Global brain ischemia in rats is associated with mitochondrial release and downregulation of Mfn2 in the cerebral cortex, but not the hippocampus.
Klacanova Katarina,Kovalska Maria,Chomova Maria,Pilchova Ivana,Tatarkova Zuzana,Kaplan Peter,Racay Peter
International journal of molecular medicine
Mitochondria are crucial for neuronal cell survival and death through their functions in ATP production and the intrinsic pathway of apoptosis. Mitochondrial dysfunction is considered to play a central role in several serious human diseases, including neurodegenerative diseases, such as Parkinson's and Alzheimer's disease and ischemic neurodegeneration. The aim of the present study was to investigate the impact of transient global brain ischemia on the expression of selected proteins involved in mitochondrial dynamics and mitochondria‑associated membranes. The main foci of interest were the proteins mitofusin 2 (Mfn2), dynamin‑related protein 1 (DRP1), voltage‑dependent anion‑selective channel 1 (VDAC1) and glucose‑regulated protein 75 (GRP75). Western blot analysis of total cell extracts and mitochondria isolated from either the cerebral cortex or hippocampus of experimental animals was performed. In addition, Mfn2 was localized intracellularly by laser scanning confocal microscopy. It was demonstrated that 15‑min ischemia, or 15‑min ischemia followed by 1, 3, 24 or 72 h of reperfusion, was associated with a marked decrease of the Mfn2 protein in mitochondria isolated from the cerebral cortex, but not in hippocampal mitochondria. Moreover, a translocation of the Mfn2 protein to the cytoplasm was documented immediately after global brain ischemia in the neurons of the cerebral cortex by laser scanning confocal microscopy. Mfn2 translocation was followed by decreased expression of Mfn2 during reperfusion. Markedly elevated levels of the VDAC1 protein were also documented in total cell extracts isolated from the hippocampus of rats after 15 min of global brain ischemia followed by 3 h of reperfusion, and from the cerebral cortex of rats after 15 min of global brain ischemia followed by 72 h of reperfusion. The mitochondrial Mfn2 release observed during the early stages of reperfusion may thus represent an important mechanism of mitochondrial dysfunction associated with neuronal dysfunction or death induced by global brain ischemia.
The ALS disease-associated mutant TDP-43 impairs mitochondrial dynamics and function in motor neurons.
Wang Wenzhang,Li Li,Lin Wen-Lang,Dickson Dennis W,Petrucelli Leonard,Zhang Teng,Wang Xinglong
Human molecular genetics
Mutations in TDP-43 lead to familial ALS. Expanding evidence suggests that impaired mitochondrial dynamics likely contribute to the selective degeneration of motor neurons in SOD1-associated ALS. In this study, we investigated whether and how TDP-43 mutations might impact mitochondrial dynamics and function. We demonstrated that overexpression of wild-type TDP-43 resulted in reduced mitochondrial length and density in neurites of primary motor neurons, features further exacerbated by ALS-associated TDP-43 mutants Q331K and M337V. In contrast, suppression of TDP-43 resulted in significantly increased mitochondrial length and density in neurites, suggesting a specific role of TDP-43 in regulating mitochondrial dynamics. Surprisingly, both TDP-43 overexpression and suppression impaired mitochondrial movement. We further showed that abnormal localization of TDP-43 in cytoplasm induced substantial and widespread abnormal mitochondrial dynamics. TDP-43 co-localized with mitochondria in motor neurons and their colocalization was enhanced by ALS associated mutant. Importantly, co-expression of mitochondrial fusion protein mitofusin 2 (Mfn2) could abolish TDP-43 induced mitochondrial dynamics abnormalities and mitochondrial dysfunction. Taken together, these data suggest that mutant TDP-43 impairs mitochondrial dynamics through enhanced localization on mitochondria, which causes mitochondrial dysfunction. Therefore, abnormal mitochondrial dynamics is likely a common feature of ALS which could be potential new therapeutic targets to treat ALS.
Mitochondrial fusion supports increased oxidative phosphorylation during cell proliferation.
Yao Cong-Hui,Wang Rencheng,Wang Yahui,Kung Che-Pei,Weber Jason D,Patti Gary J
Proliferating cells often have increased glucose consumption and lactate excretion relative to the same cells in the quiescent state, a phenomenon known as the Warburg effect. Despite an increase in glycolysis, however, here we show that non-transformed mouse fibroblasts also increase oxidative phosphorylation (OXPHOS) by nearly two-fold and mitochondrial coupling efficiency by ~30% during proliferation. Both increases are supported by mitochondrial fusion. Impairing mitochondrial fusion by knocking down mitofusion-2 (Mfn2) was sufficient to attenuate proliferation, while overexpressing Mfn2 increased proliferation. Interestingly, impairing mitochondrial fusion decreased OXPHOS but did not deplete ATP levels. Instead, inhibition caused cells to transition from excreting aspartate to consuming it. Transforming fibroblasts with the oncogene induced mitochondrial biogenesis, which further elevated OXPHOS. Notably, transformed fibroblasts continued to have elongated mitochondria and their proliferation remained sensitive to inhibition of Mfn2. Our results suggest that cell proliferation requires increased OXPHOS as supported by mitochondrial fusion.
Mitofusin 2 protects cerebellar granule neurons against injury-induced cell death.
Jahani-Asl Arezu,Cheung Eric C C,Neuspiel Margaret,MacLaurin Jason G,Fortin Andre,Park David S,McBride Heidi M,Slack Ruth S
The Journal of biological chemistry
Of the GTPases involved in the regulation of the fusion machinery, mitofusin 2 (Mfn2) plays an important role in the nervous system as point mutations of this isoform are associated with Charcot Marie Tooth neuropathy. Here, we investigate whether Mfn2 plays a role in the regulation of neuronal injury. We first examine mitochondrial dynamics following different modes of injury in cerebellar granule neurons. We demonstrate that neurons exposed to DNA damage or oxidative stress exhibit extensive mitochondrial fission, an early event preceding neuronal loss. The extent of mitochondrial fragmentation and remodeling is variable and depends on the mode and the severity of the death stimuli. Interestingly, whereas mitofusin 2 loss of function significantly induces cell death in the absence of any cell death stimuli, expression of mitofusin 2 prevents cell death following DNA damage, oxidative stress, and K+ deprivation induced apoptosis. More importantly, whereas wild-type Mfn2 and the hydrolysis-deficient mutant of Mfn2 (Mfn2(RasG12V)) function equally to promote fusion and lengthening of mitochondria, the activated Mfn2(RasG12V) mutant shows a significant increase in the protection of neurons against cell death and release of proapoptotic factor cytochrome c. These findings highlight a signaling role for Mfn2 in the regulation of apoptosis that extends beyond its role in mitochondrial fusion.
Mechanism of Activation-Induced Downregulation of Mitofusin 2 in Human Peripheral Blood T Cells.
Dasgupta Asish,Chen Kuang-Hueih,Munk Rachel B,Sasaki Carl Y,Curtis Jessica,Longo Dan L,Ghosh Paritosh
Journal of immunology (Baltimore, Md. : 1950)
Mitofusin 2 (Mfn2), a mitochondrial protein, was shown to have antiproliferative properties when overexpressed. In this article, we show that activation of resting human peripheral blood T cells caused downregulation of Mfn2 levels. This downregulation of Mfn2 was blocked by different inhibitors (mTOR inhibitor rapamycin, PI3K inhibitor LY294002, and Akt inhibitor A443654), producing cells that were arrested in the G0/G1 stage of the cell cycle. Furthermore, the activation-induced downregulation of Mfn2 preceded the entry of the cells into the cell cycle, suggesting that Mfn2 downregulation is a prerequisite for activated T cell entry into the cell cycle. Accordingly, small interfering RNA-mediated knockdown of Mfn2 resulted in increased T cell proliferation. Overexpression of constitutively active AKT resulted in the downregulation of Mfn2, which can be blocked by a proteasome inhibitor. Akt-mediated downregulation of Mfn2 was via the mTORC1 pathway because this downregulation was blocked by rapamycin, and overexpression of wild-type, but not kinase-dead mTOR, caused Mfn2 downregulation. Our data suggested that activation-induced reactive oxygen species production plays an important role in the downregulation of Mfn2. Collectively, our data suggest that the PI3K-AKT-mTOR pathway plays an important role in activation-induced downregulation of Mfn2 and subsequent proliferation of resting human T cells.
Mfn2 Ablation in the Adult Mouse Hippocampus and Cortex Causes Neuronal Death.
Han Song,Nandy Priya,Austria Quillan,Siedlak Sandra L,Torres Sandy,Fujioka Hisashi,Wang Wenzhang,Zhu Xiongwei
It is believed that mitochondrial fragmentation cause mitochondrial dysfunction and neuronal deficits in Alzheimer's disease. We recently reported that constitutive knockout of the mitochondria fusion protein mitofusin2 (Mfn2) in the mouse brain causes mitochondrial fragmentation and neurodegeneration in the hippocampus and cortex. Here, we utilize an inducible mouse model to knock out Mfn2 (Mfn2 iKO) in adult mouse hippocampal and cortical neurons to avoid complications due to developmental changes. Electron microscopy shows the mitochondria become swollen with disorganized and degenerated cristae, accompanied by increased oxidative damage 8 weeks after induction, yet the neurons appear normal at the light level. At later timepoints, increased astrocyte and microglia activation appear and nuclei become shrunken and pyknotic. Apoptosis (Terminal deoxynucleotidyl transferase dUTP nick end labeling, TUNEL) begins to occur at 9 weeks, and by 12 weeks, most hippocampal neurons are degenerated, confirmed by loss of NeuN. Prior to the loss of NeuN, aberrant cell-cycle events as marked by proliferating cell nuclear antigen (PCNA) and pHistone3 were evident in some Mfn2 iKO neurons but do not colocalize with TUNEL signals. Thus, this study demonstrated that Mfn2 ablation and mitochondrial fragmentation in adult neurons cause neurodegeneration through oxidative stress and neuroinflammation in vivo via both apoptosis and aberrant cell-cycle-event-dependent cell death pathways.
