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Francesc Palau - One of the best experts on this subject based on the ideXlab platform.
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mitochondria and calcium defects correlate with axonal dysfunction in GDAP1 related charcot marie tooth mouse model
Neurobiology of Disease, 2021Co-Authors: Azahara Civeratregon, Raul Benitez, Janet Hoenicka, Jorgina Satrústegui, Laura Dominguez, Paula Martinezvalero, Claudia Serrano, Alexander Vallmitjana, Francesc PalauAbstract:Ganglioside-induced differentiation associated protein 1 (GDAP1) gene encodes a protein of the mitochondrial outer membrane and of the mitochondrial membrane contacts with the endoplasmic reticulum (MAMs) and lysosomes. Since mutations in GDAP1 cause Charcot-Marie-Tooth, an inherited motor and sensory neuropathy, its function is essential for peripheral nerve physiology. Our previous studies showed structural and functional defects in mitochondria and their contacts when GDAP1 is depleted. Nevertheless, the underlying axonal pathophysiological events remain unclear. Here, we have used embryonic motor neurons (eMNs) cultures from GDAP1 knockout (GDAP1-/-) mice to investigate in vivo mitochondria and calcium homeostasis in the axons. We imaged mitochondrial axonal transport and we found a defective pattern in the GDAP1-/- eMNs. We also detected pathological and functional mitochondria membrane abnormalities with a drop in ATP production and a deteriorated bioenergetic status. Another consequence of the loss of GDAP1 in the soma and axons of eMNs was the in vivo increase calcium levels in both basal conditions and during recovery after neuronal stimulation with glutamate. Further, we found that glutamate-stimulation of respiration was lower in GDAP1-/- eMNs showing that the basal bioenergetics failure jeopardizes a full respiratory response and prevents a rapid return of calcium to basal levels. Together, our results demonstrate that the loss of GDAP1 critically compromises the morphology and function of mitochondria and its relationship with calcium homeostasis in the soma and axons, offering important insight into the cellular mechanisms associated with axonal degeneration of GDAP1-related CMT neuropathies and the relevance that axon length may have.
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Mitochondria-lysosome membrane contacts are defective in GDAP1-related Charcot-Marie-Tooth disease
Human molecular genetics, 2020Co-Authors: Lara Cantarero, Azahara Civera-tregón, Elena Juárez-escoto, María Rodríguez-sanz, Monica Roldan, Raul Benitez, Janet Hoenicka, Francesc PalauAbstract:Mutations in the GDAP1 gene cause Charcot-Marie-Tooth (CMT) neuropathy. GDAP1 is an atypical glutathione S-transferase (GST) of the outer mitochondrial membrane and the mitochondrial membrane contacts with the endoplasmic reticulum (MAMs). Here, we investigate the role of this GST in the autophagic flux and the membrane contact sites (MCSs) between mitochondria and lysosomes in the cellular pathophysiology of GDAP1 deficiency. We demonstrate that GDAP1 participates in basal autophagy and that its depletion affects LC3 and PI3P biology in autophagosome biogenesis and membrane trafficking from MAMs. GDAP1 also contributes to the maturation of lysosome by interacting with PYKfyve kinase, a pH-dependent master lysosomal regulator. GDAP1 deficiency causes giant lysosomes with hydrolytic activity, a delay in the autophagic lysosome reformation, and TFEB activation. Notably, we found that GDAP1 interacts with LAMP-1, which supports that GDAP1-LAMP-1 is a new tethering pair of mitochondria and lysosome membrane contacts. We observed mitochondria-lysosome MCSs in soma and axons of cultured mouse embryonic motor neurons and human neuroblastoma cells. GDAP1 deficiency reduces the MCSs between these organelles, causes mitochondrial network abnormalities, and decreases levels of cellular glutathione (GSH). The supply of GSH-MEE suffices to rescue the lysosome membranes and the defects of the mitochondrial network, but not the interorganelle MCSs nor early autophagic events. Overall, we show that GDAP1 enables the proper function of mitochondrial MCSs in both degradative and nondegradative pathways, which could explain primary insults in GDAP1-related CMT pathophysiology, and highlights new redox-sensitive targets in axonopathies where mitochondria and lysosomes are involved.
