Mfn2 Deficiency Research Articles

MFN2 Deficiency Impairs Mitochondrial Functions and PPAR Pathway During Spermatogenesis and Meiosis in Mice.

Mitochondria are highly dynamic organelles and their activity is known to be regulated by changes in morphology via fusion and fission events. However, the role of mitochondrial dynamics on cellular differentiation remains largely unknown. Here, we explored the molecular mechanism of mitochondrial fusion during spermatogenesis by generating an Mfn2 (mitofusin 2) conditional knock-out (cKO) mouse model. We found that depletion of MFN2 in male germ cells led to disrupted spermatogenesis and meiosis during which the majority of Mfn2 cKO spermatocytes did not develop to the pachytene stage. We showed that in these Mfn2 cKO spermatocytes, oxidative phosphorylation in the mitochondria was affected. In addition, RNA-Seq analysis showed that there was a significantly altered transcriptome profile in the Mfn2 deficient pachytene (or pachytene-like) spermatocytes, with a total of 262 genes up-regulated and 728 genes down-regulated, compared with wild-type (control) mice. Pathway enrichment analysis indicated that the peroxisome proliferator-activated receptor (PPAR) pathway was altered, and subsequent more detailed analysis showed that the expression of PPAR α and PPAR γ was up-regulated and down-regulated, respectively, in the MFN2 deficient pachytene (or pachytene-like) spermatocytes. We also demonstrated that there were more lipid droplets in the Mfn2 cKO cells than in the control cells. In conclusion, our study demonstrates a novel finding that MFN2 deficiency negatively affects mitochondrial functions and alters PPAR pathway together with lipid metabolism during spermatogenesis and meiosis.

PINK1‐induced phosphorylation of mitofusin 2 at serine 442 causes its proteasomal degradation and promotes cell proliferation in lung cancer and pulmonary arterial hypertension

Impaired mitochondrial fusion, due in part to decreased mitofusin 2 (Mfn2) expression, contributes to unrestricted cell proliferation and apoptosis-resistance in hyperproliferative diseases like pulmonary arterial hypertension (PAH) and non-small cell lung cancer (NSCLC). We hypothesized that Mfn2 levels are reduced due to increased proteasomal degradation of Mfn2 triggered by its phosphorylation at serine 442 (S442) and investigated the potential kinase mediators. Mfn2 expression was decreased and Mfn2 S442 phosphorylation was increased in pulmonary artery smooth muscle cells from PAH patients and in NSCLC cells. Mfn2 phosphorylation was mediated by PINK1 and protein kinase A (PKA), although only PINK1 expression was increased in these diseases. We designed a S442 phosphorylation deficient Mfn2 construct (PD-Mfn2) and a S442 constitutively phosphorylated Mfn2 construct (CP-Mfn2). The effects of these modified Mfn2 constructs on Mfn2 expression and biological function were compared with those of the wildtype Mfn2 construct (WT-Mfn2). WT-Mfn2 increased Mfn2 expression and mitochondrial fusion in both PAH and NSCLC cells resulting in increased apoptosis and decreased cell proliferation. Compared to WT-Mfn2, PD-Mfn2 caused greater Mfn2 expression, suppression of proliferation, apoptosis induction, and cell cycle arrest. Conversely, CP-Mfn2 caused only a small increase in Mfn2 expression and did not restore mitochondrial fusion, inhibit cell proliferation, or induce apoptosis. Silencing PINK1 or PKA, or proteasome blockade using MG132, increased Mfn2 expression, enhanced mitochondrial fusion and induced apoptosis. In a xenotransplantation NSCLC model, PD-Mfn2 gene therapy caused greater tumor regression than did therapy with WT-Mfn2. Mfn2 deficiency in PAH and NSCLC reflects proteasomal degradation triggered by Mfn2-S442 phosphorylation by PINK1 and/or PKA. Inhibiting Mfn2 phosphorylation has potential therapeutic benefit in PAH and lung cancer.

Osteolineage depletion of mitofusin2 enhances cortical bone formation in female mice.

