Showing papers on "Mitochondrial carrier published in 2013"
TL;DR: It is demonstrated that the efficiency of mitochondria-dependent cell growth depends on cristae shape and, accordingly, RCS assembly emerges as a link between membrane morphology and function.
905 citations
TL;DR: Red redistribution of cardiolipin serves as an ‘eat-me’ signal for the elimination of damaged mitochondria from neuronal cells as well as the engulfment of mitochondria by the autophagic system.
Abstract: Recognition of injured mitochondria for degradation by macroautophagy is essential for cellular health, but the mechanisms remain poorly understood Cardiolipin is an inner mitochondrial membrane phospholipid We found that rotenone, staurosporine, 6-hydroxydopamine and other pro-mitophagy stimuli caused externalization of cardiolipin to the mitochondrial surface in primary cortical neurons and SH-SY5Y cells RNAi knockdown of cardiolipin synthase or of phospholipid scramblase-3, which transports cardiolipin to the outer mitochondrial membrane, decreased the delivery of mitochondria to autophagosomes Furthermore, we found that the autophagy protein microtubule-associated-protein-1 light chain 3 (LC3), which mediates both autophagosome formation and cargo recognition, contains cardiolipin-binding sites important for the engulfment of mitochondria by the autophagic system Mutation of LC3 residues predicted as cardiolipin-interaction sites by computational modelling inhibited its participation in mitophagy These data indicate that redistribution of cardiolipin serves as an 'eat-me' signal for the elimination of damaged mitochondria from neuronal cells
769 citations
TL;DR: The mitochondrial lipid composition, the biosynthesis and the import of mitochondrial lipids as well as the regulation of these processes will be main issues of this review article.
667 citations
TL;DR: It is reported that the integral mitochondrial membrane protein MCUR1 (mitochondrial calcium uniporter regulator 1) binds to the MCU and promotes MCU-dependent calcium uptake to control ATP production and autophagy.
Abstract: Ca(2+) flux across the mitochondrial inner membrane regulates bioenergetics, cytoplasmic Ca(2+) signals and activation of cell death pathways. Mitochondrial Ca(2+) uptake occurs at regions of close apposition with intracellular Ca(2+) release sites, driven by the inner membrane voltage generated by oxidative phosphorylation and mediated by a Ca(2+) selective ion channel (MiCa; ref. ) called the uniporter whose complete molecular identity remains unknown. Mitochondrial calcium uniporter (MCU) was recently identified as the likely ion-conducting pore. In addition, MICU1 was identified as a mitochondrial regulator of uniporter-mediated Ca(2+) uptake in HeLa cells. Here we identified CCDC90A, hereafter referred to as MCUR1 (mitochondrial calcium uniporter regulator 1), an integral membrane protein required for MCU-dependent mitochondrial Ca(2+) uptake. MCUR1 binds to MCU and regulates ruthenium-red-sensitive MCU-dependent Ca(2+) uptake. MCUR1 knockdown does not alter MCU localization, but abrogates Ca(2+) uptake by energized mitochondria in intact and permeabilized cells. Ablation of MCUR1 disrupts oxidative phosphorylation, lowers cellular ATP and activates AMP kinase-dependent pro-survival autophagy. Thus, MCUR1 is a critical component of a mitochondrial uniporter channel complex required for mitochondrial Ca(2+) uptake and maintenance of normal cellular bioenergetics.
382 citations
TL;DR: It is shown that mitochondrial division is spatially linked to nucleoids and that a majority of these nucleoids segregate prior to division, resulting in their distribution into newly generated tips in the mitochondrial network.