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Neuroinflammation in the pathogenesis of axonal Charcot-Marie-Tooth disease caused by lack of GDAP1
Experimental neurology, 2019Co-Authors: Paula Juárez, Azahara Civera-tregón, Lara Cantarero, Janet Hoenicka, Sara Fernandez-lizarbe, Isabel Herrer, Francesc PalauAbstract:Mutations in the GDAP1 mitochondrial outer membrane gene cause Charcot-Marie-Tooth (CMT) neuropathy. Reduction or absence of GDAP1 has been associated with abnormal changes in the mitochondrial morphology and dynamics, oxidative stress and changes in calcium homeostasis. Neuroinflammation has been described in rodent models of genetic demyelinating CMT neuropathies but not in CMT primarily associated with axonopathy. Inflammatory processes have also been related to mitochondrial changes and oxidative stress in central neurodegenerative disorders. Here we investigated the presence of neuroinflammation in the axonal neuropathy of the GDAP1-/- mice. We showed by transcriptome profile of spinal cord and the in vivo detection of activated phagocytes that the absence of GDAP1 is associated with upregulation of inflammatory pathways. We observed reactive gliosis in spinal cord with increase of the astroglia markers GFAP and S100B, and the microglia marker IBA1. Additionally, we found significant increase of inflammatory mediators such as TNF-α and pERK, and C1qa and C1qb proteins of the complement system. Importantly, we observed an increased expression of CD206 and CD86 as M2 and M1 microglia and macrophage response markers, respectively, in GDAP1-/- mice. These inflammatory changes were also associated with abnormal molecular changes in synapses. In summary, we demonstrate that inflammation in spinal cord and sciatic nerve, but not in brain and cerebellum, is part of the pathophysiology of axonal GDAP1-related CMT.
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Calcium Deregulation and Mitochondrial Bioenergetics in GDAP1-Related CMT Disease.
International journal of molecular sciences, 2019Co-Authors: Paloma González-sánchez, Francesc Palau, Araceli Del Arco, Jorgina SatrústeguiAbstract:The pathology of Charcot-Marie-Tooth (CMT), a disease arising from mutations in different genes, has been associated with an impairment of mitochondrial dynamics and axonal biology of mitochondria. Mutations in ganglioside-induced differentiation-associated protein 1 (GDAP1) cause several forms of CMT neuropathy, but the pathogenic mechanisms involved remain unclear. GDAP1 is an outer mitochondrial membrane protein highly expressed in neurons. It has been proposed to play a role in different aspects of mitochondrial physiology, including mitochondrial dynamics, oxidative stress processes, and mitochondrial transport along the axons. Disruption of the mitochondrial network in a neuroblastoma model of GDAP1-related CMT has been shown to decrease Ca2+ entry through the store-operated calcium entry (SOCE), which caused a failure in stimulation of mitochondrial respiration. In this review, we summarize the different functions proposed for GDAP1 and focus on the consequences for Ca2+ homeostasis and mitochondrial energy production linked to CMT disease caused by different GDAP1 mutations.
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CMT-linked loss-of-function mutations in GDAP1 impair store-operated Ca2+ entry-stimulated respiration.
Scientific reports, 2017Co-Authors: Paloma González-sánchez, Francesc Palau, David Pla-martin, Eduardo Calpena, Carlos B. Rueda, Paula Martinez-valero, Araceli Del Arco, Jorgina SatrústeguiAbstract:GDAP1 is an outer mitochondrial membrane protein involved in Charcot-Marie-Tooth (CMT) disease. Lack of GDAP1 gives rise to altered mitochondrial networks and endoplasmic reticulum (ER)-mitochondrial interactions resulting in a decreased ER-Ca2+ levels along with a defect on store-operated calcium entry (SOCE) related to a misallocation of mitochondria to subplasmalemmal sites. The defect on SOCE is mimicked by MCU silencing or mitochondrial depolarization, which prevent mitochondrial calcium uptake. Ca2+ release from de ER and Ca2+ inflow through SOCE in neuroblastoma cells result in a Ca2+-dependent upregulation of respiration which is blunted in GDAP1 silenced cells. Reduced SOCE in cells with CMT recessive missense mutations in the α-loop of GDAP1, but not dominant mutations, was associated with smaller SOCE-stimulated respiration. These cases of GDAP1 deficiency also resulted in a decreased ER-Ca2+ levels which may have pathological implications. The results suggest that CMT neurons may be under energetic constraints upon stimulation by Ca2+ mobilization agonists and point to a potential role of perturbed mitochondria-ER interaction related to energy metabolism in forms of CMT caused by some of the recessive or null mutations of GDAP1.