Mitochondria are essential organelles that form highly complex, interconnected dynamic networks inside cells. The GTPase mitofusin 2 (MFN2) is a highly conserved outer mitochondrial membrane protein involved in the regulation of mitochondrial morphology, which can affect various metabolic and signaling functions. The role of mitochondria in bone formation remains unclear. Since MFN2 levels increase during osteoblast (OB) differentiation, we investigated the role of MFN2 in the osteolineage by crossing mice bearing floxed Mfn2 alleles with those bearing Prx-cre to generate cohorts of conditional knock out (cKO) animals. By ex vivo microCT, cKO female mice, but not males, display an increase in cortical thickness at 8, 18, and 30weeks, compared to wild-type (WT) littermate controls. However, the cortical anabolic response to mechanical loading was not different between genotypes. To address how Mfn2 deficiency affects OB differentiation, bone marrow-derived mesenchymal stromal cells (MSCs) from both wild-type and cKO mice were cultured in osteogenic media with different levels of β-glycerophosphate. cKO MSCs show increased mineralization and expression of multiple markers of OB differentiation only at the lower dose. Interestingly, despite showing the expected mitochondrial rounding and fragmentation due to loss of MFN2, cKO MSCs have an increase in oxygen consumption during the first 7days of OB differentiation. Thus, in the early phases of osteogenesis, MFN2 restrains oxygen consumption thereby limiting differentiation and cortical bone accrual during homeostasis in vivo.

Mitofusin 2 in Macrophages Links Mitochondrial ROS Production, Cytokine Release, Phagocytosis, Autophagy, and Bactericidal Activity

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. Mfn2floxed;CreLysM 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.

Deficient Endoplasmic Reticulum-Mitochondrial Phosphatidylserine Transfer Causes Liver Disease

Non-alcoholic fatty liver is the most common liver disease worldwide. Here, we show that the mitochondrial protein mitofusin 2 (Mfn2) protects against liver disease. Reduced Mfn2 expression was detected in liver biopsies from patients with non-alcoholicsteatohepatitis (NASH). Moreover, reduced Mfn2 levels were detected in mouse models of steatosis or NASH, and its re-expression in a NASH mouse model ameliorated the disease. Liver-specific ablation of Mfn2 in mice provoked inflammation, triglycerideaccumulation, fibrosis, and liver cancer. We demonstrate that Mfn2 binds phosphatidylserine (PS) and can specifically extract PS into membrane domains, favoring PS transfer to mitochondria and mitochondrial phosphatidylethanolamine (PE) synthesis. Consequently, hepatic Mfn2 deficiency reduces PS transfer and phospholipid synthesis, leadingto endoplasmic reticulum (ER) stress and the development of a NASH-like phenotype and liver cancer. Ablation of Mfn2 in liver reveals that disruption of ER-mitochondrial PS transfer is a new mechanism involved in the development of liver disease.

Upregulation of Mitofusin2 in neurons slows down skeletal muscles atrophy in ALS disease and aging model

Skeletal muscles undergo atrophy in response to diseases and aging. Mitochondria, the source of cellular energy, were found to be fragmented in experimental models of ALS disease and aging. However, whether and how mitochondrial fragmentation plays a role in the progression of disease and aging is not clear. Mitofusin2 (Mfn2) is a key regulator of mitochondrial fusion which is crucial for the maintenance of mitochondrial morphology. To test whether Mfn2 reduction in motor neurons contributed to the disease progression in SOD1G93A mice or aged mice, we generated transgenic mice overexpressing Mfn2 specifically in neurons under the control of the Thy1.2 promoter (TMFN mice). Mfn2 overexpression was confirmed in spinal cords but not skeletal muscles. By crossing Mfn2 overexpressed mice with SOD1G93A mice, we obtained double transgenic Mfn2/SOD1G93A mice. Interestingly, mitochondrial fragmentation was abolished in motor neurons of Mfn2/SOD1G93A mice even in the disease endstage. Strikingly, characteristic neuromuscular junction (NMJ) denervation, collapse, and dysfunction in SOD1G93A mice were also greatly alleviated in Mfn2/SOD1G93A mice. The dramatic protective effect of neuronal Mfn2 in SOD1G93A mice was further indicated by the greatly delayed symptom onset of more than 60 days and the complete prevention of behavioral deficits at 120 days old, a very late stage immediately before endstage onset.A similar protective effect was observed in aged mice. The Mfn2 neuronal overexpression mice suffered less neuromuscular synaptic dysfunction and skeletal muscle atrophy than their non‐transgenic litter mates even at 22 months old, which in human years is equivalent to the 70s. Taken together, our study provides evidence suggesting that Mfn2 deficiency induced mitochondrial fragmentation in motor neurons is likely an important mechanism leading to NMJ dysfunction and skeletal muscle atrophy in ALS and aging, and that Mfn2 could be a common therapeutic target for skeletal muscle atrophy.Support or Funding InformationThis study is supported by grants from the US NIH (1R01NS097679 and 1R01NS089604), US Alzheimer's Association (AARG‐17‐499682), and US Association for Frontotemporal Degeneration and Alzheimer's Drug Discovery Foundation (20161206).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Melatonin prevents adverse myocardial infarction remodeling via Notch1/Mfn2 pathway