Abstract: Mitochondria generate most of the energy used by cells, and they also play key roles in cellular growth, death, and differentiation. They are evolutionarily derived from bacteria and have retained their own DNA and protein translation system, but they are also dependent on the cell for their growth and replication. A significant portion of the outer membrane of a mitochondrion is in contact with the endoplasmic reticulum (ER)—an organelle that is the starting point for the synthesis of secreted proteins, and is also critical for the synthesis of lipids and other organelles. Recent work suggests that mitochondria–ER contact points mark sites of mitochondrial division, but it is unclear exactly how this process occurs. Here, Murley et al. use the budding yeast and model organism Saccharomyces cerevisiae to show that at mitochondrial division sites, a multiprotein complex called ERMES promotes the formation of ER–mitochondrial contact points, while an evolutionarily conserved enzyme, Gem1, antagonizes these contacts to aid mitochondrial segregation. The contact points are found adjacent to nucleoids (which are complexes of mitochondrial DNA and proteins)—an observation suggesting that ER-associated mitochondrial division evolved to help distribute nucleoids between newly formed mitochondria. The present study also reveals a novel role for the conserved protein Gem1 and could lead researchers to reinvestigate the functions of Miro1/2—the equivalent of Gem1 in higher eukaryotes. Miro1/2 is thought to connect mitochondria to motor proteins, which transports them through the cell along microtubules. Dysfunction of Miro1/2 reduces the mobility of mitochondria, and the work of Murley et al. suggests that this could be a consequence of enhanced contacts between mitochondria and the ER.
280 citations
TL;DR: These findings identify Num1 as a key component of a mitochondria–ER–cortex anchor, which is termed “MECA,” that functions in parallel with mitochondrial dynamics to distribute and position the essential mitochondrial network.
Abstract: To elucidate the functional roles of mitochondrial dynamics in vivo, we identified genes that become essential in cells lacking the dynamin-related proteins Fzo1 and Dnm1, which are required for mitochondrial fusion and division, respectively. The screen identified Num1, a cortical protein implicated in mitochondrial division and distribution that also functions in nuclear migration. Our data indicate that Num1, together with Mdm36, forms a physical tether that robustly anchors mitochondria to the cell cortex but plays no direct role in mitochondrial division. Our analysis indicates that Num1-dependent anchoring is essential for distribution of the static mitochondrial network in fzo1 dnm1 cells. Consistently, expression of a synthetic mitochondria–cortex tether rescues the viability of fzo1 dnm1 num1 cells. We find that the cortical endoplasmic reticulum (ER) also is a constituent of the Num1 mitochondria–cortex tether, suggesting an active role for the ER in mitochondrial positioning in cells. Thus, taken together, our findings identify Num1 as a key component of a mitochondria–ER–cortex anchor, which we termed “MECA,” that functions in parallel with mitochondrial dynamics to distribute and position the essential mitochondrial network.
161 citations
TL;DR: The present findings unveil the missing step of the cardiolipin synthesis pathway in mitochondria as well as the flexibile regulation of phospholipid biosynthesis to respond to compromised CDP-DAG synthesis in mitochondia.
138 citations
TL;DR: Two membrane protein assembly factors C3orf1 and TMEM126B are identified, not found in the mature complex, that help this process by putting together two membrane arm subcomplexes of Mitochondrial respiratory complex I.
Abstract: Mitochondrial respiratory complex I is a product of both the nuclear and mitochondrial genomes. The integration of seven subunits encoded in mitochondrial DNA into the inner membrane, their association with 14 nuclear-encoded membrane subunits, the construction of the extrinsic arm from 23 additional nuclear-encoded proteins, iron–sulfur clusters, and flavin mononucleotide cofactor require the participation of assembly factors. Some are intrinsic to the complex, whereas others participate transiently. The suppression of the expression of the NDUFA11 subunit of complex I disrupted the assembly of the complex, and subcomplexes with masses of 550 and 815 kDa accumulated. Eight of the known extrinsic assembly factors plus a hydrophobic protein, C3orf1, were associated with the subcomplexes. The characteristics of C3orf1, of another assembly factor, TMEM126B, and of NDUFA11 suggest that they all participate in constructing the membrane arm of complex I.
129 citations
TL;DR: The ubiquitin-proteasome system persistently removes a fraction of intermembrane space proteins under physiological conditions, acting as a negative regulator in the biogenesis of this class of proteins.