Laia Pedrola - One of the best experts on this subject based on the ideXlab platform.
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cell expression of GDAP1 in the nervous system and pathogenesis of charcot marie tooth type 4a disease
Journal of Cellular and Molecular Medicine, 2008Co-Authors: Laia Pedrola, Antonio Espert, Erich E. Sirkowski, Steven S. Scherer, Isabel Fariñas, Teresa Valdessanchez, Maribel Sanchezpiris, Francesc PalauAbstract:Mutations in the mitochondrial protein GDAP1 are the cause of Charcot-Marie-Tooth type 4A disease (CMT4A), a severe form of peripheral neuropathy associated with either demyelinating, axonal or intermediate pheno-types. GDAP1 is located in the outer mitochondrial membrane and it seems that may be related with the mitochondrial network dynamics. We are interested to define cell expression in the nervous system and the effect of mutations in mitochondrial morphology and pathogenesis of the disease. We investigated GDAP1 expression in the nervous system and dorsal root ganglia (DRG) neuron cultures. GDAP1 is expressed in motor and sensory neurons of the spinal cord and other large neurons such as cerebellar Purkinje neurons, hippocampal pyramidal neurons, mitral neurons of the olfactory bulb and cortical pyramidal neurons. The lack of GDAP1 staining in the white matter and nerve roots suggested that glial cells do not express GDAP1. In DRG cultures satellite cells and Schwann cells were GDAP1-negative. Overexpression of GDAP1-induced fragmentation of mitochondria suggesting a role of GDAP1 in the fission pathway of the mitochondrial dynamics. Missense mutations showed two different patterns: most of them induced mitochondrial fragmentation but the T157P mutation showed an aggregation pattern. Whereas null mutations of GDAP1 should be associated with loss of function of the protein, missense mutations may act through different pathogenic mechanisms including a dominant-negative effect, suggesting that different molecular mechanisms may underlay the pathogenesis of CMT4A.
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A novel Met116Thr mutation in the GDAP1 gene in a Polish family with the axonal recessive Charcot-Marie-Tooth type 4 disease.
Journal of the neurological sciences, 2005Co-Authors: Dagmara Kabzińska, Francesc Palau, Andrzej Kochanski, Laia Pedrola, Hanna Drac, Katarzyna Rowińska-marcińska, Barbara Ryniewicz, Irena Hausmanowa-petrusewiczAbstract:Mutations in the gene coding for ganglioside-induced differentiation-associated protein-1 (GDAP1), which maps to chromosome 8q21, have been described in families with autosomal recessive Charcot-Marie-Tooth disease (CMT4A). Interestingly, some mutations in the GDAP1 gene have been reported in the demyelinating form of CMT1 disease, whereas others were found in patients with the axonal type of CMT disease. So far, 23 mutations in the GDAP1 gene have been reported in patients of different ethnic origins. In this study we report a novel mutation Met116Thr in the GDAP1 gene identified in a three generation Polish family with axonal CMT4.
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GDAP1 the protein causing charcot marie tooth disease type 4a is expressed in neurons and is associated with mitochondria
Human Molecular Genetics, 2005Co-Authors: Antonio Espert, Laia Pedrola, Reyes Claramunt, Michael E. Shy, Francesc PalauAbstract:Mutations in GDAP1, the ganglioside-induced differentiation-associated protein 1 gene, cause Charcot-Marie-Tooth (CMT) type 4A, a severe autosomal recessive form of neuropathy associated with either demye-linating or axonal phenotypes. Here, we demonstrate that GDAP1 has far greater expression in neurons than in myelinating Schwann cells. We investigated cell localization of GDAP1 in a human neuroblastoma cell line by means of transient overexpression and co-localization with organelle markers in COS-7 cells and by western blot analysis of subcell fractions with anti-GDAP1 polyclonal antibodies. We observed that GDAP1 is localized in mitochondria. We also show that C-terminal transmembrane domains are necessary for the correct localization in mitochondria; however, missense mutations do not change the mitochondrial pattern of the wild-type protein. Our findings suggest that CMT4A disease is in fact a mitochondrial neuropathy mainly involving axons and represents a disease belonging to the new category of mitochondrial disorders caused by mutations in nuclear genes. We postulate that GDAP1 may be related to the maintenance of the mitochondrial network.