Mitochondrial dysfunction is linked with myocardial infarction (MI), a disorder in which Notch1 has attracted increasing attention. However, the involvement of Notch1 in mitochondrial impairment after an MI is poorly understood, as is the role of mitochondrial fusion-associated protein 2 (Mfn2). Moreover, whether melatonin potentiates the Notch1/Mfn2 pathway in post-MI cardiac damage remains unclear. In our study, small interfering RNAs against Notch1 or Mfn2 and Jagged1 peptide were delivered via intramyocardial injection. At 3 days after these treatments, MI was induced by ligation of the anterior descending branch. We found that this ablation of Notch1 or Mfn2 aggravated post-MI injury, including worsened mitochondrial damage and increased generation of reactive oxygen species (ROS). In contrast, Jagged1 improved mitochondrial structure and function, decreased ROS production and attenuated post-MI injury. Interestingly, though Mfn2 expression was mildly regulated by Notch1 signaling in myocardium, Mfn2 deficiency nearly eliminated the cardioprotection by Jagged1, as evidenced by suppressed cardiac function, aggravated myocardial fibrosis, increased cell apoptosis, worsened mitochondrial impairment and enhanced oxidative stress. These observations revealed that Mfn2 plays an indispensable role in protection against MI-induced injury by Notch1. The mechanism might involve disrupting a damaging cycle of mitochondrial damage and ROS generation. Furthermore, melatonin activated Notch1 signaling and increased Mfn2 expression were reversed by luzindole, a nonselective antagonist of the melatonin receptor. Notably, melatonin attenuated post-MI injury in normal mice, but not in mice deficient in Notch1 or Mfn2. These results demonstrate that melatonin attenuates post-MI injury via the Notch1/Mfn2 pathway in a receptor-dependent manner.

Mfn2 deficiency links age-related sarcopenia and impaired autophagy to activation of an adaptive mitophagy pathway.

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 Affects Embryo Development via Mitochondrial Dysfunction and Apoptosis.

BackgroundGrowth factors, energy sources, and mitochondrial function strongly affect embryo growth and development in vitro. The biological role and prospective significance of the mitofusin gene Mfn2 in the development of preimplantation embryos remain poorly understood. Our goal is to profile the role of Mfn2 in mouse embryos and determine the underlying mechanism of Mfn2 function in embryo development.MethodsWe transfected Mfn2-siRNA into 2-cell fertilized eggs and then examined the expression of Mfn2, the anti-apoptotic protein Bcl-2, and the apoptosis-promoting protein Bax by Western blot. Additionally, we determined the blastocyst formation rate and measured ATP levels, mtDNA levels, mitochondrial membrane potential (ΔΨm), and apoptosis in all of the embryos.ResultsThe results indicate that the Mfn2 and Bcl-2 levels were markedly decreased, whereas Bax levels were increased in the T group (embryos transfected with Mfn2-siRNA) compared with the C group (embryos transfected with control-siRNA). The blastocyst formation rate was significantly decreased in the T group. The ATP content and the relative amounts of mtDNA and cDNA in the T group were significantly reduced compared with the C group. In the T group, ΔΨm and Ca2+ levels were reduced, and the number of apoptotic cells was increased.ConclusionLow in vitro expression of Mfn2 attenuates the blastocyst formation rate and cleavage speed in mouse zygotes and causes mitochondrial dysfunction, as confirmed by the ATP and mtDNA levels and mitochondrial membrane potential. Mfn2 deficiency induced apoptosis through the Bcl-2/Bax and Ca2+ pathways. These findings indicate that Mfn2 could affect preimplantation embryo development through mitochondrial function and cellular apoptosis.

