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Gerald W Dorn - One of the best experts on this subject based on the ideXlab platform.

  • the tethering function of mitofusin2 controls osteoclast differentiation by modulating the ca2 nfatc1 axis
    Journal of Biological Chemistry, 2020
    Co-Authors: Anna Ballard, Gerald W Dorn, Rong Zeng, Allahdad Zarei, Christine Shao, Linda Cox, Hui Yan, Antonietta Franco, Roberta X Faccio
    Abstract:

    Dynamic regulation of the mitochondrial network by mitofusins (MFNs) modulates energy production, cell survival, and many intracellular signaling events, including calcium handling. However, the relative importance of specific mitochondrial functions and their dependence on MFNs vary greatly among cell types. Osteoclasts have many mitochondria, and increased mitochondrial biogenesis and oxidative phosphorylation enhance bone resorption, but little is known about the mitochondrial network or MFNs in osteoclasts. Because expression of each MFN isoform increases with osteoclastogenesis, we conditionally deleted MFN1 and MFN2 (double conditional KO (dcKO)) in murine osteoclast precursors, finding that this increased bone mass in young female mice and abolished osteoclast precursor differentiation into mature osteoclasts in vitro. Defective osteoclastogenesis was reversed by overexpression of MFN2 but not MFN1; therefore, we generated mice lacking only MFN2 in osteoclasts. MFN2-deficient female mice had increased bone mass at 1 year and resistance to Receptor Activator of NF-κB Ligand (RANKL)-induced osteolysis at 8 weeks. To explore whether MFN-mediated tethering or mitophagy is important for osteoclastogenesis, we overexpressed MFN2 variants defective in either function in dcKO precursors and found that, although mitophagy was dispensable for differentiation, tethering was required. Because the master osteoclastogenic transcriptional regulator nuclear factor of activated T cells 1 (NFATc1) is calcium-regulated, we assessed calcium release from the endoplasmic reticulum and store-operated calcium entry and found that the latter was blunted in dcKO cells. Restored osteoclast differentiation by expression of intact MFN2 or the mitophagy-defective variant was associated with normalization of store-operated calcium entry and NFATc1 levels, indicating that MFN2 controls mitochondrion–endoplasmic reticulum tethering in osteoclasts.

  • Restoring mitofusin balance prevents axonal degeneration in a Charcot-Marie-Tooth type 2A model
    The Journal of clinical investigation, 2019
    Co-Authors: Yueqin Zhou, Antonietta Franco, Sharon Carmona, A.k.m.g. Muhammad, Shaughn Bell, Jesse Landeros, Michael Vazquez, Gerald W Dorn
    Abstract:

    Mitofusin-2 (MFN2) is a mitochondrial outer-membrane protein that plays a pivotal role in mitochondrial dynamics in most tissues, yet mutations in MFN2, which cause Charcot-Marie-Tooth disease type 2A (CMT2A), primarily affect the nervous system. We generated a transgenic mouse model of CMT2A that developed severe early onset vision loss and neurological deficits, axonal degeneration without cell body loss, and cytoplasmic and axonal accumulations of fragmented mitochondria. While mitochondrial aggregates were labeled for mitophagy, mutant MFN2 did not inhibit Parkin-mediated degradation, but instead had a dominant negative effect on mitochondrial fusion only when MFN1 was at low levels, as occurs in neurons. Finally, using a transgenic approach, we found that augmenting the level of MFN1 in the nervous system in vivo rescued all phenotypes in mutant MFN2R94Q-expressing mice. These data demonstrate that the MFN1/MFN2 ratio is a key determinant of tissue specificity in CMT2A and indicate that augmentation of MFN1 in the nervous system is a viable therapeutic strategy for the disease.

  • abrogating mitochondrial dynamics in mouse hearts accelerates mitochondrial senescence
    Cell Metabolism, 2017
    Co-Authors: Moshi Song, Antonietta Franco, Julie Fleischer, Lihong Zhang, Gerald W Dorn
    Abstract:

    Summary Mitochondrial fusion and fission are critical to heart health; genetically interrupting either is rapidly lethal. To understand whether it is loss of, or the imbalance between, fusion and fission that underlies observed cardiac phenotypes, we engineered mice in which Mfn-mediated fusion and Drp1-mediated fission could be concomitantly abolished. Compared to fusion-defective MFN1/Mfn2 cardiac knockout or fission-defective Drp1 cardiac knockout mice, MFN1/Mfn2/Drp1 cardiac triple-knockout mice survived longer and manifested a unique pathological form of cardiac hypertrophy. Over time, however, combined abrogation of fission and fusion provoked massive progressive mitochondrial accumulation that severely distorted cardiomyocyte sarcomeric architecture. Mitochondrial biogenesis was not responsible for mitochondrial superabundance, whereas mitophagy was suppressed despite impaired mitochondrial proteostasis. Similar but milder defects were observed in aged hearts. Thus, cardiomyopathies linked to dynamic imbalance between fission and fusion are temporarily mitigated by forced mitochondrial adynamism at the cost of compromising mitochondrial quantity control and accelerating mitochondrial senescence.

  • THE ADULT MURINE HEART IS PROTECTED AGAINST ISCHEMIA-REPERFUSION INJURY IN THE ABSENCE OF BOTH MITOFUSIN (MFN) PROTEINS
    Heart, 2014
    Co-Authors: Andrew R. Hall, Yun Chen, Gerald W Dorn, N Burke, Rk Dongworth, Derek J. Hausenloy
    Abstract:

    Background Cardiac-specific ablation of both MFN1 and Mfn2 in the adult heart results in mitochondrial fragmentation and a lethal cardiomyopathy after about 6 weeks. The effect of combined MFN1 and Mfn2 deletion on the susceptibility to acute ischemia-reperfusion injury (IRI) and subsequent calcium overload is not known, and is investigated in this study. Methods and Results Cardiac-specific ablation of both MFN1 and Mfn2 (DKO) was initiated in mice aged 5 weeks using 5 days administration of tamoxifen (MerCreMer), resulting in total knockout of both these proteins at the age of 10 weeks. These mice were subjected to in vivo myocardial ischemia (30 mins) followed by 24 hrs reperfusion before myocardial infarct size was determined. The sustained MI size in the DKO mice was 50% smaller than that in the WT control mice. These findings were associated with decreased MPTP opening susceptibility (assessed by calcium-induced mitochondrial swelling), reduced mitochondrial calcium overload after simulated IR (assessed by Rhod2 staining) and impaired mitochondrial respiration in the DKO hearts when compared to WT control. Conclusions We have shown that the adult murine heart deficient in both MFN1 and Mfn2 was protected against acute IRI, a finding which was associated with defects in mitochondrial function and reduced mitochondrial calcium overload. This data suggests that the Mitofusins may be therapeutic targets for cardioprotection.

  • mitochondrial fusion is essential for organelle function and cardiac homeostasis
    Circulation Research, 2011
    Co-Authors: Yun Chen, Gerald W Dorn
    Abstract:

    Rationale: Mitochondria constitute 30% of myocardial mass. Mitochondrial fusion and fission appear essential for health of most tissues. Mitochondrial fission occurs in neonatal cardiomycyte and is implicated in cardiomyocyte death. Mitochondrial fusion has not been observed in postmitotic myocytes of adult hearts, and its occurrence and function in this context are controversial. Objective: Determine the consequences on organelle and organ function of disrupting cardiomyocyte mitochondrial fusion in vivo. Methods and Results: The murine MFN1 and mfn2 genes, encoding mitofusins (Mfn) 1 and 2 that mediate mitochondrial tethering and outer mitochondrial membrane fusion, were interrupted by Cre-mediated excision of essential exons in neonatal (Nkx2.5-Cre) and adult (MYH6 modified estrogen receptor-Cre-modified estrogen receptor plus tamoxifen or Raloxifene) hearts. Embryonic combined MFN1/Mfn2 ablation was lethal after e9.5. Conditional combined MFN1/Mfn2 ablation in adult hearts induced mitochondrial fragmentation, cardiomyocyte and mitochondrial respiratory dysfunction, and rapidly progressive and lethal dilated cardiomyopathy. Before heart failure developed, cardiomyocyte shortening and calcium cycling were unaffected by absence of MFN1 and Mfn2. Based on the time course over which fusion-defective mitochondrial size decreases, a mitochondrial fusion/fission cycle in adult mouse hearts occurs approximately every 16 days. Conclusions: Mitochondrial fusion in adult cardiac myocytes is necessary to maintain normal mitochondrial morphology and is essential for normal cardiac respiratory and contractile function. Interruption of mitochondrial fusion causes lethal cardiac failure at a time corresponding to 3 or 4 cycles of unopposed mitochondrial fission.