Abstract: Mitochondrial precursor proteins are synthesized in the cytosol and subsequently imported into mitochondria. The import of mitochondrial intermembrane space proteins is coupled with their oxidative folding and governed by the mitochondrial intermembrane space import and assembly (MIA) pathway. The cytosolic steps that precede mitochondrial import are not well understood. We identified a role for the ubiquitin-proteasome system in the biogenesis of intermembrane space proteins. Interestingly, the function of the ubiquitin-proteasome system is not restricted to conditions of mitochondrial protein import failure. The ubiquitin-proteasome system persistently removes a fraction of intermembrane space proteins under physiological conditions, acting as a negative regulator in the biogenesis of this class of proteins. Thus, the ubiquitin-proteasome system plays an important role in determining the levels of proteins targeted to the intermembrane space of mitochondria.
121 citations
TL;DR: It is suggested that the composition of cristae is preserved during fusion of mitochondria and that cristsae with mixed OXPHOS complexes are only slowly and successively formed by restricted diffusion of inner membrane proteins into existing cristAE.
Abstract: Mitochondria are involved in cellular energy supply, signaling and apoptosis. Their ability to fuse and divide provides functional and morphological flexibility and is a key feature in mitochondrial quality maintenance. To study the impact of mitochondrial fusion/fission on the reorganization of inner membrane proteins, oxidative phosphorylation (OXPHOS) complexes in mitochondria of different HeLa cells were tagged with fluorescent proteins (GFP and DsRed-HA), and cells were fused by polyethylene glycol treatment. Redistribution of the tagged OXPHOS complexes was then followed by means of immunoelectron microscopy, two color super-resolution fluorescence microscopy and single molecule tracking. In contrast to outer membrane and matrix proteins, which mix quickly and homogeneously upon mitochondrial fusion, the mixing of inner membrane proteins was decelerated. Our data suggest that the composition of cristae is preserved during fusion of mitochondria and that cristae with mixed OXPHOS complexes are only slowly and successively formed by restricted diffusion of inner membrane proteins into existing cristae. The resulting transitory mosaic composition of the inner mitochondrial membrane illuminates mitochondrial heterogeneity and potentially is linked to local differences in function and membrane potential.
118 citations
TL;DR: It is suggested that Fis1 acts as a mitochondrial receptor in the recruitment of mitochondrial morphology protein in mammalian cells through interaction with TBC1D15, a mammalian homologue of Dnm1.
Abstract: In yeast, C-tail-anchored mitochondrial outer membrane protein Fis1 recruits the mitochondrial-fission-regulating GTPase Dnm1 to mitochondrial fission sites. However, the function of its mammalian homologue remains enigmatic because it has been reported to be dispensable for the mitochondrial recruitment of Drp1, a mammalian homologue of Dnm1. We identified TBC1D15 as a Fis1-binding protein in HeLa cell extracts. Immunoprecipitation revealed that Fis1 efficiently interacts with TBC1D15 but not with Drp1. Bacterially expressed Fis1 and TBC1D15 formed a direct and stable complex. Exogenously expressed TBC1D15 localized mainly in cytoplasm in HeLa cells, but when coexpressed with Fis1 it localized to mitochondria. Knockdown of TBC1D15 induced highly developed mitochondrial network structures similar to the effect of Fis1 knockdown, suggesting that the TBC1D15 and Fis1 are associated with the regulation of mitochondrial morphology independently of Drp1. These data suggest that Fis1 acts as a mitochondrial receptor in the recruitment of mitochondrial morphology protein in mammalian cells.
TL;DR: It is suggested that mathematical modeling of mitochondria that incorporates accurate information about the organelle's internal architecture can lead to a better understanding of its diverse functions.
TL;DR: This work identifies two novel proteins that play a role in mitochondrial lipid homeostasis and describes a role of Mdm10 in ergosterol trafficking.