Mitofusin 2-deficiency suppresses cell proliferation through disturbance of autophagy.

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.

Conditional knockout of proximal tubule mitofusin 2 accelerates recovery and improves survival after renal ischemia.

Proximal tubule (PT) cells are critical targets of acute ischemic injury. Elimination of the mitochondrial fusion protein mitofusin 2 (Mfn2) sensitizes PT cells to apoptosis in vitro. However, the role of PT Mfn2 in ischemic AKI in vivo is unknown. To test its role, we evaluated the effects of conditional KO of PT Mfn2 (cKO-PT-Mfn2) on animal survival after transient bilateral renal ischemia associated with severe AKI. Forty-eight hours after ischemia, 28% of control mice survived compared with 86% of cKO-PT-Mfn2 animals (P<0.001 versus control). Although no significant differences in histologic injury score, apoptosis, or necrosis were detected between genotypes, cKO-PT-Mfn2 kidneys exhibited a 3.5-fold increase in cell proliferation restricted to the intrarenal region with Mfn2 deletion. To identify the signals responsible for increased proliferation, primary PT cells with Mfn2 deficiency were subjected to stress by ATP depletion in vitro. Compared with normal Mfn2 expression, Mfn2 deficiency significantly increased PT cell proliferation and persistently activated extracellular signal-regulated kinase 1/2 (ERK1/2) during recovery from stress. Furthermore, stress and Mfn2 deficiency decreased the interaction between Mfn2 and Ras detected by immunoprecipitation, and purified Mfn2 dose-dependently decreased Ras activity in a cell-free assay. Ischemia in vivo also reduced the Mfn2-RAS interaction and increased both RAS and p-ERK1/2 activity in the renal cortical homogenates of cKO-PT-Mfn2 mice. Our results suggest that, in contrast to its proapoptotic effects in vitro, selective PT Mfn2 deficiency accelerates recovery of renal function and enhances animal survival after ischemic AKI in vivo, partly by increasing Ras-ERK-mediated cell proliferation.

Mitofusin 2 deficiency leads to oxidative stress that contributes to insulin resistance in rat skeletal muscle cells

Mitofusin 2 (Mfn2) is a dynamin-like protein anchored in the outer mitochondrial membrane that plays a crucial role in ensuring optimal mitochondrial morphological homeostasis. It has been shown that reduced expression of Mfn2 is associated with insulin resistance, but the mechanism is still unclear. We investigated whether Mfn2 deficiency leads to impaired insulin sensitivity via elevated oxidative stress. L6 skeletal muscle cells were treated with palmitate and Mfn2 expression was repressed by transfection with antisense Mfn2. Levels of antioxidant enzymes, reactive oxygen species (ROS), the phosphorylation of c-Jun N-terminal Kinase (JNK) and nuclear factor-κB (NF-κB) and the mitochondrial membrane potential (Δψm) were measured. The results showed palmitate-induced insulin resistance of skeletal muscle cells was accompanied by Mfn2 repression. Meanwhile, the cells had decreased Δψm and activity of antioxidant enzymes which could increase production of ROS, phosphorylation of JNK and NF-κB. When Mfn2 was up-regulated in palmitate-treated cells, oxidative stress and insulin resistance was alleviated. Furthermore, knock-down of Mfn2 in control cells enhanced oxidative stress. Mfn2 deficiency led to increased superoxide concentration and activation of JNK as well as NF-κB associated with insulin signaling. In conclusion, Mfn2 is a potent repressor for oxidative stress and regulation of Mfn2 expression may prove to be a potential method to circumvent insulin resistance.