David C Chan - One of the best experts on this subject based on the ideXlab platform.

  • mitochondrial dynamics is a distinguishing feature of skeletal muscle fiber types and regulates organellar compartmentalization
    Cell Metabolism, 2015
    Co-Authors: Prashant Mishra, Grigor Varuzhanyan, Anh H Pham, David C Chan
    Abstract:

    Skeletal muscle fibers differentiate into specific fiber types with distinct metabolic properties determined by their reliance on oxidative phosphorylation (OXPHOS). Using in vivo approaches, we find that OXPHOS-dependent fibers, compared to glycolytic fibers, contain elongated mitochondrial networks with higher fusion rates that are dependent on the mitofusins MFN1 and Mfn2. Switching of a glycolytic fiber to an oxidative IIA type is associated with elongation of mitochondria, suggesting that mitochondrial fusion is linked to metabolic state. Furthermore, we reveal that mitochondrial proteins are compartmentalized to discrete domains centered around their nuclei of origin. The domain dimensions are dependent on fiber type and are regulated by the mitochondrial dynamics proteins MFN1, Mfn2, and Mff. Our results indicate that mitochondrial dynamics is tailored to fiber type physiology and provides a rationale for the segmental defects characteristic of aged and diseased muscle fibers.

  • loss of mfn2 results in progressive retrograde degeneration of dopaminergic neurons in the nigrostriatal circuit
    Human Molecular Genetics, 2012
    Co-Authors: Anh H Pham, Shuxia Meng, Quynh N Chu, David C Chan
    Abstract:

    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.

  • complementation between mouse MFN1 and mfn2 protects mitochondrial fusion defects caused by cmt2a disease mutations
    Journal of Cell Biology, 2007
    Co-Authors: Scott A. Detmer, David C Chan
    Abstract:

    Mfn2, an oligomeric mitochondrial protein important for mitochondrial fusion, is mutated in Charcot-Marie-Tooth disease (CMT) type 2A, a peripheral neuropathy characterized by axonal degeneration. In addition to homooligomeric complexes, Mfn2 also associates with MFN1, but the functional significance of such heterooligomeric complexes is unknown. Also unknown is why Mfn2 mutations in CMT2A lead to cell type–specific defects given the widespread expression of Mfn2. In this study, we show that homooligomeric complexes formed by many Mfn2 disease mutants are nonfunctional for mitochondrial fusion. However, wild-type MFN1 complements mutant Mfn2 through the formation of heterooligomeric complexes, including complexes that form in trans between mitochondria. Wild-type Mfn2 cannot complement the disease alleles. Our results highlight the functional importance of MFN1–Mfn2 heterooligomeric complexes and the close interplay between the two mitofusins in the control of mitochondrial fusion. Furthermore, they suggest that tissues with low MFN1 expression are vulnerable in CMT2A and that methods to increase MFN1 expression in the peripheral nervous system would benefit CMT2A patients.

  • disruption of fusion results in mitochondrial heterogeneity and dysfunction
    Journal of Biological Chemistry, 2005
    Co-Authors: Hsiuchen Chen, Anne Chomyn, David C Chan
    Abstract:

    Mitochondria undergo continual cycles of fusion and fission, and the balance of these opposing processes regulates mitochondrial morphology. Paradoxically, cells invest many resources to maintain tubular mitochondrial morphology, when reducing both fusion and fission simultaneously achieves the same end. This observation suggests a requirement for mitochondrial fusion, beyond maintenance of organelle morphology. Here, we show that cells with targeted null mutations in MFN1 or Mfn2 retained low levels of mitochondrial fusion and escaped major cellular dysfunction. Analysis of these mutant cells showed that both homotypic and heterotypic interactions of Mfns are capable of fusion. In contrast, cells lacking both MFN1 and Mfn2 completely lacked mitochondrial fusion and showed severe cellular defects, including poor cell growth, widespread heterogeneity of mitochondrial membrane potential, and decreased cellular respiration. Disruption of OPA1 by RNAi also blocked all mitochondrial fusion and resulted in similar cellular defects. These defects in Mfn-null or OPA1-RNAi mammalian cells were corrected upon restoration of mitochondrial fusion, unlike the irreversible defects found in fzo yeast. In contrast, fragmentation of mitochondria, without severe loss of fusion, did not result in such cellular defects. Our results showed that key cellular functions decline as mitochondrial fusion is progressively abrogated.