Abstract: Summary The yeast mitochondrial outer membrane (MOM) protein Mdm10 is involved in at least three different processes: (1) association of mitochondria with the endoplasmic reticulum and mitochondrial lipid homeostasis (2) membrane assembly of MOM proteins, and (3) inheritance and morphogenesis of mitochondria. To decipher the precise role of Mdm10 in mitochondrial function, we screened for high-copy suppressors of the severe growth defect of the mdm10Δ mutant. We identified two novel mitochondrial proteins (open reading frames YOR228c and YLR253w) that we named Mdm10 complementing protein (Mcp) 1 and Mcp2. Overexpression of Mcp1 or Mcp2 restores the alterations in morphology and stability of respiratory chain complexes of mitochondria devoid of Mdm10, but the observed defect in assembly of MOM proteins is not rescued. Lipid analysis demonstrates that elevated levels of Mcp1 and Mcp2 restore the alterations in mitochondrial phospholipid and ergosterol homeostasis in cells lacking Mdm10. Collectively, this work identifies two novel proteins that play a role in mitochondrial lipid homeostasis and describes a role of Mdm10 in ergosterol trafficking.
TL;DR: The findings behind the recent claim that OPA1 mediates adrenergic control of lipolysis by functioning as a cytosolic A-kinase anchoring protein (AKAP) on the hemimembrane that envelops the lipid droplet are reviewed.
TL;DR: It is proposed that Nm23-H4 acts as a lipid-dependent mitochondrial switch with dual function in phosphotransfer serving local GTP supply and cardiolipin transfer for apoptotic signaling and putative other functions.
TL;DR: The contribution of the cell cortex protein, Num1, to mitochondrial partitioning in yeast is studied to provide new insights into the coupling of mitochondrial dynamics, immobilization, and retention during inheritance.
Abstract: Summary During the cell cycle each organelle has to be faithfully partitioned to the daughter cells. However, the mechanisms controlling organellar inheritance remain poorly understood. We studied the contribution of the cell cortex protein, Num1, to mitochondrial partitioning in yeast. Live-cell microscopy revealed that Num1 is required for attachment of mitochondria to the cell cortex and retention in mother cells. Electron tomography of anchoring sites revealed plasma membrane invaginations directly contacting the mitochondrial outer membrane. Expression of chimeric plasma membrane tethers rescued mitochondrial fission defects in Δ num1 and Δ mdm36 mutants. These findings provide new insights into the coupling of mitochondrial dynamics, immobilization, and retention during inheritance.
TL;DR: A novel role for Pic2 in copper import into mitochondria is proposed and for the first time a mitochondrial carrier family protein capable of importing copper into the matrix is identified.
TL;DR: Interestingly, RNAi-mediated ablation of three outer membrane proteins of unknown function resulted in a collapse of the network-like mitochondrion of procyclic cells and for the first time identified factors that control mitochondrial shape in T. brucei.
TL;DR: The current understanding of how activating and inhibitory interactions within this family lead to the activation and oligomerization of MOMP effectors Bax and Bak, which result in membrane permeabilization is discussed.
Abstract: Although mitochondria are usually considered as supporters of life, they are also involved in cellular death. Mitochondrial outer membrane permeabilization (MOMP) is a crucial event during apoptosis because it causes the release of proapoptotic factors from the mitochondrial intermembrane space to the cytosol. MOMP is mainly controlled by the Bcl-2 family of proteins, which consists of both proapoptotic and antiapoptotic members. We discuss the current understanding of how activating and inhibitory interactions within this family lead to the activation and oligomerization of MOMP effectors Bax and Bak, which result in membrane permeabilization. The order of events leading to MOMP is then highlighted step by step, emphasizing recent discoveries regarding the formation of Bax/Bak pores on the outer mitochondrial membrane. Besides the Bcl-2 proteins, the mitochondrial organelle contributes to and possibly regulates MOMP, because mitochondrial resident proteins and membrane lipids are prominently involved in the process.
TL;DR: It is reported that the MIA pathway is involved in the transport of mitochondrial inner membrane protein Tim22, thereby broadening the known functions of MIA to the biogenesis of inner membrane proteins.