MitoQ protects dopaminergic neurons in a 6-OHDA induced PD model by enhancing Mfn2-dependent mitochondrial fusion via activation of PGC-1α.
Xi Ye,Feng Dayun,Tao Kai,Wang Ronglin,Shi Yajun,Qin Huaizhou,Murphy Michael P,Yang Qian,Zhao Gang
Biochimica et biophysica acta. Molecular basis of disease
Parkinson's disease (PD) is characterized by the degeneration of dopaminergic neurons in the substantia nigra compacta (SNc). Although mitochondrial dysfunction is the critical factor in the pathogenesis of PD, the underlying molecular mechanisms are not well understood, and as a result, effective medical interventions are lacking. Mitochondrial fission and fusion play important roles in the maintenance of mitochondrial function and cell viability. Here, we investigated the effects of MitoQ, a mitochondria-targeted antioxidant, in 6-hydroxydopamine (6-OHDA)-induced in vitro and in vivo PD models. We observed that 6-OHDA enhanced mitochondrial fission by decreasing the expression of Mfn1, Mfn2 and OPA1 as well as by increasing the expression of Drp1 in the dopaminergic (DA) cell line SN4741. Notably, MitoQ treatment particularly upregulated the Mfn2 protein and mRNA levels and promoted mitochondrial fusion in the presence of 6-OHDA in a Mfn2-dependent manner. In addition, MitoQ also stabilized mitochondrial morphology and function in the presence of 6-OHDA, which further suppressed the formation of reactive oxygen species (ROS), as well as ameliorated mitochondrial fragmentation and cellular apoptosis. Moreover, the activation of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) was attributed to the upregulation of Mfn2 induced by MitoQ. Consistent with these findings, administration of MitoQ in 6-OHDA-treated mice significantly rescued the decrease of Mfn2 expression and the loss of DA neurons in the SNc. Taken together, our findings suggest that MitoQ protects DA neurons in a 6-OHDA induced PD model by activating PGC-1α to enhance Mfn2-dependent mitochondrial fusion.
Depletion of Mitofusin-2 Causes Mitochondrial Damage in Cisplatin-Induced Neuropathy.
Bobylev Ilja,Joshi Abhijeet R,Barham Mohammed,Neiss Wolfram F,Lehmann Helmar C
Sensory neuropathy is a relevant side effect of the antineoplastic agent cisplatin. Mitochondrial damage is assumed to play a critical role in cisplatin-induced peripheral neuropathy, but the pathomechanisms underlying cisplatin-induced mitotoxicity and neurodegeneration are incompletely understood. In an animal model of cisplatin-induced neuropathy, we determined in detail the extent and spatial distribution of mitochondrial damage during cisplatin treatment. Changes in the total number of axonal mitochondria during cisplatin treatment were assessed in intercostal nerves from transgenic mice that express cyan fluorescent protein. Further, we explored the impact of cisplatin on the expression of nuclear encoded molecules of mitochondrial fusion and fission, including mitofusin-2 (MFN2), optic atrophy 1 (OPA1), and dynamin-related protein 1 (DRP1). Cisplatin treatment resulted in a loss of total mitochondrial mass in axons and in an abnormal mitochondrial morphology including atypical enlargement, increased vacuolization, and loss of cristae. These changes were observed in distal and proximal nerve segments and were more prominent in axons than in Schwann cells. Transcripts of fusion and fission proteins were reduced in distal nerve segments. Significant reduced expression levels of the fusion protein MFN2 was detected in nerves of cisplatin-exposed animals. In summary, we provide for the first time an evidence that cisplatin alters mitochondrial dynamics in peripheral nerves. Loss of MFN2, previously implicated in the pathogenesis of other neurodegenerative diseases, also contributes to the pathogenesis in cisplatin-induced neuropathy.
Mitofusin 2 in POMC neurons connects ER stress with leptin resistance and energy imbalance.
Schneeberger Marc,Dietrich Marcelo O,Sebastián David,Imbernón Mónica,Castaño Carlos,Garcia Ainhoa,Esteban Yaiza,Gonzalez-Franquesa Alba,Rodríguez Ignacio Castrillón,Bortolozzi Analía,Garcia-Roves Pablo M,Gomis Ramon,Nogueiras Ruben,Horvath Tamas L,Zorzano Antonio,Claret Marc
Mitofusin 2 (MFN2) plays critical roles in both mitochondrial fusion and the establishment of mitochondria-endoplasmic reticulum (ER) interactions. Hypothalamic ER stress has emerged as a causative factor for the development of leptin resistance, but the underlying mechanisms are largely unknown. Here, we show that mitochondria-ER contacts in anorexigenic pro-opiomelanocortin (POMC) neurons in the hypothalamus are decreased in diet-induced obesity. POMC-specific ablation of Mfn2 resulted in loss of mitochondria-ER contacts, defective POMC processing, ER stress-induced leptin resistance, hyperphagia, reduced energy expenditure, and obesity. Pharmacological relieve of hypothalamic ER stress reversed these metabolic alterations. Our data establish MFN2 in POMC neurons as an essential regulator of systemic energy balance by fine-tuning the mitochondrial-ER axis homeostasis and function. This previously unrecognized role for MFN2 argues for a crucial involvement in mediating ER stress-induced leptin resistance.
Association between Mitofusin 2 Gene Polymorphisms and Late-Onset Alzheimer's Disease in the Korean Population.
Kim Young Jong,Park Jin Kyung,Kang Won Sub,Kim Su Kang,Han Changsu,Na Hae Ri,Park Hae Jeong,Kim Jong Woo,Kim Young Youl,Park Moon Ho,Paik Jong-Woo
OBJECTIVE:Mitochondrial dysfunction is a prominent and early feature of Alzheimer's disease (AD). The morphologic changes observed in the AD brain could be caused by a failure of mitochondrial fusion mechanisms. The aim of this study was to investigate whether genetic polymorphisms of two genes involved in mitochondrial fusion mechanisms, optic atrophy 1 () and mitofusin 2 (), were associated with AD in the Korean population by analyzing genotypes and allele frequencies. METHODS:One coding single nucleotide polymorphism (SNP) in the , rs1042837, and two coding SNPs in the , rs7624750 and rs9851685, were compared between 165 patients with AD (83 men and 82 women, mean age 72.3±4.41) and 186 healthy control subjects (82 men and 104 women, mean age 76.5±5.98). RESULTS:Among these three SNPs, rs1042837 showed statistically significant differences in allele frequency, and genotype frequency in the co-dominant 1 model and in the dominant model. CONCLUSION:These results suggest that the rs1042837 polymorphism in may be involved in the pathogenesis of AD.
TDP-43 interacts with mitochondrial proteins critical for mitophagy and mitochondrial dynamics.
Davis Stephani A,Itaman Sheed,Khalid-Janney Christopher M,Sherard Justin A,Dowell James A,Cairns Nigel J,Gitcho Michael A
Transactive response DNA-binding protein of 43 kDa (TDP-43) functions as a heterogeneous nuclear ribonucleoprotein and is the major pathological protein in frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis/motor neuron disease (ALS/MND). TDP-43 pathology may also be present as a comorbidity in approximately 20-50% of sporadic Alzheimer's disease cases. In a mouse model of MND, full-length TDP-43 increases association with the mitochondria and blocking the TDP-43/mitochondria interaction ameliorates motor dysfunction. Utilizing a proteomics screen, several mitochondrial TDP-43-interacting partners were identified, including voltage-gated anion channel 1 (VDAC1) and prohibitin 2 (PHB2), a crucial mitophagy receptor. Overexpression of TDP-43 led to an increase in PHB2 whereas TDP-43 knockdown reduced PHB2 expression in cells treated with carbonyl cyanide m-chlorophenylhydrazone (CCCP), an inducer of mitophagy. These results suggest that TDP-43 expression contributes to metabolism and mitochondrial function however we show no change in bioenergetics when TDP-43 is overexpressed and knocked down in HEK293T cells. Furthermore, the fusion protein mitofusin 2 (MFN2) interacts in complex with TDP-43 and selective expression of human TDP-43 in the hippocampus and cortex induced an age-dependent change in Mfn2 expression. Mitochondria morphology is altered in 9-month-old mice selectively expressing TDP-43 in an APP/PS1 background compared with APP/PS1 littermates. We further confirmed TDP-43 localization to the mitochondria using immunogold labeled TDP-43 transmission electron microscopy (TEM) and mitochondrial isolation methods There was no increase in full-length TDP-43 localized to the mitochondria in APP/PS1 mice compared to wild-type (littermates); however, using C- and N-terminal-specific TDP-43 antibodies, the N-terminal (27 kDa, N27) and C-terminal (30 kDa, C30) fragments of TDP-43 are greatly enriched in mitochondrial fractions. In addition, when the mitochondrial peptidase (PMPCA) is overexpressed there is an increase in the N-terminal fragment (N27). These results suggest that TDP-43 processing may contribute to metabolism and mitochondrial function.
PINK1-phosphorylated mitofusin 2 is a Parkin receptor for culling damaged mitochondria.
