filter, and also regulatory EMRE, MICU1, and MICU2 subunits. To properly carry out physiological functions, the Uniporter must stay closed in resting conditions, becoming open only when stimulated by intracellular Ca2+ signals. This Ca2+-dependent activation, known to be mediated by MICU subunits, is not well understood. Here, we demonstrate that the DIME-aspartate mediates a Ca2+-modulated electrostatic interaction with MICU1, forming an MICU1 contact interface with a nearby Ser residue at the cytoplasmic entrance of the MCU pore. A mutagenesis screen of MICU1 identifies two highly-conserved Arg residues that might contact the DIME-Asp. Perturbing MCU-MICU1 interactions elicits unregulated, constitutive Ca2+ flux into mitochondria. These results indicate that MICU1 confers Ca2+-dependent gating of the Uniporter by blocking/unblocking MCU.
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Proteolytic control of the mitochondrial calcium Uniporter complex.
Proceedings of the National Academy of Sciences of the United States of America, 2017Co-Authors: Chen-wei Tsai, Ping-chieh Pao, Charles B Phillips, Carole Williams, Christopher Miller, Matthew J. Ranaghan, Ming-feng TsaiAbstract:The mitochondrial calcium Uniporter is a Ca2+-activated Ca2+ channel complex mediating mitochondrial Ca2+ uptake, a process crucial for Ca2+ signaling, bioenergetics, and cell death. The Uniporter is composed of the pore-forming MCU protein, the gatekeeping MICU1 and MICU2 subunits, and EMRE, a single-pass membrane protein that links MCU and MICU1 together. As a bridging subunit required for channel function, EMRE could paradoxically inhibit Uniporter complex formation if expressed in excess. Here, we show that mitochondrial mAAA proteases AFG3L2 and SPG7 rapidly degrade unassembled EMRE using the energy of ATP hydrolysis. Once EMRE is incorporated into the complex, its turnover is inhibited >15-fold. Protease-resistant EMRE mutants produce Uniporter subcomplexes that induce constitutive Ca2+ leakage into mitochondria, a condition linked to debilitating neuromuscular disorders in humans. The results highlight the dynamic nature of Uniporter subunit assembly, which must be tightly regulated to ensure proper mitochondrial responses to intracellular Ca2+ signals.
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dual functions of a small regulatory subunit in the mitochondrial calcium Uniporter complex
eLife, 2016Co-Authors: Ming-feng Tsai, Chen-wei Tsai, Charles B Phillips, Matthew J. Ranaghan, Carole Willliams, Christopher MillerAbstract:Mitochondrial Ca(2+) uptake, a process crucial for bioenergetics and Ca(2+) signaling, is catalyzed by the mitochondrial calcium Uniporter. The Uniporter is a multi-subunit Ca(2+)-activated Ca(2+) channel, with the Ca(2+) pore formed by the MCU protein and Ca(2+)-dependent activation mediated by MICU subunits. Recently, a mitochondrial inner membrane protein EMRE was identified as a Uniporter subunit absolutely required for Ca(2+) permeation. However, the molecular mechanism and regulatory purpose of EMRE remain largely unexplored. Here, we determine the transmembrane orientation of EMRE, and show that its known MCU-activating function is mediated by the interaction of transmembrane helices from both proteins. We also reveal a second function of EMRE: to maintain tight MICU regulation of the MCU pore, a role that requires EMRE to bind MICU1 using its conserved C-terminal polyaspartate tail. This dual functionality of EMRE ensures that all transport-competent Uniporters are tightly regulated, responding appropriately to a dynamic intracellular Ca(2+) landscape.