Abstract: The mitochondrial intermembrane space assembly (MIA) pathway is generally considered to be dedicated to the redox-dependent import and biogenesis of proteins localized to the intermembrane space of mitochondria. The oxidoreductase Mia40 is a central component of the pathway responsible for the transfer of disulfide bonds to intermembrane space precursor proteins, causing their oxidative folding. Here we present the first evidence that the function of Mia40 is not restricted to the transport and oxidative folding of intermembrane space proteins. We identify Tim22, a multispanning membrane protein and core component of the TIM22 translocase of inner membrane, as a protein with cysteine residues undergoing oxidation during Tim22 biogenesis. We show that Mia40 is involved in the biogenesis and complex assembly of Tim22. Tim22 forms a disulfide-bonded intermediate with Mia40 upon import into mitochondria. Of interest, Mia40 binds the Tim22 precursor also via noncovalent interactions. We propose that Mia40 not only is responsible for disulfide bond formation, but also assists the Tim22 protein in its integration into the inner membrane of mitochondria.
TL;DR: A finding that the Ant-induced DCD is suppressed by reduced cytosolic protein synthesis suggests a proteostatic crosstalk between mitochondria and the cytosol, which may play an important role for cell survival during aging.
Abstract: Mitochondria are intracellular organelles involved in ATP synthesis, apoptosis, calcium signaling, metabolism, and the synthesis of critical metabolic cofactors. Mitochondrial dysfunction is associated with age-related degenerative diseases. How mitochondrial dysfunction causes cell degeneration is not well understood. Recent studies have shown that mutations in the adenine nucleotide translocase (Ant) cause aging-dependent degenerative cell death (DCD) in yeast, which is sequentially manifested by inner membrane stress, mitochondrial DNA (mtDNA) loss, and progressive loss of cell viability. Ant is an abundant protein primarily involved in ADP/ATP exchange across the mitochondrial inner membrane. It also mediates basal proton leak and regulates the mitochondrial permeability transition pore. Missense mutations in the human Ant1 cause several degenerative diseases which are commonly manifested by fractional mtDNA deletions. Multiple models have been proposed to explain the Ant1-induced pathogenesis. Studies from yeast have suggested that in addition to altered nucleotide transport properties, the mutant proteins cause a global stress on the inner membrane. The mutant proteins likely interfere with general mitochondrial biogenesis in a dominant-negative manner, which secondarily destabilizes mtDNA. More recent work revealed that the Ant-induced DCD is suppressed by reduced cytosolic protein synthesis. This finding suggests a proteostatic crosstalk between mitochondria and the cytosol, which may play an important role for cell survival during aging.
TL;DR: The import of mitochondrial precursor proteins is summarized and four different systems for protein import, processing and assembly show further connections to the respiratory chain.
01 Jan 2013
TL;DR: Mitochondrial biogenesis relies on the transport of hundreds of different precursor proteins from the cytosol into the different mitochondrial subcompartments, where the presequences are often removed by the matrix processing peptidase.
Abstract: Mitochondrial biogenesis relies on the transport of hundreds of different precursor proteins from the cytosol into the different mitochondrial subcompartments. Proteins destined to the mitochondrial matrix contain aminoterminal matrix targeting sequences, also called presequences, which are recognized by cytosol-exposed receptors on the mitochondrial outer membrane. Translocation complexes in the outer membrane, the TOM complex, and in the inner membrane, the TIM23 complex, then thread these precursor proteins into the matrix, where the presequences are often removed by the matrix processing peptidase. Proteins of the outer membrane, the intermembrane space, and the inner membrane can contain internal targeting signals which allow them to embark on specific sorting routes to reach their final location.
TL;DR: High-resolution fluorescence mapping of a channel-facing transmembrane segment (TMS2) of Tim23 from Saccharomyces cerevisiae shows how the energized state of the membrane drives functionally relevant structural dynamics in membrane proteins coupled to processes such as channel gating.