Chen Yun,Dorn Gerald W
Science (New York, N.Y.)
Senescent and damaged mitochondria undergo selective mitophagic elimination through mechanisms requiring two Parkinson's disease factors, the mitochondrial kinase PINK1 (PTEN-induced putative kinase protein 1; PTEN is phosphatase and tensin homolog) and the cytosolic ubiquitin ligase Parkin. The nature of the PINK-Parkin interaction and the identity of key factors directing Parkin to damaged mitochondria are unknown. We show that the mitochondrial outer membrane guanosine triphosphatase mitofusin (Mfn) 2 mediates Parkin recruitment to damaged mitochondria. Parkin bound to Mfn2 in a PINK1-dependent manner; PINK1 phosphorylated Mfn2 and promoted its Parkin-mediated ubiqitination. Ablation of Mfn2 in mouse cardiac myocytes prevented depolarization-induced translocation of Parkin to the mitochondria and suppressed mitophagy. Accumulation of morphologically and functionally abnormal mitochondria induced respiratory dysfunction in Mfn2-deficient mouse embryonic fibroblasts and cardiomyocytes and in Parkin-deficient Drosophila heart tubes, causing dilated cardiomyopathy. Thus, Mfn2 functions as a mitochondrial receptor for Parkin and is required for quality control of cardiac mitochondria.
Neuronal degeneration and cognitive impairment can be prevented via the normalization of mitochondrial dynamics.
Ishikawa Kaori,Yamamoto Satoshi,Hattori Satoko,Nishimura Naoya,Matsumoto Hirokazu,Miyakawa Tsuyoshi,Nakada Kazuto
Neuronal cells possess a certain degree of plasticity to recover from cell damage. When the stress levels are higher than their plasticity capabilities, neuronal degeneration is triggered. However, the factors correlated to the plasticity capabilities need to be investigated. In this study, we generated a novel mouse model that able to express in an inducible manner a dominant-negative form of MFN2, a mitochondrial fusion factor. We then compared the phenotype of the mice continuously expressing the mutated MFN2 with that of the mice only transiently expressing it. Remarkably, the phenotypes of the group transiently expressing mutant MFN2 could be divided into 3 types: equivalent to what was observed in the continuous expression group, intermediate between the continuous expression group and the control group, and equivalent to the control group. In particular, in the continuous expression group, we observed remarkable hyperactivity and marked cognitive impairments, which were not seen, or were very mild in the transient expression group. These results indicate that abnormal mitochondrial dynamics lead to stress, triggering neuron degeneration; therefore, the neurodegeneration progression can be prevented via the normalization of the mitochondrial dynamics. Since the availability of mouse models suitable for the reproduction of both neurodegeneration and recovery at least partially is very limited, our mouse model can be a useful tool to investigate neuronal plasticity mechanisms and neurodegeneration.
Mul1 restrains Parkin-mediated mitophagy in mature neurons by maintaining ER-mitochondrial contacts.
Puri Rajat,Cheng Xiu-Tang,Lin Mei-Yao,Huang Ning,Sheng Zu-Hang
Chronic mitochondrial stress associates with major neurodegenerative diseases. Recovering stressed mitochondria constitutes a critical step of mitochondrial quality control and thus energy maintenance in early stages of neurodegeneration. Here, we reveal Mul1-Mfn2 pathway that maintains neuronal mitochondrial integrity under stress conditions. Mul1 deficiency increases Mfn2 activity that triggers the first phasic mitochondrial hyperfusion and also acts as an ER-Mito tethering antagonist. Reduced ER-Mito coupling leads to increased cytoplasmic Ca load that activates calcineurin and induces the second phasic Drp1-dependent mitochondrial fragmentation and mitophagy. Overexpressing Mfn2, but not Mfn1, mimics Mul1-deficient phenotypes, while expressing PTPIP51, an ER-Mito anchoring protein, suppresses Parkin-mediated mitophagy. Thus, by regulating mitochondrial morphology and ER-Mito contacts, Mul1-Mfn2 pathway plays an early checkpoint role in maintaining mitochondrial integrity. Our study provides new mechanistic insights into neuronal mitochondrial maintenance under stress conditions, which is relevant to several major neurodegenerative diseases associated with mitochondrial dysfunction and altered ER-Mito interplay.
Oxidative stress-induced mitophagy is suppressed by the miR-106b-93-25 cluster in a protective manner.
Zhang Cheng,Nie Pengqing,Zhou Chunliu,Hu Yue,Duan Suling,Gu Meijia,Jiang Dongxu,Wang Yunfu,Deng Zixin,Chen Jincao,Chen Shi,Wang Lianrong
Cell death & disease
Increased reactive oxygen species levels in the mitochondrial matrix can induce Parkin-dependent mitophagy, which selectively degrades dysfunctional mitochondria via the autolysosome pathway. Phosphorylated mitofusin-2 (MFN2), a receptor of parkin RBR E3 ubiquitin-protein ligase (Parkin), interacts with Parkin to promote the ubiquitination of mitochondrial proteins; meanwhile, the mitophagy receptors Optineurin (OPTN) and nuclear dot protein 52 (NDP52) are recruited to damaged mitochondria to promote mitophagy. However, previous studies have not investigated changes in the levels of OPTN, MFN2, and NDP52 during Parkin-mediated mitophagy. Here, we show that mild and sustained hydrogen peroxide (HO) stimulation induces Parkin-dependent mitophagy accompanied by downregulation of the mitophagy-associated proteins OPTN, NDP52, and MFN2. We further demonstrate that HO promotes the expression of the miR-106b-93-25 cluster and that miR-106b and miR-93 synergistically inhibit the translation of OPTN, NDP52, and MFN2 by targeting their 3' untranslated regions. We further reveal that compromised phosphorylation of MYC proto-oncogene protein (c-Myc) at threonine 58 (T58) (producing an unstable form of c-Myc) caused by reduced nuclear glycogen synthase kinase-3 beta (GSK3β) levels contributes to the promotion of miR-106b-93-25 cluster expression upon HO induction. Furthermore, miR-106b-mediated and miR-93-mediated inhibition of mitophagy-associated proteins (OPTN, MFN2, and NDP52) restrains cell death by controlling excessive mitophagy. Our data suggest that microRNAs (miRNAs) targeting mitophagy-associated proteins maintain cell survival, which is a novel mechanism of mitophagy control. Thus, our findings provide mechanistic insight into how miRNA-mediated regulation alters the biological process of mitophagy.
Resveratrol stimulates mitochondrial fusion by a mechanism requiring mitofusin-2.
Robb Ellen L,Moradi Fereshteh,Maddalena Lucas A,Valente Andrew J F,Fonseca Joao,Stuart Jeffrey A
Biochemical and biophysical research communications
Resveratrol (RES) is a plant-derived stilbene associated with a wide range of health benefits. Mitochondria are a key downstream target of RES, and in some cell types RES promotes mitochondrial biogenesis, altered cellular redox status, and a shift toward oxidative metabolism. Mitochondria exist as a dynamic network that continually remodels via fusion and fission processes, and the extent of fusion is related to cellular redox status and metabolism. We investigated RES's effects on mitochondrial network morphology in several cell lines using a quantitative approach to measure the extent of network fusion. 48 h continuous treatment with 10-20 μM RES stimulated mitochondrial fusion in C2C12 myoblasts, PC3 cancer cells, and mouse embryonic fibroblasts stimulated significant increases in fusion in all instances, resulting in larger and more highly branched mitochondrial networks. Mitofusin-2 (Mfn2) is a key protein facilitating mitochondrial fusion, and its expression was also stimulated by RES. Using Mfn2-null cells we demonstrated that RES's effects on mitochondrial fusion, cellular respiration rates, and cell growth are all dependent upon the presence of Mfn2. Taken together, these results demonstrate that Mfn2 and mitochondrial fusion are affected by RES in ways that appear to relate to RES's known effects on cellular metabolism and growth.
Mitofusin 2 is necessary for striatal axonal projections of midbrain dopamine neurons.
Lee Seungmin,Sterky Fredrik H,Mourier Arnaud,Terzioglu Mügen,Cullheim Staffan,Olson Lars,Larsson Nils-Göran
Human molecular genetics
Mitochondrial dysfunction is implicated in aging and degenerative disorders such as Parkinson's disease (PD). Continuous fission and fusion of mitochondria shapes their morphology and is essential to maintain oxidative phosphorylation. Loss-of-function mutations in PTEN-induced kinase1 (PINK1) or Parkin cause a recessive form of PD and have been linked to altered regulation of mitochondrial dynamics. More specifically, the E3 ubiquitin ligase Parkin has been shown to directly regulate the levels of mitofusin 1 (Mfn1) and Mfn2, two homologous outer membrane large GTPases that govern mitochondrial fusion, but it is not known whether this is of relevance for disease pathophysiology. Here, we address the importance of Mfn1 and Mfn2 in midbrain dopamine (DA) neurons in vivo by characterizing mice with DA neuron-specific knockout of Mfn1 or Mfn2. We find that Mfn1 is dispensable for DA neuron survival and motor function. In contrast, Mfn2 DA neuron-specific knockouts develop a fatal phenotype with reduced weight, locomotor disturbances and death by 7 weeks of age. Mfn2 knockout DA neurons have spherical and enlarged mitochondria with abnormal cristae and impaired respiratory chain function. Parkin does not translocate to these defective mitochondria. Surprisingly, Mfn2 DA neuron-specific knockout mice have normal numbers of midbrain DA neurons, whereas there is a severe loss of DA nerve terminals in the striatum, accompanied by depletion of striatal DA levels. These results show that Mfn2, but not Mfn1, is required for axonal projections of DA neurons in vivo.