  • mitofusins MFN1 and mfn2 coordinately regulate mitochondrial fusion and are essential for embryonic development
    Journal of Cell Biology, 2003
    Co-Authors: Hsiuchen Chen, Scott A. Detmer, Andrew J Ewald, Erik E Griffin, Scott E Fraser, David C Chan
    Abstract:

    Mitochondrial morphology is determined by a dynamic equilibrium between organelle fusion and fission, but the significance of these processes in vertebrates is unknown. The mitofusins, MFN1 and Mfn2, have been shown to affect mitochondrial morphology when overexpressed. We find that mice deficient in either MFN1 or Mfn2 die in midgestation. However, whereas Mfn2 mutant embryos have a specific and severe disruption of the placental trophoblast giant cell layer, MFN1-deficient giant cells are normal. Embryonic fibroblasts lacking MFN1 or Mfn2 display distinct types of fragmented mitochondria, a phenotype we determine to be due to a severe reduction in mitochondrial fusion. Moreover, we find that MFN1 and Mfn2 form homotypic and heterotypic complexes and show, by rescue of mutant cells, that the homotypic complexes are functional for fusion. We conclude that MFN1 and Mfn2 have both redundant and distinct functions and act in three separate molecular complexes to promote mitochondrial fusion. Strikingly, a subset of mitochondria in mutant cells lose membrane potential. Therefore, mitochondrial fusion is essential for embryonic development, and by enabling cooperation between mitochondria, has protective effects on the mitochondrial population.

Katsuyoshi Mihara - One of the best experts on this subject based on the ideXlab platform.

  • identification of a novel protein that regulates mitochondrial fusion by modulating mitofusin mfn protein function
    Journal of Cell Science, 2006
    Co-Authors: Yuka Eura, Naotada Ishihara, Toshihiko Oka, Katsuyoshi Mihara
    Abstract:

    Mitofusin proteins 1 and 2 (MFN1 and Mfn2, respectively) of the mammalian mitochondrial outer membrane are homologues of Drosophila FZO and yeast Fzo1, and both are essential for GTP-dependent mitochondrial fusion. We identified a 55-kDa Mfn-binding protein named MIB. It is a member of the medium-chain dehydrogenase/reductase protein superfamily, and has a conserved coenzyme-binding domain (CBD). The majority of MIB is localized in the cytoplasm but a small amount is associated with mitochondria. Exogenous expression of MIB in HeLa cells induced mitochondrial fragmentation, which was prevented by coexpression of MFN1, suggesting a functional interaction of MIB with Mfn proteins; the GGVG sequence in the CBD of MIB is essential for its function. By contrast, MIB knockdown resulted in growth arrest of the cells, although apoptotic sensitivity was not affected by either its knockdown or its overexpression. Furthermore, MIB knockdown induced a large extension of mitochondrial network structures. By contrast, a double knockdown of MIB and MFN1 resulted in mitochondrial fragmentation and reversal of the growth arrest, the morphology and growth phenotype induced by knockdown of MFN1 alone, again suggesting that MIB modulates MFN1 function. Together, these findings suggest that MIB is essential for cellular function by regulating mitochondrial membrane dynamics in cooperation with Mfn proteins.

  • mitofusin 1 and 2 play distinct roles in mitochondrial fusion reactions via gtpase activity
    Journal of Cell Science, 2004
    Co-Authors: Naotada Ishihara, Yuka Eura, Katsuyoshi Mihara
    Abstract:

    The mammalian homologues of yeast and Drosophila Fzo, mitofusin (Mfn) 1 and 2, are both essential for mitochondrial fusion and maintenance of mitochondrial morphology. Though the GTPase domain is required for Mfn protein function, the molecular mechanisms of the GTPase-dependent reaction as well as the functional division of the two Mfn proteins are unknown. To examine the function of Mfn proteins, tethering of mitochondrial membranes was measured in vitro by fluorescence microscopy using green fluorescence protein- or red fluorescent protein-tagged and MFN1-expressing mitochondria, or by immunoprecipitation using mitochondria harboring HA- or FLAG-tagged Mfn proteins. These experiments revealed that MFN1-harboring mitochondria were efficiently tethered in a GTP-dependent manner, whereas Mfn2-harboring mitochondria were tethered with only low efficiency. Sucrose density gradient centrifugation followed by co-immunoprecipitation revealed that MFN1 produced oligomerized ∼250 kDa and ∼450 kDa complexes in a GTP-dependent manner. The ∼450 kDa complex contained oligomerized MFN1 from distinct apposing membranes (docking complex), whereas the ∼250 kDa complex was composed of MFN1 present on the same membrane or in the membrane-solubilized state (cis complex). These results were also confirmed using blue-native PAGE. MFN1 exhibited higher activity for this reaction than Mfn2. Purified recombinant MFN1 exhibited ∼eightfold higher GTPase activity than Mfn2. These findings indicate that the two Mfn proteins have distinct activities, and suggest that MFN1 is mainly responsible for GTP-dependent membrane tethering.