Abstract: Tim23, the central subunit of the TIM23 protein-translocation complex, forms a voltage-gated channel in the mitochondrial inner membrane (MIM), an energy-conserving membrane that generates a proton-motive force to drive vital processes. Using high-resolution fluorescence mapping of a channel-facing transmembrane segment (TMS2) of Tim23 from Saccharomyces cerevisiae, we demonstrate that changes in the energized state of the MIM cause marked structural alterations in the channel region. In an energized membrane, TMS2 forms a continuous α-helix that is inaccessible to the aqueous intermembrane space (IMS). Upon depolarization, the helical periodicity of TMS2 is disrupted, and the channel becomes exposed to the IMS. Kinetic measurements confirm that changes in TMS2 conformation coincide with depolarization. These results reveal how the energized state of the membrane drives functionally relevant structural dynamics in membrane proteins coupled to processes such as channel gating.
TL;DR: Tom70 and Djp1 are identified as crucial players in the biogenesis of Mim1, providing a unique case of specificity between a cochaperone and its substrate protein.
Abstract: A special group of mitochondrial outer membrane proteins spans the membrane once, exposing soluble domains to both sides of the membrane. These proteins are synthesized in the cytosol and then inserted into the membrane by an unknown mechanism. To identify proteins that are involved in the biogenesis of the single-span model protein Mim1, we performed a high-throughput screen in yeast. Two interesting candidates were the cytosolic cochaperone Djp1 and the mitochondrial import receptor Tom70. Our results indeed demonstrate a direct interaction of newly synthesized Mim1 molecules with Tom70. We further observed lower steady-state levels of Mim1 in mitochondria from djp1Δ and tom70 tom71Δ cells and massive mislocalization of overexpressed GFP-Mim1 to the endoplasmic reticulum in the absence of Djp1. Importantly, these phenotypes were observed specifically for the deletion of DJP1 and were not detected in mutant cells lacking any of the other cytosolic cochaperones of the Hsp40 family. Furthermore, the djp1Δ tom70Δ tom71Δ triple deletion resulted in a severe synthetic sick/lethal growth phenotype. Taking our results together, we identified Tom70 and Djp1 as crucial players in the biogenesis of Mim1. Moreover, the involvement of Djp1 provides a unique case of specificity between a cochaperone and its substrate protein.
TL;DR: It is found that the intramitochondrial pool of p53 is located in the intermembrane space and the matrix, and mitochondrial p53 localized in the matrix promotes the assembly of F₁F₀-ATP synthase.
Abstract: We and others previously reported that endogenous p53 can be located at mitochondria in the absence of stress, suggesting that p53 has a role in the normal physiology of this organelle The aim of this study was to characterize in unstressed cells the intramitochondrial localization of p53 and identify new partners and functions of p53 in mitochondria We find that the intramitochondrial pool of p53 is located in the intermembrane space and the matrix Of note, unstressed HCT116 p53+/+ cells simultaneously show increased O₂ consumption and decreased mitochondrial superoxide production compared with their p53-null counterpart This data was confirmed by stable H1299 cell lines expressing low levels of p53 specifically targeted to the matrix Using immunoprecipitation and mass spectrometry, we identified the oligomycin sensitivity-conferring protein (OSCP), a subunit of the F₁F₀-ATP synthase complex, as a new partner of endogenous p53, specifically interacting with p53 localized in the matrix Interestingly, this interaction seems implicated in mitochondrial p53 localization Moreover, p53 localized in the matrix promotes the assembly of F₁F₀-ATP synthase Taking into account that deregulations of mitochondrial respiration and reactive oxygen species production are tightly linked to cancer development, we suggest that mitochondrial p53 may be an important regulator of normal mitochondrial and cellular physiology, potentially exerting tumor suppression activity inside mitochondria
TL;DR: It is demonstrated that Higd-1a inhibits Opa1 cleavage and is required for mitochondrial fusion by virtue of its interaction with Opa2, a key element in fusion of the inner membrane.