Loss of mitofusin 2 links beta-amyloid-mediated mitochondrial fragmentation and Cdk5-induced oxidative stress in neuron cells.
Park Junghyung,Choi Hoonsung,Min Ju-Sik,Kim Bokyung,Lee Sang-Rae,Yun Jong Won,Choi Myung-Sook,Chang Kyu-Tae,Lee Dong-Seok
Journal of neurochemistry
Mitochondrial dysfunction is implicated in age-related degenerative disorders such as Alzheimer's disease (AD). Maintenance of mitochondrial dynamics is essential for regulating mitochondrial function. Aβ oligomers (AβOs), the typical cause of AD, lead to mitochondrial dysfunction and neuronal loss. AβOs have been shown to induce mitochondrial fragmentation, and their inhibition suppresses mitochondrial dysfunction and neuronal cell death. Oxidative stress is one of the earliest hallmarks of AD. Cyclin-dependent kinase 5 (Cdk5) may cause oxidative stress by disrupting the antioxidant system, including Prx2. Cdk5 is also regarded as a modulator of mitochondrial fission; however, a precise mechanistic link between Cdk5 and mitochondrial dynamics is lacking. We estimated mitochondrial morphology and alterations in mitochondrial morphology-related proteins in Neuro-2a (N2a) cells stably expressing the Swedish mutation of amyloid precursor protein (APP), which is known to increase AβO production. We demonstrated that mitochondrial fragmentation by AβOs accompanies reduced mitofusin 1 and 2 (Mfn1/2) levels. Interestingly, the Cdk5 pathway, including phosphorylation of the Prx2-related oxidative stress, has been shown to regulate Mfn1 and Mfn2 levels. Furthermore, Mfn2, but not Mfn1, over-expression significantly inhibits the AβO-mediated cell death pathway. Therefore, these results indicate that AβO-mediated oxidative stress triggers mitochondrial fragmentation via decreased Mfn2 expression by activating Cdk5-induced Prx2 phosphorylation. Mitochondrial fragmentation induced by amyloid-beta oligomer (AβOs) which is generated from the Swedish mutation of amyloid precursor protein (APP) accompanies reduced Mfn1/2 levels. Interestingly, the Cdk5 pathway, including phosphorylation of the Prx2-related oxidative stress, has been shown to regulate Mfn1/2. Furthermore, Mfn2 over-expression significantly inhibits the AβO-mediated neuronal cells death pathway, but not Mfn1 over-expression. Therefore, these results indicate that AβO-mediated oxidative stress triggers mitochondrial fragmentation via decreased Mfn2 expression by activating Cdk5-induced Prx2 phosphorylation. ATP, adenosine triphosphate; Bax, Bcl-2-associated X protein; Bcl-2, B-cell lymphoma 2; Cdk5, Cyclin-dependent kinase; Cyt C, cytochrome C; Mfn2, mitofusin 2; Prx2, peroxiredoxin 2; ROS, reactive oxygen species.
Mitofusin 2-deficiency suppresses cell proliferation through disturbance of autophagy.
Ding Yanhong,Gao Han,Zhao Lifang,Wang Xian,Zheng Ming
Mitofusin2 (Mfn2), a mitochondrial outer membrane protein serving primarily as a mitochondrial fusion protein, has multiple functions in regulating cell biological processes. Defects of Mfn2 were found in diabetes, obesity, and neurodegenerative diseases. In the present study, we found that knockdown of Mfn2 by shRNA led to impaired autophagic degradation, inhibited mitochondrial oxygen consumption rate and cell glycolysis, reduced ATP production, and suppressed cell proliferation. Inhibition of autophagic degradation mimicked Mfn2-deficiency mediated cell proliferation suppression, while enhancement of autophagosome maturation restored the suppressed cell proliferation by Mfn2-deficiency. Thus, our findings revealed the role of Mfn2 in regulating cell proliferation and mitochondrial metabolism, and shed new light on understanding the mechanisms of Mfn2 deficiency related diseases.
Mitofusin 2 is required to maintain mitochondrial coenzyme Q levels.
Mourier Arnaud,Motori Elisa,Brandt Tobias,Lagouge Marie,Atanassov Ilian,Galinier Anne,Rappl Gunter,Brodesser Susanne,Hultenby Kjell,Dieterich Christoph,Larsson Nils-Göran
The Journal of cell biology
Mitochondria form a dynamic network within the cell as a result of balanced fusion and fission. Despite the established role of mitofusins (MFN1 and MFN2) in mitochondrial fusion, only MFN2 has been associated with metabolic and neurodegenerative diseases, which suggests that MFN2 is needed to maintain mitochondrial energy metabolism. The molecular basis for the mitochondrial dysfunction encountered in the absence of MFN2 is not understood. Here we show that loss of MFN2 leads to impaired mitochondrial respiration and reduced ATP production, and that this defective oxidative phosphorylation process unexpectedly originates from a depletion of the mitochondrial coenzyme Q pool. Our study unravels an unexpected and novel role for MFN2 in maintenance of the terpenoid biosynthesis pathway, which is necessary for mitochondrial coenzyme Q biosynthesis. The reduced respiratory chain function in cells lacking MFN2 can be partially rescued by coenzyme Q10 supplementation, which suggests a possible therapeutic strategy for patients with diseases caused by mutations in the Mfn2 gene.
Mitochondrial redox-driven mitofusin 2 S-glutathionylation promotes neuronal necroptosis via disrupting ER-mitochondria crosstalk in cadmium-induced neurotoxicity.
Che Lin,Yang Chuan-Li,Chen Yu,Wu Zi-Li,Du Ze-Bang,Wu Jia-Shen,Gan Cong-Ling,Yan Si-Ping,Huang Jing,Guo Ni-Jun,Lin Yu-Chun,Lin Zhong-Ning
Reactive oxygen species (ROS)-mediated endoplasmic reticulum (ER) stress and mitochondrial dysfunction are known to affect the structural and functional damage in the neural system. Cadmium (Cd) is an environmental contaminant that is widely found in numerous environmental matrices and exhibits potential neurotoxic risk. However, it remains unclear how mitochondrial redox status induces, and whether Cd destabilizes, the ER-mitochondria crosstalk to have a toxic effect on the nervous system. Herein, in our present study, bioinformatics analysis revealed an important role of protein interaction and mitochondrial machinery in brain samples from Alzheimer's disease (AD) patients. Furthermore, we established a neurotoxicity model in vivo and in vitro induced by cadmium chloride (CdCl). We demonstrated that CdCl exposure disrupts the balance in mitochondrial redox represented by enhanced mitochondrial ROS (mitoROS) levels, which enhance mitofusin 2 (Mfn2) S-glutathionylation and interrupt the mitochondria-associated ER membranes (MAMs) for crosstalk between the ER and mitochondria to induce neuronal necroptosis. Mechanistically, it was shown that CdCl exposure significantly enhances the mitochondria-associated degradation (MAD) of Mfn2 via S-glutathionylation, which inhibits Mfn2 localization to the MAMs and subsequently leads to the formation of the RIPK1-RIPK3-p-MLKL complex (a key component of the necrosome) at MAMs, to promote neuronal necroptosis. Furthermore, the glutaredoxin 1 (Grx1) catalyzed and Mfn2 overexpression restored S-glu-Mfn2, MAMs perturbation, necrosome formation, and necroptosis in neurons induced by CdCl exposure in vitro. Moreover, the intervention with antioxidants to reduce mitochondrial redox, such as N-acetyl-l-cysteine (NAC) and mitochondria-targeted antioxidant Mito-TEMPO, reduced the S-glutathionylation of Mfn2 involved in the antagonism of CdCl-induced necroptosis and neurotoxicity in vivo and in vitro. Taken together, our results are the first time to demonstrate that S-glutathionylation of Mfn2 promotes neuronal necroptosis via disruption of ER-mitochondria crosstalk in CdCl-induced neurotoxicity, providing the novel mechanistic insight into how hazardous chemical-induced adverse effects in various organs and tissues could be interpreted by intraorganellar pathways under the control of MAMs components in neurons.
Mitofusin 2, a mitochondria-ER tethering protein, facilitates osteoclastogenesis by regulating the calcium-calcineurin-NFATc1 axis.
Jung Suhan,Kwon Jun-Oh,Kim Min Kyung,Song Min-Kyoung,Kim Bongjun,Lee Zang Hee,Kim Hong-Hee
Biochemical and biophysical research communications
Mitofusin 2 (Mfn2) is a mitochondrial outer membrane protein that participates in tethering mitochondria to the ER. Mitochondria-ER tethering has been demonstrated to play important roles in many cellular activities by regulating homeostasis of metabolites and calcium. Intracellular calcium signaling is crucial for the differentiation of osteoclasts, the bone-resorbing cells. In this study, we investigated whether Mfn2 plays a role in osteoclastogenesis by receptor activator of nuclear factor kappa B (RANKL) in primary cells. We found that RANKL increased Mfn2 expression during osteoclast formation from mouse bone marrow-derived macrophages (BMMs). When Mfn2 expression was suppressed in BMMs by using a siRNA-mediated gene knock-down system, osteoclast differentiation and activity of mature osteoclasts were reduced. Mfn2 knock-down also decreased the RANKL-mediated induction of NFATc1, the key transcription factor for osteoclast gene expression, without affecting c-Fos level. This effect on NFATc1 was associated with decreased calcium oscillation and calcineurin activity in Mfn2-deficient osteoclasts. Taken together, our results indicate that Mfn2 positively contributes to RANKL-induced osteoclast differentiation by regulating the calcium-calcieurin-NFATc1 axis, raising the importance of a previously under-recognized role of mitochondria in osteoclastogenesis.