  • Mitofusin 1 and 2 play distinct roles in mitochondrial fusion reactions via GTPase activity.
    Journal of cell science, 2004
    Co-Authors: Naotada Ishihara, Yuka Eura, Katsuyoshi Mihara
    Abstract:

    The mammalian homologues of yeast and Drosophila Fzo, mitofusin (Mfn) 1 and 2, are both essential for mitochondrial fusion and maintenance of mitochondrial morphology. Though the GTPase domain is required for Mfn protein function, the molecular mechanisms of the GTPase-dependent reaction as well as the functional division of the two Mfn proteins are unknown. To examine the function of Mfn proteins, tethering of mitochondrial membranes was measured in vitro by fluorescence microscopy using green fluorescence protein- or red fluorescent protein-tagged and MFN1-expressing mitochondria, or by immunoprecipitation using mitochondria harboring HA- or FLAG-tagged Mfn proteins. These experiments revealed that MFN1-harboring mitochondria were efficiently tethered in a GTP-dependent manner, whereas Mfn2-harboring mitochondria were tethered with only low efficiency. Sucrose density gradient centrifugation followed by co-immunoprecipitation revealed that MFN1 produced oligomerized approximately 250 kDa and approximately 450 kDa complexes in a GTP-dependent manner. The approximately 450 kDa complex contained oligomerized MFN1 from distinct apposing membranes (docking complex), whereas the approximately 250 kDa complex was composed of MFN1 present on the same membrane or in the membrane-solubilized state (cis complex). These results were also confirmed using blue-native PAGE. MFN1 exhibited higher activity for this reaction than Mfn2. Purified recombinant MFN1 exhibited approximately eightfold higher GTPase activity than Mfn2. These findings indicate that the two Mfn proteins have distinct activities, and suggest that MFN1 is mainly responsible for GTP-dependent membrane tethering.

  • two mitofusin proteins mammalian homologues of fzo with distinct functions are both required for mitochondrial fusion
    Journal of Biochemistry, 2003
    Co-Authors: Yuka Eura, Naotada Ishihara, Sadaki Yokota, Katsuyoshi Mihara
    Abstract:

    Mitochondria are dynamic organelles that undergo frequent fission and fusion or branching. Although these morphologic changes are considered crucial for cellular functions, the underlying mechanisms remain elusive, especially in mammalian cells. We characterized two rat mitochondrial outer membrane proteins, MFN1 and Mfn2, with distinct tissue expressions, that are homologous to Drosophila Fzo, a GTPase involved in mitochondrial fusion. Expression of the GTPase-domain mutant of Mfn2 (Mfn2(K109T)) in HeLa cells induced mitochondrial fragmentation in which Mfn2(K109T) localized at the restricted domains. Immuno-electronmicroscopy revealed that Mfn2(K109T) was concentrated at the contact domains between adjacent mitochondria, suggesting that fusion of the outer membrane was arrested at some intermediate step. MFN1 expression induced highly connected tubular network structures depending on the functional GTPase domain. The MFN1-induced tubular networks were suppressed by co-expression with Mfn2. In vivo depletion of either isoform by RNA interference revealed that both are required to maintain normal mitochondrial morphology. The fusion of differentially-labeled mitochondria in HeLa cells subjected to depletion of either Mfn isoform and subsequent cell fusion by hemagglutinating virus of Japan revealed that both proteins have distinct functions in mitochondrial fusion. We conclude that the two Mfn isoforms cooperate in mitochondrial fusion in mammalian cells.

Carla M. Koehler - One of the best experts on this subject based on the ideXlab platform.