Mitochondrial Genome Linearization Is a Causative Factor for Cardiomyopathy in Mice and Drosophila

Mitofusin (Mfn)2 redundantly promotes mitochondrial outer membrane tethering and organelle fusion with Mfn1, and uniquely functions as the mitochondrial receptor for Parkin during PTEN-induced putative kinase 1 (PINK1)-Parkin-mediated mitophagy. Selective deletion of Mfn2 with retention of Mfn1 preserves mitochondrial fusion while rendering damaged mitochondria resistant to normal quality control culling mechanisms. Consequently, neuron and cardiomyocyte-specific Mfn2 gene ablation is associated with accumulation of damaged mitochondria and organ dysfunction. Here, we determined how mitochondrial DNA (mtDNA) damage contributes to cardiomyopathy in Mfn2-deficient hearts. RNA sequencing of Mfn2-deficient hearts revealed increased expression of some nuclear-encoded mitochondrial genes, but mitochondrial-encoded transcripts were not upregulated in parallel and mtDNA content was decreased. Ultra-deep sequencing of mtDNA showed no increase in single nucleotide mutations, but copy number variations representing insertion-deletion (in-del) mutations were induced over time by cardiomyocyte-specific Mfn2 deficiency. Double-strand mtDNA breaks in the form of in-dels were confirmed by polymerase chain reaction, and in the form of linear mitochondrial genomes were identified by southern blot analysis. Linearization of Drosophila cardiomyocyte mtDNA using conditional cardiomyocyte-specific expression of mitochondrial targeted XhoI recapitulated the cardiomyopathy of Mfn2-deficient mouse hearts. This is the first description of mitochondrial genome linearization as a causative factor in cardiomyopathy. One of the consequences of interrupting mitochondrial culling by the PINK1-Mfn2-Parkin mechanism is an increase in mtDNA double-stranded breaks, which adversely impact mitochondrial function and DNA replication.

Mitofusin 2-Containing Mitochondrial-Reticular Microdomains Direct Rapid Cardiomyocyte Bioenergetic Responses Via Interorganelle Ca 2+ Crosstalk

Mitochondrial Ca(2+) uptake is essential for the bioenergetic feedback response through stimulation of Krebs cycle dehydrogenases. Close association of mitochondria to the sarcoplasmic reticulum (SR) may explain efficient mitochondrial Ca(2+) uptake despite low Ca(2+) affinity of the mitochondrial Ca(2+) uniporter. However, the existence of such mitochondrial Ca(2+) microdomains and their functional role are presently unresolved. Mitofusin (Mfn) 1 and 2 mediate mitochondrial outer membrane fusion, whereas Mfn2 but not Mfn1 tethers endoplasmic reticulum to mitochondria in noncardiac cells. To elucidate roles for Mfn1 and 2 in SR-mitochondrial tethering, Ca(2+) signaling, and bioenergetic regulation in cardiac myocytes. Fruit fly heart tubes deficient of the Drosophila Mfn ortholog MARF had increased contraction-associated and caffeine-sensitive Ca(2+) release, suggesting a role for Mfn in SR Ca(2+) handling. Whereas cardiac-specific Mfn1 ablation had no effects on murine heart function or Ca(2+) cycling, Mfn2 deficiency decreased cardiomyocyte SR-mitochondrial contact length by 30% and reduced the content of SR-associated proteins in mitochondria-associated membranes. This was associated with decreased mitochondrial Ca(2+) uptake (despite unchanged mitochondrial membrane potential) but increased steady-state and caffeine-induced SR Ca(2+) release. Accordingly, Ca(2+)-induced stimulation of Krebs cycle dehydrogenases during β-adrenergic stimulation was hampered in Mfn2-KO but not Mfn1-KO myocytes, evidenced by oxidation of the redox states of NAD(P)H/NAD(P)(+) and FADH(2)/FAD. Physical tethering of SR and mitochondria via Mfn2 is essential for normal interorganelle Ca(2+) signaling in the myocardium, consistent with a requirement for SR-mitochondrial Ca(2+) signaling through microdomains in the cardiomyocyte bioenergetic feedback response to physiological stress.