Critical reappraisal confirms that Mitofusin 2 is an endoplasmic reticulum-mitochondria tether.
Naon Deborah,Zaninello Marta,Giacomello Marta,Varanita Tatiana,Grespi Francesca,Lakshminaranayan Sowmya,Serafini Annalisa,Semenzato Martina,Herkenne Stephanie,Hernández-Alvarez Maria Isabel,Zorzano Antonio,De Stefani Diego,Dorn Gerald W,Scorrano Luca
Proceedings of the National Academy of Sciences of the United States of America
The discovery of the multiple roles of mitochondria-endoplasmic reticulum (ER) juxtaposition in cell biology often relied upon the exploitation of Mitofusin (Mfn) 2 as an ER-mitochondria tether. However, this established Mfn2 function was recently questioned, calling for a critical re-evaluation of Mfn2's role in ER-mitochondria cross-talk. Electron microscopy and fluorescence-based probes of organelle proximity confirmed that ER-mitochondria juxtaposition was reduced by constitutive or acute Mfn2 deletion. Functionally, mitochondrial uptake of Ca released from the ER was reduced following acute Mfn2 ablation, as well as in Mfn2 cells overexpressing the mitochondrial calcium uniporter. Mitochondrial Ca uptake rate and extent were normal in isolated Mfn2 liver mitochondria, consistent with the finding that acute or chronic Mfn2 ablation or overexpression did not alter mitochondrial calcium uniporter complex component levels. Hence, Mfn2 stands as a bona fide ER-mitochondria tether whose ablation decreases interorganellar juxtaposition and communication.
MiR-195 dependent roles of mitofusin2 in the mitochondrial dysfunction of hippocampal neurons in SAMP8 mice.
Zhang Rui,Zhou Huimin,Jiang Lei,Mao Yueran,Cui Ximing,Xie Bing,Cui Dongsheng,Wang Hui,Zhang Qingfu,Xu Shunjiang
Abnormal gene expression, including mRNAs, and microRNAs (miRNA), have been identified in the development of Alzheimer's disease (AD). Although mitofusin2 (mfn2) has been found to be down-regulated in the neurons from hippocampus and cortex in AD patients, little is known about its roles and the regulatory mechanisms in the pathogenesis of AD. This study was performed to investigate the roles of mfn2 protein and its upstream regulatory mechanism in the progression of AD using a senescence accelerated mouse prone-8 (SAMP8) model. The results of quantitative real-time PCR and western blot revealed that mfn2 expression displayed a consistent decrease with aging in the hippocampus of SAMP8 than did age-matched SAMR1 mice. The luciferase activity assay combined with mutational analysis confirmed the binding site of miR-195 to the 3' -untranslated region (3'-UTR) of mfn2 mRNA. Furthermore, miR-195 inhibitor or antigomir induced the higher level expression of mfn2 protein in vitro and in vivo. In addition, exogenous expression of miR-195 decreased the mitochondrial membrane potential (MMP) of the HT-22 cells by targeting mfn2. In conclusion, these results indicated that deregulation of mfn2 might be involved in mitochondrial dysfunction during the progression of AD, and its decreased expression was regulated at least in part by miR-195 in AD mice. The abnormal expression of miR-195 played a potential role in mitochondrial disorder by targeting mfn2 in hippocampus of SAMP8 mice. Therefore, upregulation of mfn2 protein by inhibiting miR-195 might be a potential new therapeutic strategy for treatment of AD.
Mitofusin 2 in Macrophages Links Mitochondrial ROS Production, Cytokine Release, Phagocytosis, Autophagy, and Bactericidal Activity.
Tur Juan,Pereira-Lopes Selma,Vico Tania,Marín Eros A,Muñoz Juan P,Hernández-Alvarez Maribel,Cardona Pere-Joan,Zorzano Antonio,Lloberas Jorge,Celada Antonio
Mitofusin 2 (Mfn2) plays a major role in mitochondrial fusion and in the maintenance of mitochondria-endoplasmic reticulum contact sites. Given that macrophages play a major role in inflammation, we studied the contribution of Mfn2 to the activity of these cells. Pro-inflammatory stimuli such as lipopolysaccharide (LPS) induced Mfn2 expression. The use of the Mfn2 and Mfn1 myeloid-conditional knockout (KO) mouse models reveals that Mfn2 but not Mfn1 is required for the adaptation of mitochondrial respiration to stress conditions and for the production of reactive oxygen species (ROS) upon pro-inflammatory activation. Mfn2 deficiency specifically impairs the production of pro-inflammatory cytokines and nitric oxide. In addition, the lack of Mfn2 but not Mfn1 is associated with dysfunctional autophagy, apoptosis, phagocytosis, and antigen processing. Mfn2 mice fail to be protected from Listeria, Mycobacterium tuberculosis, or LPS endotoxemia. These results reveal an unexpected contribution of Mfn2 to ROS production and inflammation in macrophages.
Mitofusin 2 regulates neutrophil adhesive migration and the actin cytoskeleton.
Zhou Wenqing,Hsu Alan Y,Wang Yueyang,Syahirah Ramizah,Wang Tianqi,Jeffries Jacob,Wang Xu,Mohammad Haroon,Seleem Mohamed N,Umulis David,Deng Qing
Journal of cell science
Neutrophils rely on glycolysis for energy production. How mitochondria regulate neutrophil function is not fully understood. Here, we report that mitochondrial outer membrane protein Mitofusin 2 (MFN2) regulates neutrophil homeostasis and chemotaxis -deficient neutrophils are released from the hematopoietic tissue, trapped in the vasculature in zebrafish embryos, and not capable of chemotaxis. Consistent with this, human neutrophil-like cells that are deficient for MFN2 fail to arrest on activated endothelium under sheer stress or perform chemotaxis on 2D surfaces. Deletion of MFN2 results in a significant reduction of neutrophil infiltration to the inflamed peritoneal cavity in mice. Mechanistically, MFN2-deficient neutrophil-like cells display disrupted mitochondria-ER interaction, heightened intracellular Ca levels and elevated Rac activation after chemokine stimulation. Restoring a mitochondria-ER tether rescues the abnormal Ca levels, Rac hyperactivation and chemotaxis defect resulting from MFN2 depletion. Finally, inhibition of Rac activation restores chemotaxis in MFN2-deficient neutrophils. Taken together, we have identified that MFN2 regulates neutrophil migration via maintaining the mitochondria-ER interaction to suppress Rac activation, and uncovered a previously unrecognized role of MFN2 in regulating cell migration and the actin cytoskeleton.This article has an associated First Person interview with the first authors of the paper.
Mitofusin 2 Regulates Axonal Transport of Calpastatin to Prevent Neuromuscular Synaptic Elimination in Skeletal Muscles.
Wang Luwen,Gao Ju,Liu Jingyi,Siedlak Sandra L,Torres Sandy,Fujioka Hisashi,Huntley Mikayla L,Jiang Yinfei,Ji Haiyan,Yan Tingxiang,Harland Micah,Termsarasab Pichet,Zeng Sophia,Jiang Zhen,Liang Jingjing,Perry George,Hoppel Charles,Zhang Cheng,Li Hu,Wang Xinglong
Skeletal muscles undergo atrophy in response to diseases and aging. Here we report that mitofusin 2 (Mfn2) acts as a dominant suppressor of neuromuscular synaptic loss to preserve skeletal muscles. Mfn2 is reduced in spinal cords of transgenic SOD1 and aged mice. Through preserving neuromuscular synapses, increasing neuronal Mfn2 prevents skeletal muscle wasting in both SOD1 and aged mice, whereas deletion of neuronal Mfn2 produces neuromuscular synaptic dysfunction and skeletal muscle atrophy. Neuromuscular synaptic loss after sciatic nerve transection can also be alleviated by Mfn2. Mfn2 coexists with calpastatin largely in mitochondria-associated membranes (MAMs) to regulate its axonal transport. Genetic inactivation of calpastatin abolishes Mfn2-mediated protection of neuromuscular synapses. Our results suggest that, as a potential key component of a novel and heretofore unrecognized mechanism of cytoplasmic protein transport, Mfn2 may play a general role in preserving neuromuscular synapses and serve as a common therapeutic target for skeletal muscle atrophy.
Neuronal Mitochondria Modulation of LPS-Induced Neuroinflammation.