  • rapid degradation of mutant slc25a46 by the ubiquitin proteasome system results in MFN1 2 mediated hyperfusion of mitochondria
    Molecular Biology of the Cell, 2017
    Co-Authors: Janos Steffen, Ajay A. Vashisht, Jijun Wan, Joanna C. Jen, Steven M. Claypool, James A. Wohlschlegel, Carla M. Koehler
    Abstract:

    SCL25A46 is a mitochondrial carrier protein that surprisingly localizes to the outer membrane and is distantly related to Ugo1. Here we show that a subset of SLC25A46 interacts with mitochondrial dynamics components and the MICOS complex. Decreased expression of SLC25A46 results in increased stability and oligomerization of MFN1 and MFN2 on mitochondria, promoting mitochondrial hyperfusion. A mutation at L341P causes rapid degradation of SLC25A46, which manifests as a rare disease, pontocerebellar hypoplasia. The E3 ubiquitin ligases MULAN and MARCH5 coordinate ubiquitylation of SLC25A46 L341P, leading to degradation by organized activities of P97 and the proteasome. Whereas outer mitochondrial membrane-associated degradation is typically associated with apoptosis or a specialized type of autophagy termed mitophagy, SLC25A46 degradation operates independently of activation of outer membrane stress pathways. Thus SLC25A46 is a new component in mitochondrial dynamics that serves as a regulator for MFN1/2 oligomerization. Moreover, SLC25A46 is selectively degraded from the outer membrane independently of mitophagy and apoptosis, providing a framework for mechanistic studies in the proteolysis of outer membrane proteins.

  • Rapid degradation of mutant SLC25A46 by the ubiquitin-proteasome system results in MFN1/2-mediated hyperfusion of mitochondria
    Molecular biology of the cell, 2017
    Co-Authors: Janos Steffen, Ajay A. Vashisht, Jijun Wan, Joanna C. Jen, Steven M. Claypool, James A. Wohlschlegel, Carla M. Koehler
    Abstract:

    SCL25A46 is a mitochondrial carrier protein that surprisingly localizes to the outer membrane and is distantly related to Ugo1. Here we show that a subset of SLC25A46 interacts with mitochondrial dynamics components and the MICOS complex. Decreased expression of SLC25A46 results in increased stability and oligomerization of MFN1 and MFN2 on mitochondria, promoting mitochondrial hyperfusion. A mutation at L341P causes rapid degradation of SLC25A46, which manifests as a rare disease, pontocerebellar hypoplasia. The E3 ubiquitin ligases MULAN and MARCH5 coordinate ubiquitylation of SLC25A46 L341P, leading to degradation by organized activities of P97 and the proteasome. Whereas outer mitochondrial membrane-associated degradation is typically associated with apoptosis or a specialized type of autophagy termed mitophagy, SLC25A46 degradation operates independently of activation of outer membrane stress pathways. Thus SLC25A46 is a new component in mitochondrial dynamics that serves as a regulator for MFN1/2 oligomerization. Moreover, SLC25A46 is selectively degraded from the outer membrane independently of mitophagy and apoptosis, providing a framework for mechanistic studies in the proteolysis of outer membrane proteins.

Suzanne Hoppins - One of the best experts on this subject based on the ideXlab platform.

  • identification of a mitofusin specificity region that confers unique activities to MFN1 and mfn2
    Molecular Biology of the Cell, 2019
    Co-Authors: Stephanie R Sloat, B N Whitley, E A Engelhart, Suzanne Hoppins
    Abstract:

    Mitochondrial structure can be maintained at steady state or modified in response to changes in cellular physiology. This is achieved by the coordinated regulation of dynamic properties including mitochondrial fusion, division, and transport. Disease states, including neurodegeneration, are associated with defects in these processes. In vertebrates, two mitofusin paralogues, MFN1 and Mfn2, are required for efficient mitochondrial fusion. The mitofusins share a high degree of homology and have very similar domain architecture, including an amino terminal GTPase domain and two extended helical bundles that are connected by flexible regions. MFN1 and Mfn2 are nonredundant and are both required for mitochondrial outer membrane fusion. However, the molecular features that make these proteins functionally distinct are poorly defined. By engineering chimeric proteins composed of MFN1 and Mfn2, we discovered a region that contributes to isoform-specific function (mitofusin isoform-specific region [MISR]). MISR confers unique fusion activity and mitofusin-specific nucleotide-dependent assembly properties. We propose that MISR functions in higher-order oligomerization either directly, as an interaction interface, or indirectly through conformational changes.