Mitofusin 2 (Mfn2) links mitochondrial and endoplasmic reticulum function with insulin signaling and is essential for normal glucose homeostasis

Mitochondria are dynamic organelles that play a key role in energy conversion. Optimal mitochondrial function is ensured by a quality-control system tightly coupled to fusion and fission. In this connection, mitofusin 2 (Mfn2) participates in mitochondrial fusion and undergoes repression in muscle from obese or type 2 diabetic patients. Here, we provide in vivo evidence that Mfn2 plays an essential role in metabolic homeostasis. Liver-specific ablation of Mfn2 in mice led to numerous metabolic abnormalities, characterized by glucose intolerance and enhanced hepatic gluconeogenesis. Mfn2 deficiency impaired insulin signaling in liver and muscle. Furthermore, Mfn2 deficiency was associated with endoplasmic reticulum stress, enhanced hydrogen peroxide concentration, altered reactive oxygen species handling, and active JNK. Chemical chaperones or the antioxidant N-acetylcysteine ameliorated glucose tolerance and insulin signaling in liver-specific Mfn2 KO mice. This study provides an important description of a unique unexpected role of Mfn2 coordinating mitochondria and endoplasmic reticulum function, leading to modulation of insulin signaling and glucose homeostasis in vivo.

Abstract P230: Orchestration of Cardiac Mitochondrial Biogenesis by Mitofusin2

Mitochondria generate energy and play central roles governing cell signaling and programmed death. As an energy dependant tissue, the heart is more affected by mitochondrial dysfunction than other tissues. Mitochondrial fusion and fission are essential for normal morphology and function. Mitofusins Mfn1 and Mfn2 are the key components to process mitochondrial fusion. Previous data shows that Mfn1 and Mfn2 are functional redundant in skeletal muscle and the drosophila heart tube. But the role for the mitochondrial fusion proteins in mammalian cardiac myocytes is unknown. The functional role of mitofusins in the mammalian heart was investigated by conditional gene deletion of Mfn1 and Mfn2 in mice. Mice with myh6Cre-directed inactivation of Mfn1 exhibited normal cardiac structure and function. In contrast, ablation of Mfn2 in mouse heart caused progressive dilated cardiomyopathy. At 6 weeks old, the cardiac structure and function were normal, but at 16 weeks old, Mfn2 null mice developed severe cardiac dilation and impaired contractile performance at baseline and in response to β-adrenergic stimulation without induction of programmed cell death. Cardiomyocytes from young Mfn2 null mice had normal excitation-contraction coupling, whereas older cardiomyocytes showed decreased fractional shortening with normal calcium cycling. mRNA resequencing of Mfn2 null hearts revealed that cardiac dysfunction was associated with induction of nuclear-encoded mitochondrial genes. Mfn2 deficient cardiac mitochondria displayed normal function of the mitochondrial permeability transition, respiration and expression of respiration chain complexes, but increased mitochondrial oxidant stress. Remarkably, the development of cardiomyopathy paralleled dramatic (60-80%) loss of mitochondria, measured both as the ratio of mitochondrial DNA to nuclear DNA, and as μg protein/mg cardiac protein. Conclusion: 1. Mfn1 and Mfn2 are not redundant in the heart.Mfn1 is dispensable for normal cardiac development and function. 2. Mfn2 deficiency reacts in a dilated cariomyopathy caused not by a failure of mitochondrial function, but by a crisis of mitochondrial biogenesis.

The Charcot–Marie–Tooth type 2A gene product, Mfn2, up-regulates fuel oxidation through expression of OXPHOS system

Mitofusin-2 (Mfn2) is a mitochondrial membrane protein that participates in mitochondrial fusion in mammalian cells and mutations in the Mfn2 gene cause Charcot-Marie-Tooth neuropathy type 2A. Here, we show that Mfn2 loss-of-function inhibits pyruvate, glucose and fatty acid oxidation and reduces mitochondrial membrane potential, whereas Mfn2 gain-of-function increases glucose oxidation and mitochondrial membrane potential. As to the mechanisms involved, we have found that Mfn2 loss-of-function represses nuclear-encoded subunits of OXPHOS complexes I, II, III and V, whereas Mfn2 overexpression induced the subunits of complexes I, IV and V. Obesity-induced Mfn2 deficiency in rat skeletal muscle was also associated with a decrease in the subunits of complexes I, II, III and V. In addition, the effect of Mfn2 overexpression on mitochondrial metabolism was mimicked by a truncated Mfn2 mutant that is inactive as a mitochondrial fusion protein. Our results indicate that Mfn2 triggers mitochondrial energization, at least in part, by regulating OXPHOS expression through signals that are independent of its role as a mitochondrial fusion protein.