Harland Micah,Torres Sandy,Liu Jingyi,Wang Xinglong
The Journal of neuroscience : the official journal of the Society for Neuroscience
Neuronal mitochondria dysfunction and neuroinflammation are two prominent pathological features increasingly realized as important pathogenic mechanisms for neurodegenerative diseases. However, little attempt has been taken to investigate the likely interactions between them. Mitofusin2 (Mfn2) is a mitochondrial outer membrane protein regulating mitochondrial fusion, a dynamic process essential for mitochondrial function. To explore the significance of neuronal mitochondria in the regulation of neuroinflammation, male and female transgenic mice with forced overexpression of Mfn2 specifically in neurons were intraperitoneally injected with lipopolysaccharide (LPS), a widely used approach to model neurodegeneration-associated neuroinflammation. Remarkably, LPS-induced lethality was almost completely abrogated in neuronal Mfn2 overexpression mice. Compared with nontransgenic wild-type mice, mice with neuronal Mfn2 overexpression also exhibited alleviated bodyweight loss, behavioral sickness, and myocardial dysfunction. LPS-induced release of IL-1β but not TNF-α was further found greatly inhibited in the CNS of mice with neuronal Mfn2 overexpression, whereas peripheral inflammatory responses in the blood, heart, lung, and spleen remained unchanged. At the cellular and molecular levels, neuronal Mfn2 suppressed the activation of microglia, prevented LPS-induced mitochondrial fragmentation in neurons, and importantly, upregulated the expression of CX3CL1, a unique chemokine constitutively produced by neurons to suppress microglial activation. Together, these results reveal an unrecognized possible role of neuronal mitochondria in the regulation of microglial activation, and propose neuronal Mfn2 as a likely mechanistic linker between neuronal mitochondria dysfunction and neuroinflammation in neurodegeneration. Our study suggests that Mfn2 in neurons contributes to the regulation of neuroinflammation. Based on the remarkable suppression of LPS-induced neuroinflammation and neurodegeneration-associated mitochondrial dysfunction and dynamic abnormalities by neuronal Mfn2, this study centered on Mfn2-mediated neuroinflammation reveals novel molecular mechanisms that are involved in both mitochondrial dysfunction and neuroinflammation in neurodegenerative diseases. The pharmacological targeting of Mfn2 may present a novel treatment for neuroinflammation-associated diseases.
Mitofusin 2 but not mitofusin 1 mediates Bcl-XL-induced mitochondrial aggregation.
Du Mengyan,Yu Si,Su Wenhua,Zhao Mengxin,Yang Fangfang,Liu Yangpei,Mai Zihao,Wang Yong,Wang Xiaoping,Chen Tongsheng
Journal of cell science
Bcl-2 family proteins, as central players of the apoptotic program, participate in regulation of the mitochondrial network. Here, a quantitative live-cell fluorescence resonance energy transfer (FRET) two-hybrid assay was used to confirm the homo-/hetero-oligomerization of mitofusins 2 and 1 (MFN2 and MFN1), and also demonstrate the binding of MFN2 to MFN1 with 1:1 stoichiometry. A FRET two-hybrid assay for living cells co-expressing CFP-labeled Bcl-XL (an anti-apoptotic Bcl-2 family protein encoded by ) and YFP-labeled MFN2 or MFN1 demonstrated the binding of MFN2 or MFN1 to Bcl-XL with 1:1 stoichiometry. Neither MFN2 nor MFN1 bound with monomeric Bax in healthy cells, but both MFN2 and MFN1 bind to punctate Bax (pro-apoptotic Bcl-2 family protein) during apoptosis. Oligomerized Bak (also known as BAK1; a pro-apoptotic Bcl-2 family protein) only associated with MFN1 but not MFN2. Moreover, co-expression of Bcl-XL with MFN2 or MFN1 had the same anti-apoptotic effect as the expression of Bcl-XL alone to staurosporine-induced apoptosis, indicating the Bcl-XL has its full anti-apoptotic ability when complexed with MFN2 or MFN1. However, knockdown of MFN2 but not MFN1 reduced mitochondrial aggregation induced by overexpression of Bcl-XL, indicating that MFN2 but not MFN1 mediates Bcl-XL-induced mitochondrial aggregation.
[Mitochondrial fusion protein Mfn2 and cardiovascular diseases].
Yu Hai-Yi,Guo Yan-Hong,Gao Wei
Sheng li ke xue jin zhan [Progress in physiology]
Mitofusin 2 (Mfn2) is a mitochondrial dynamin-related protein involved in the mitochondrial fusion reaction and is also connected to an altered mitochondrial energy supply. Mfn2 is a signaling molecule, plays an important role in cell proliferation, differentiation and apoptosis, which participates in the pathophysiology of several cardiovascular diseases, such as hypertension, restenosis after angioplasty, atherosclerosis, cardiac hypertrophy, and cardiac oxidative stress injury. Regulating the mitochondria-related metabolism, Mfn2 affects diabetes and insulin resistance pathogenesis. In addition, Mfn2 could be an important biomarker and therapeutic target molecule for cardiovascular diseases.
The soluble form of Bax regulates mitochondrial fusion via MFN2 homotypic complexes.
Hoppins Suzanne,Edlich Frank,Cleland Megan M,Banerjee Soojay,McCaffery J Michael,Youle Richard J,Nunnari Jodi
In mammals, fusion of the mitochondrial outer membrane is controlled by two DRPs, MFN1 and MFN2, that function in place of a single outer membrane DRP, Fzo1 in yeast. We addressed the significance of two mammalian outer membrane fusion DRPs using an in vitro mammalian mitochondrial fusion assay. We demonstrate that heterotypic MFN1-MFN2 trans complexes possess greater efficacy in fusion as compared to homotypic MFN1 or MFN2 complexes. In addition, we show that the soluble form of the proapoptotic Bcl2 protein, Bax, positively regulates mitochondrial fusion exclusively through homotypic MFN2 trans complexes. Together, these data demonstrate functional and regulatory distinctions between MFN1 and MFN2 and provide insight into their unique physiological roles.
The distinctive role of tau and amyloid beta in mitochondrial dysfunction through alteration in Mfn2 and Drp1 mRNA Levels: A comparative study in Drosophila melanogaster.
Abtahi Seyedeh Leila,Masoudi Raheleh,Haddadi Mohammad
Alzheimer's disease (AD) is one of the most common forms of neurodegenerative diseases. Aggregation of Aβ42 and hyperphosphorylated tau are two major hallmarks of AD. Whether different forms of tau (soluble or hyperphosphorylated) or Aβ are the main culprit in the events observed in AD is still under investigation. Here, we examined the effect of wild-type, prone to hyperphosphorylation and hyperphosphorylated tau, and also Aβ42 peptide on the brain antioxidant defense system and two mitochondrial genes, Marf (homologous to human MFN2) and Drp1 involved in mitochondrial dynamics in transgenic Drosophila melanogaster. AD is an age associated disease. Therefore, the activity of antioxidant agents, CAT, SOD, and GSH levels and the mRNA levels of Marf and Drp1 were assessed in different time points of the flies lifespan. Reduction in cognitive function and antioxidant activity was observed in all transgenic flies at any time point. The most and the least effect on the eye phenotype was exerted by hyperphosphorylated tau and Aβ42, respectively. In addition, the most remarkable alteration in Marf and Drp1 mRNA levels was observed in transgenic flies expressing hyperphosphorylated tau when pan neuronal expression of transgenes was applied. However, when the disease causing gene expression was confined to the mushroom body, Marf and Drp1 mRNA levels alteration was more prominent in tau and tau transgenic flies, respectively. In conclusion, in spite of antioxidant deficiency caused by different types of tau and Aβ42, it seems that tau exerts more toxic effect on the eye phenotype and mitochondrial genes regulation (Marf and Drp1). Moreover, different mechanisms seem to be involved in mitochondrial genes dysregulation when Aβ or various forms of tau are expressed.
Loss of Mfn2 results in progressive, retrograde degeneration of dopaminergic neurons in the nigrostriatal circuit.
Pham Anh H,Meng Shuxia,Chu Quynh N,Chan David C
Human molecular genetics
Mitochondria continually undergo fusion and fission, and these dynamic processes play a major role in regulating mitochondrial function. Studies of several genes associated with familial Parkinson's disease (PD) have implicated aberrant mitochondrial dynamics in the disease pathology, but the importance of these processes in dopaminergic neurons remains poorly understood. Because the mitofusins Mfn1 and Mfn2 are essential for mitochondrial fusion, we deleted these genes from a subset of dopaminergic neurons in mice. Loss of Mfn2 results in a movement defect characterized by reduced activity and rearing. In open field tests, Mfn2 mutants show severe, age-dependent motor deficits that can be rescued with L-3,4 dihydroxyphenylalanine. These motor deficits are preceded by the loss of dopaminergic terminals in the striatum. However, the loss of dopaminergic neurons in the midbrain occurs weeks after the onset of these motor and striatal deficits, suggesting a retrograde mode of neurodegeneration. In our conditional knockout strategy, we incorporated a mitochondrially targeted fluorescent reporter to facilitate tracking of mitochondria in the affected neurons. Using an organotypic slice culture system, we detected fragmented mitochondria in the soma and proximal processes of these neurons. In addition, we found markedly reduced mitochondrial mass and transport, which may contribute to the neuronal loss. These effects are specific for Mfn2, as the loss of Mfn1 yielded no corresponding defects in the nigrostriatal circuit. Our findings indicate that perturbations of mitochondrial dynamics can cause nigrostriatal defects and may be a risk factor for the neurodegeneration in PD.
When MFN2 (mitofusin 2) met autophagy: A new age for old muscles.
Sebastián David,Zorzano Antonio
A long-standing quest is to define the mechanisms responsible for the mitochondrial dysfunction and accumulation of damaged mitochondria that occur during aging. Indeed, those defects are considered major contributors to the aging process. We have analyzed the effect of aging on the muscle expression of Mfn2 and the impact of Mfn2 ablation on muscle function. Our findings reveal that Mfn2 is repressed in muscle during aging, and that is a determinant for the inhibition of autophagy, and mitochondrial quality control, which lead to the accumulation of damaged mitochondria.
Mfn2 ablation causes an oxidative stress response and eventual neuronal death in the hippocampus and cortex.