  • identification of a mitofusin specificity region that confers unique activities to MFN1 and mfn2
    bioRxiv, 2019
    Co-Authors: Stephanie R Sloat, B N Whitley, E A Engelhart, Suzanne Hoppins
    Abstract:

    ABSTRACT Mitochondrial structure can be maintained at steady state or modified in response to changes in cellular physiology. This is achieved by the coordinated regulation of dynamic properties including mitochondrial fusion, division and transport. Disease states, including neurodegeneration, are associated with defects in these processes. In vertebrates, two Mitofusin paralogs, MFN1 and Mfn2, are required for efficient mitochondrial fusion. The Mitofusins share a high degree of homology and have very similar domain architecture, including an amino terminal GTPase domain and two extended helical bundles that are connected by flexible regions. MFN1 and Mfn2 are non-redundant and are both required for mitochondrial outer membrane fusion. However, the molecular features that make these proteins functionally distinct are poorly defined. By engineering chimeric proteins composed of MFN1 and Mfn2, we discovered a region that contributes to isoform-specific function (Mitofusin Isoform Specific Region – MISR). MISR confers unique fusion activity and Mitofusin specific nucleotide-dependent assembly properties. We propose that MISR functions in higher order oligomerization either directly, as an interaction interface, or indirectly through conformational changes.

  • a catalytic domain variant of mitofusin requiring a wildtype paralog for function uncouples mitochondrial outer membrane tethering and fusion
    Journal of Biological Chemistry, 2019
    Co-Authors: Emily A Engelhart, Suzanne Hoppins
    Abstract:

    Mitofusins (Mfns) are dynamin-related GTPases that mediate mitochondrial outer-membrane fusion, a process that is required for mitochondrial and cellular health. In MFN1 and Mfn2 paralogs, a conserved phenylalanine (Phe-202 (MFN1) and Phe-223 (Mfn2)) located in the GTPase domain on a conserved β strand is part of an aromatic network in the core of this domain. To gain insight into the poorly understood mechanism of Mfn-mediated membrane fusion, here we characterize a Mitofusin mutant variant etiologically linked to Charcot–Marie–Tooth syndrome. From analysis of mitochondrial structure in cells and mitochondrial fusion in vitro, we found that conversion of Phe-202 to leucine in either MFN1 or Mfn2 diminishes the fusion activity of heterotypic complexes with both MFN1 and Mfn2 and abolishes fusion activity of homotypic complexes. Using coimmunoprecipitation and native gel analysis, we further dissect the steps of mitochondrial fusion and demonstrate that the mutant variant has normal tethering activity but impaired higher-order nucleotide-dependent assembly. The defective coupling of tethering to membrane fusion observed here suggests that nucleotide-dependent self-assembly of Mitofusin is required after tethering to promote membrane fusion.

  • the soluble form of bax regulates mitochondrial fusion via mfn2 homotypic complexes
    Molecular Cell, 2011
    Co-Authors: Suzanne Hoppins, Megan M Cleland, Richard J Youle, Michael J Mccaffery, Frank Edlich, Soojay Banerjee, Jodi Nunnari
    Abstract:

    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.

  • coassembly of mgm1 isoforms requires cardiolipin and mediates mitochondrial inner membrane fusion
    Journal of Cell Biology, 2009
    Co-Authors: Rachel M Devay, Suzanne Hoppins, Lenin Dominguezramirez, Laura L Lackner, Henning Stahlberg, Jodi Nunnari
    Abstract:

    Two dynamin-related protein (DRP) families are essential for fusion of the outer and inner mitochondrial membranes, Fzo1 (yeast)/MFN1/Mfn2 (mammals) and Mgm1 (yeast)/Opa1 (mammals), respectively. Fzo1/Mfns possess two medial transmembrane domains, which place their critical GTPase and coiled-coil domains in the cytosol. In contrast, Mgm1/Opa1 are present in cells as long (l) isoforms that are anchored via the N terminus to the inner membrane, and short (s) isoforms were predicted to be soluble in the intermembrane space. We addressed the roles of Mgm1 isoforms and how DRPs function in membrane fusion. Our analysis indicates that in the absence of a membrane, l- and s-Mgm1 both exist as inactive GTPase monomers, but that together in trans they form a functional dimer in a cardiolipin-dependent manner that is the building block for higher-order assemblies.