Abstract: The activity and morphology of mitochondria are maintained by dynamic fusion and fission processes regulated by a group of proteins residing in, or attached to, their inner and outer membranes. Hypoxia-induced gene domain protein-1a (Higd-1a)/HIMP1-a/HIG1, a mitochondrial inner membrane protein, plays a role in cell survival under hypoxic conditions. In the present study, we showed that Higd-1a depletion resulted in mitochondrial fission, depletion of mtDNA, disorganization of cristae, and growth retardation. We demonstrated that Higd-1a functions by specifically binding to Optic atrophy 1 (Opa1), a key element in fusion of the inner membrane. In the absence of Higd-1a, Opa1 was cleaved, resulting in the loss of its long isoforms and accumulation of small soluble forms. The small forms of Opa1 do not interact with Higd-1a, suggesting that a part of Opa1 in or proximal to the membrane is required for that interaction. Opa1 cleavage, mitochondrial fission, and cell death induced by dissipation of the mitochondrial membrane potential were significantly inhibited by ectopic expression of Higd-1a. Furthermore, growth inhibition due to Higd-1a depletion could be overcome by overexpression of a noncleavable form of Opa1. Collectively, our observations demonstrate that Higd-1a inhibits Opa1 cleavage and is required for mitochondrial fusion by virtue of its interaction with Opa1.
TL;DR: The TIM23 complex is altered in mitochondria lacking the IMP subunit Imp1 although none of the TIM23 components contains a bipartite presequence, revealing carboxy-terminal processing as a new mechanism in the biogenesis of the mitochondrial inner membrane.
Abstract: Many proteins destined for the inner mitochondrial membrane are imported via amino-terminal targeting sequences that are subsequently cleaved. Ieva et al. identify an alternative mechanism for mitochondrial protein import that depends on a carboxyl-terminal targeting sequence.
TL;DR: It is suggested that a monomer and the N-terminal 12 amino acids are necessary for mitochondrial localization ofDJ-1 mutants and that conformation change induced by C106 oxidation or by E18 mutation leads to translocation of DJ-1 into mitochondria.
Abstract: DJ-1 is a novel oncogene and also a causative gene for familial Parkinson’s disease (park7). DJ-1 has multiple functions that include transcriptional regulation, anti-oxidative reaction and chaperone and mitochondrial regulation. Mitochondrial dysfunction is observed in DJ-1-knockout mice and fry, and mitochondrial DJ-1 is more protective against oxidative stress-induced cell death. Although translocation of DJ-1 into mitochondria is enhanced by oxidative stress that leads to oxidation of cysteine 106 (C106) of DJ-1, the characteristics of mitochondrial DJ-1 and the mechanism by which DJ-1 is translocated into mitochondria are poorly understood. In this study, immunostaining, co-immunoprecipitation, cell fractionation and pull-down experiments showed that mutants of glutamine 18 (E18) DJ-1 are localized in mitochondria and do not make homodimers. Likewise, DJ-1 with mutations of two cysteines located in the dimer interface, C46S and C53A, and pathogenic mutants, M26I and L166P DJ-1, were found to be localized in mitochondria and not to make homodimers. Mutant DJ-1 harboring both E18A and C106S, in which C106 is not oxidized, was also localized in mitochondria, indicating that oxidation of C106 is important but not essential for mitochondrial localization of DJ-1. It should be noted that E18A DJ-1 was translocated from mitochondria to the cytoplasm when mitochondrial membrane potential was reduced by treatment of cells with CCCP, an uncoupler of the oxidative phosphorylation system in mitochondria. Furthermore, deletion or substitution of the N-terminal 12 amino acids in DJ-1 resulted in re-localization of E18A, M26I and L166P DJ-1 from mitochondria into the cytoplasm. These findings suggest that a monomer and the N-terminal 12 amino acids are necessary for mitochondrial localization of DJ-1 mutants and that conformation change induced by C106 oxidation or by E18 mutation leads to translocation of DJ-1 into mitochondria.
TL;DR: It is determined that cells defective in mitochondrial inheritance give rise to dead buds that go through cytokinesis normally, showing no evidence of a proposed cell-cycle mitochondrial inheritance checkpoint.