Jiang Sirui,Nandy Priya,Wang Wenzhang,Ma Xiaopin,Hsia Jeffrey,Wang Chunyu,Wang Zhenlian,Niu Mengyue,Siedlak Sandra L,Torres Sandy,Fujioka Hisashi,Xu Ying,Lee Hyoung-Gon,Perry George,Liu Jun,Zhu Xiongwei
BACKGROUND:Mitochondria are the organelles responsible for energy metabolism and have a direct impact on neuronal function and survival. Mitochondrial abnormalities have been well characterized in Alzheimer Disease (AD). It is believed that mitochondrial fragmentation, due to impaired fission and fusion balance, likely causes mitochondrial dysfunction that underlies many aspects of neurodegenerative changes in AD. Mitochondrial fission and fusion proteins play a major role in maintaining the health and function of these important organelles. Mitofusion 2 (Mfn2) is one such protein that regulates mitochondrial fusion in which mutations lead to the neurological disease. METHODS:To examine whether and how impaired mitochondrial fission/fusion balance causes neurodegeneration in AD, we developed a transgenic mouse model using the CAMKII promoter to knockout neuronal Mfn2 in the hippocampus and cortex, areas significantly affected in AD. RESULTS:Electron micrographs of neurons from these mice show swollen mitochondria with cristae damage and mitochondria membrane abnormalities. Over time the Mfn2 cKO model demonstrates a progression of neurodegeneration via mitochondrial morphological changes, oxidative stress response, inflammatory changes, and loss of MAP2 in dendrites, leading to severe and selective neuronal death. In this model, hippocampal CA1 neurons were affected earlier and resulted in nearly total loss, while in the cortex, progressive neuronal death was associated with decreased cortical size. CONCLUSIONS:Overall, our findings indicate that impaired mitochondrial fission and fusion balance can cause many of the neurodegenerative changes and eventual neuron loss that characterize AD in the hippocampus and cortex which makes it a potential target for treatment strategies for AD.
Mfn2 protects dopaminergic neurons exposed to paraquat both in vitro and in vivo: Implications for idiopathic Parkinson's disease.
Zhao Fanpeng,Wang Wenzhang,Wang Chunyu,Siedlak Sandra L,Fujioka Hisashi,Tang Beisha,Zhu Xiongwei
Biochimica et biophysica acta. Molecular basis of disease
Mitochondrial dynamics and quality control play a critical role in the maintenance of mitochondrial homeostasis and function. Pathogenic mutations of many genes associated with familial Parkinson's disease (PD) caused abnormal mitochondrial dynamics, suggesting a likely involvement of disturbed mitochondrial fission/fusion in the pathogenesis of PD. In this study, we focused on the potential role of mitochondrial fission/fusion in idiopathic PD patients and in neuronal cells and animals exposed to paraquat (PQ), a commonly used herbicide and PD-related neurotoxin, as models for idiopathic PD. Significantly increased expression of dynamin-like protein 1 (DLP1) and a trend towards reduced expression of Mfn1 and Mfn2 were noted in the substantia nigra tissues from idiopathic PD cases. Interestingly, PQ treatment led to similar changes in the expression of fission/fusion proteins both in vitro and in vivo which was accompanied by extensive mitochondrial fragmentation and mitochondrial dysfunction. Blockage of PQ-induced mitochondrial fragmentation by Mfn2 overexpression protected neurons against PQ-induced mitochondrial dysfunction in vitro. More importantly, PQ-induced oxidative damage and stress signaling as well as selective loss of dopaminergic (DA) neurons in the substantia nigra and axonal terminals in striatum was also inhibited in transgenic mice overexpressing hMfn2. Overall, our study demonstrated that disturbed mitochondrial dynamics mediates PQ-induced mitochondrial dysfunction and neurotoxicity both in vitro and in vivo and is also likely involved in the pathogenesis of idiopathic PD which make them a promising therapeutic target for PD treatment.
Mfn2 downregulation in excitotoxicity causes mitochondrial dysfunction and delayed neuronal death.
Martorell-Riera Alejandro,Segarra-Mondejar Marc,Muñoz Juan P,Ginet Vanessa,Olloquequi Jordi,Pérez-Clausell Jeús,Palacín Manuel,Reina Manuel,Puyal Julien,Zorzano Antonio,Soriano Francesc X
The EMBO journal
Mitochondrial fusion and fission is a dynamic process critical for the maintenance of mitochondrial function and cell viability. During excitotoxicity neuronal mitochondria are fragmented, but the mechanism underlying this process is poorly understood. Here, we show that Mfn2 is the only member of the mitochondrial fusion/fission machinery whose expression is reduced in in vitro and in vivo models of excitotoxicity. Whereas in cortical primary cultures, Drp1 recruitment to mitochondria plays a primordial role in mitochondrial fragmentation in an early phase that can be reversed once the insult has ceased, Mfn2 downregulation intervenes in a delayed mitochondrial fragmentation phase that progresses even when the insult has ceased. Downregulation of Mfn2 causes mitochondrial dysfunction, altered calcium homeostasis, and enhanced Bax translocation to mitochondria, resulting in delayed neuronal death. We found that transcription factor MEF2 regulates basal Mfn2 expression in neurons and that excitotoxicity-dependent degradation of MEF2 causes Mfn2 downregulation. Thus, Mfn2 reduction is a late event in excitotoxicity and its targeting may help to reduce excitotoxic damage and increase the currently short therapeutic window in stroke.
Regulation of ER-mitochondria contacts by Parkin via Mfn2.
Basso Valentina,Marchesan Elena,Peggion Caterina,Chakraborty Joy,von Stockum Sophia,Giacomello Marta,Ottolini Denis,Debattisti Valentina,Caicci Federico,Tasca Elisabetta,Pegoraro Valentina,Angelini Corrado,Antonini Angelo,Bertoli Alessandro,Brini Marisa,Ziviani Elena
Parkin, an E3 ubiquitin ligase and a Parkinson's disease (PD) related gene, translocates to impaired mitochondria and drives their elimination via autophagy, a process known as mitophagy. Mitochondrial pro-fusion protein Mitofusins (Mfn1 and Mfn2) were found to be a target for Parkin mediated ubiquitination. Mfns are transmembrane GTPase embedded in the outer membrane of mitochondria, which are required on adjacent mitochondria to mediate fusion. In mammals, Mfn2 also forms complexes that are capable of tethering mitochondria to endoplasmic reticulum (ER), a structural feature essential for mitochondrial energy metabolism, calcium (Ca) transfer between the organelles and Ca dependent cell death. Despite its fundamental physiological role, the molecular mechanisms that control ER-mitochondria cross talk are obscure. Ubiquitination has recently emerged as a powerful tool to modulate protein function, via regulation of protein subcellular localization and protein ability to interact with other proteins. Ubiquitination is also a reversible mechanism, which can be actively controlled by opposing ubiquitination-deubiquitination events. In this work we found that in Parkin deficient cells and parkin mutant human fibroblasts, the tether between ER and mitochondria is decreased. We identified the site of Parkin dependent ubiquitination and showed that the non-ubiquitinatable Mfn2 mutant fails to restore ER-mitochondria physical and functional interaction. Finally, we took advantage of an established in vivo model of PD to demonstrate that manipulation of ER-mitochondria tethering by expressing an ER-mitochondria synthetic linker is sufficient to rescue the locomotor deficit associated to an in vivo Drosophila model of PD.
Role of mitofusin 2 (Mfn2) in controlling cellular proliferation.
Chen Kuang-Hueih,Dasgupta Asish,Ding Jinhui,Indig Fred E,Ghosh Paritosh,Longo Dan L
FASEB journal : official publication of the Federation of American Societies for Experimental Biology
It has been reported that Mitofusin2 (Mfn2) inhibits cell proliferation when overexpressed. We wanted to study the role of endogenous Mfn2 in cell proliferation, along with the structural features of Mfn2 that influence its mitochondrial localization and control of cell proliferation. Mfn2-knockdown clones of a B-cell lymphoma cell line BJAB exhibited an increased rate of cell proliferation. A 2-fold increase in cell proliferation was also observed in Mfn2-knockout mouse embryonic fibroblast (MEF) cells as compared with the control wild-type cells, and the proliferative advantage of the knockout MEF cells was blocked on reintroduction of the Mfn2 gene. Mfn2 exerts its antiproliferative effect by acting as an effector molecule of Ras, resulting in the inhibition of the Ras-Raf-ERK signaling pathway. Furthermore, both the N-terminal (aa 1-264) and the C-terminal (aa 265-757) fragments of Mfn2 blocked cell proliferation through distinct mechanisms: the N-terminal-mediated inhibition was due to its interaction with Raf-1, whereas the C-terminal fragment of Mfn2 inhibited cell proliferation by interacting with Ras. The inhibition of proliferation by the N-terminal fragment was independent of its mitochondrial localization. Collectively, our data provide new insights regarding the role of Mfn2 in controlling cellular proliferation.
Mfn2 is Required for Mitochondrial Development and Synapse Formation in Human Induced Pluripotent Stem Cells/hiPSC Derived Cortical Neurons.
Fang Du,Yan Shijun,Yu Qing,Chen Doris,Yan Shirley ShiDu
Mitochondria are essential dynamic organelles for energy production. Mitochondria dynamically change their shapes tightly coupled to fission and fusion. Imbalance of fission and fusion can cause deficits in mitochondrial respiration, morphology and motility. Mfn2 (mitofusin 2), a mitochondrial membrane protein that participates in mitochondrial fusion in mammalian cells, contributes to the maintenance and operation of the mitochondrial network. Due to lack of applicable model systems, the mechanisms and involvement of mitochondria in neurogenesis in human brain cells have not been well explored. Here, by employing the human induced pluripotent stem cells (hiPSCs) differentiation system, we fully characterized mitochondrial development, neurogenesis and synapse formation in hiPSCs-derived cortical neurons. Differentiation of hiPSCs to cortical neurons with extended period demonstrates mature neurophysiology characterization and functional synaptic network formation. Mitochondrial respiration, morphology and motility in the differentiated neurons also exhibit pronounced development during differentiation. Mfn2 knock-down results in deficits in mitochondrial metabolism and network, neurogenesis and synapse formation, while Mfn2 overexpression enhances mitochondrial bioenergetics and functions, and promotes the differentiation and maturation of neurons. Together, our data indicate that Mfn2 is essential for human mitochondrial development in neuronal maturation and differentiation, which will enhance our understanding of the role of Mfn2 in neurogenesis.
MFN2 ameliorates cell apoptosis in a cellular model of Parkinson's disease induced by rotenone.
Yang Yang,Xue Liu-Jun,Xue Xiao,Ou Zhou,Jiang Teng,Zhang Ying-Dong
Experimental and therapeutic medicine
A number of studies indicated that apoptosis, a specific type of programmed cell death, contributed to the loss of dopaminergic neurons during progression of Parkinson's disease (PD). Previously, the authors of the present study demonstrated that apoptosis of dopaminergic neurons was mainly achieved via the mitochondria-mediated apoptosis pathway, however, the precise molecular mechanisms remain to be elucidated. The present study aimed to determine whether mitofusin-2 (MFN2), a mitochondrial protein, participated in the apoptosis of dopaminergic neurons in a cellular model of PD induced by rotenone. The present study demonstrated that the expression of MFN2 was relatively stable following treatment with rotenone. Lentiviral knockdown and overexpression experiments for the first time, to the best of the authors knowledge, revealed that MFN2 prevented rotenone-induced cell death by amelioration of apoptosis. These results revealed a protective role of MFN2 against apoptosis in an model of PD and may be used to establish MFN2 as a potential therapeutic target for the treatment of this disease.
MFN2-related neuropathies: Clinical features, molecular pathogenesis and therapeutic perspectives.
Stuppia Giulia,Rizzo Federica,Riboldi Giulietta,Del Bo Roberto,Nizzardo Monica,Simone Chiara,Comi Giacomo P,Bresolin Nereo,Corti Stefania
Journal of the neurological sciences
Mitofusin 2 (MFN2) is a GTPase dynamin-like protein of the outer mitochondrial membrane, encoded in the nuclear genome by the MFN2 gene located on the short (p) arm of chromosome 1. MFN2 protein is involved in several intracellular pathways, but is mainly involved in a network that has an essential role in several mitochondrial functions, including fusion, axonal transport, interorganellar communication and mitophagy. Mutations in the gene encoding MFN2 are associated with Charcot-Marie-Tooth disease type 2A (CMT2A), a neurological disorder characterized by a wide clinical phenotype that involves the central and peripheral nervous system. Here, we present the clinical, genetic and neuropathological features of human diseases associated with MFN2 mutations. We also report proposed pathogenic mechanisms through which MFN2 mutations likely contribute to the development of neurodegeneration. MFN2-related disorders may occur more frequently than previously considered, and they may represent a paradigm for the study of the defective mitochondrial dynamics that seem to play a significant role in the molecular and cellular pathogenesis of common neurodegenerative diseases; thus they may also lead to the identification of related therapeutic targets.
Mfn2 Overexpression Attenuates MPTP Neurotoxicity In Vivo.
Zhao Fanpeng,Austria Quillan,Wang Wenzhang,Zhu Xiongwei
International journal of molecular sciences
Mitochondrial dysfunction represents a critical event in the pathogenesis of Parkinson's disease (PD). Increasing evidence demonstrates that disturbed mitochondrial dynamics and quality control play an important role in mitochondrial dysfunction in PD. Our previous study demonstrated that MPP induces mitochondrial fragmentation in vitro. In this study, we aimed to assess whether blocking MPTP-induced mitochondrial fragmentation by overexpressing Mfn2 affords neuroprotection in vivo. We found that the significant loss of dopaminergic neurons in the substantia nigra (SN) induced by MPTP treatment, as seen in wild-type littermate control mice, was almost completely blocked in mice overexpressing Mfn2 (hMfn2 mice). The dramatic reduction in dopamine neuronal fibers and dopamine levels in the striatum caused by MPTP administration was also partially inhibited in hMfn2 mice. MPTP-induced oxidative stress and inflammatory response in the SN and striatum were significantly alleviated in hMfn2 mice. The impairment of motor function caused by MPTP was also blocked in hMfn2 mice. Overall, our work demonstrates that restoration of mitochondrial dynamics by Mfn2 overexpression protects against neuronal toxicity in an MPTP-based PD mouse model, which supports the modulation of mitochondrial dynamics as a potential therapeutic target for PD treatment.
Mfn2 deficiency links age-related sarcopenia and impaired autophagy to activation of an adaptive mitophagy pathway.
Sebastián David,Sorianello Eleonora,Segalés Jessica,Irazoki Andrea,Ruiz-Bonilla Vanessa,Sala David,Planet Evarist,Berenguer-Llergo Antoni,Muñoz Juan Pablo,Sánchez-Feutrie Manuela,Plana Natàlia,Hernández-Álvarez María Isabel,Serrano Antonio L,Palacín Manuel,Zorzano Antonio
The EMBO journal
Mitochondrial dysfunction and accumulation of damaged mitochondria are considered major contributors to aging. However, the molecular mechanisms responsible for these mitochondrial alterations remain unknown. Here, we demonstrate that mitofusin 2 (Mfn2) plays a key role in the control of muscle mitochondrial damage. We show that aging is characterized by a progressive reduction in Mfn2 in mouse skeletal muscle and that skeletal muscle Mfn2 ablation in mice generates a gene signature linked to aging. Furthermore, analysis of muscle Mfn2-deficient mice revealed that aging-induced Mfn2 decrease underlies the age-related alterations in metabolic homeostasis and sarcopenia. Mfn2 deficiency reduced autophagy and impaired mitochondrial quality, which contributed to an exacerbated age-related mitochondrial dysfunction. Interestingly, aging-induced Mfn2 deficiency triggers a ROS-dependent adaptive signaling pathway through induction of HIF1α transcription factor and BNIP3. This pathway compensates for the loss of mitochondrial autophagy and minimizes mitochondrial damage. Our findings reveal that Mfn2 repression in muscle during aging is a determinant for the inhibition of mitophagy and accumulation of damaged mitochondria and triggers the induction of a mitochondrial quality control pathway.
Mfn2 ubiquitination by PINK1/parkin gates the p97-dependent release of ER from mitochondria to drive mitophagy.
McLelland Gian-Luca,Goiran Thomas,Yi Wei,Dorval Geneviève,Chen Carol X,Lauinger Nadine D,Krahn Andrea I,Valimehr Sepideh,Rakovic Aleksandar,Rouiller Isabelle,Durcan Thomas M,Trempe Jean-François,Fon Edward A
Despite their importance as signaling hubs, the function of mitochondria-ER contact sites in mitochondrial quality control pathways remains unexplored. Here we describe a mechanism by which Mfn2, a mitochondria-ER tether, gates the autophagic turnover of mitochondria by PINK1 and parkin. Mitochondria-ER appositions are destroyed during mitophagy, and reducing mitochondria-ER contacts increases the rate of mitochondrial degradation. Mechanistically, parkin/PINK1 catalyze a rapid burst of Mfn2 phosphoubiquitination to trigger p97-dependent disassembly of Mfn2 complexes from the outer mitochondrial membrane, dissociating mitochondria from the ER. We additionally demonstrate that a major portion of the facilitatory effect of p97 on mitophagy is epistatic to Mfn2 and promotes the availability of other parkin substrates such as VDAC1. Finally, we reconstitute the action of these factors on Mfn2 and VDAC1 ubiquitination in a cell-free assay. We show that mitochondria-ER tethering suppresses mitophagy and describe a parkin-/PINK1-dependent mechanism that regulates the destruction of mitochondria-ER contact sites.
MFN2 couples glutamate excitotoxicity and mitochondrial dysfunction in motor neurons.
Wang Wenzhang,Zhang Fan,Li Li,Tang Fangqiang,Siedlak Sandra L,Fujioka Hisashi,Liu Yingchao,Su Bo,Pi Yan,Wang Xinglong
The Journal of biological chemistry
Mitochondrial dysfunction plays a central role in glutamate-evoked neuronal excitotoxicity, and mitochondrial fission/fusion dynamics are essential for mitochondrial morphology and function. Here, we establish a novel mechanistic linker among glutamate excitotoxicity, mitochondrial dynamics, and mitochondrial dysfunction in spinal cord motor neurons. Ca(2+)-dependent activation of the cysteine protease calpain in response to glutamate results in the degradation of a key mitochondrial outer membrane fusion regulator, mitofusin 2 (MFN2), and leads to MFN2-mediated mitochondrial fragmentation preceding glutamate-induced neuronal death. MFN2 deficiency impairs mitochondrial function, induces motor neuronal death, and renders motor neurons vulnerable to glutamate excitotoxicity. Conversely, MFN2 overexpression blocks glutamate-induced mitochondrial fragmentation, mitochondrial dysfunction, and/or neuronal death in spinal cord motor neurons both in vitro and in mice. The inhibition of calpain activation also alleviates glutamate-induced excitotoxicity of mitochondria and neurons. Overall, these results suggest that glutamate excitotoxicity causes mitochondrial dysfunction by impairing mitochondrial dynamics via calpain-mediated MFN2 degradation in motor neurons and thus present a molecular mechanism coupling glutamate excitotoxicity and mitochondrial dysfunction.