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Evolutionary and Structural Analyses of GDAP1, Involved in Charcot-Marie-Tooth Disease, Characterize a Novel Class of Glutathione Transferase-Related Genes
Molecular biology and evolution, 2003Co-Authors: Francesc Palau, Laia Pedrola, Antonio Marco, Ana Cuesta, Ignacio MarínAbstract:Mutations in the Ganglioside-induced differentiation-associated protein-1 (GDAP1) gene cause autosomal recessive Charcot-Marie-Tooth disease type 4A. The protein encoded by GDAP1 shows clear similarity to glutathione transferases (also known as glutathione S-transferases or GSTs). The human genome contains a paralog of GDAP1 called GDAP1L1. Using comparative genomics, we show that orthologs of GDAP1 and GDAP1L1 are found in mammals, birds, amphibians, and fishes. Likely orthologs of those genes in invertebrates and a low but consistent similarity with some plant and eubacterial genes have also been found. We demonstrate that GDAP1 and GDAP1L1 do not belong to any of the known classes of GST genes. In addition to having distinctive sequences, GDAP1 and its relatives are also characterized by an extended region in GST domain II, absent in most other GSTs, and by a C-terminal end predicted to contain transmembrane domains. Mutations affecting any of those characteristic domains are known to cause Charcot-Marie-Tooth disease. These features define the GDAP1 class of GST-like proteins.
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Mutations in GDAP1: autosomal recessive CMT with demyelination and axonopathy.
Neurology, 2002Co-Authors: Eva Nelis, Francesc Palau, Laia Pedrola, Ana Cuesta, Sevim Erdem, P. Van Den Bergh, M.-c. Belpaire-dethiou, Chantal Ceuterick, V. Van Gerwen, A.a.w.m. Gabreëls-festenAbstract:Background: Mutations in the ganglioside-induced differentiation-associated protein 1 gene (GDAP1) were recently shown to be responsible for autosomal recessive (AR) demyelinating Charcot-Marie-Tooth disease (CMT) type 4A (CMT4A) as well as AR axonal CMT with vocal cord paralysis. Methods: The coding region of GDAP1 was screened for the presence of mutations in seven families with AR CMT in which the patients were homozygous for markers of the CMT4A locus at chromosome 8q21.1. Results: A nonsense mutation was detected in exon 5 (c.581C>G, S194X), a 1-bp deletion in exon 6 (c.786delG, G262fsX284), and a missense mutation in exon 6 (c.844C>T, R282C). Conclusions: Mutations in GDAP1 are a frequent cause of AR CMT. They result in an early-onset, severe clinical phenotype. The range of nerve conduction velocities (NCV) is variable. Some patients have normal or near normal NCV, suggesting an axonal neuropathy, whereas others have severely slowed NCV compatible with demyelination. The peripheral nerve biopsy findings are equally variable and show features of demyelination and axonal degeneration.
Victor Lopez Del Amo - One of the best experts on this subject based on the ideXlab platform.
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A Drosophila model of GDAP1 function reveals the involvement of insulin signalling in the mitochondria-dependent neuromuscular degeneration.
Biochimica et biophysica acta. Molecular basis of disease, 2017Co-Authors: Victor Lopez Del Amo, Martina Palomino-schätzlein, Marta Seco-cervera, José Luis García-giménez, Federico V. Pallardó, Antonio Pineda-lucena, Máximo Ibo GalindoAbstract:Abstract Charcot-Marie-Tooth disease is a rare peripheral neuropathy for which there is no specific treatment. Some forms of Charcot-Marie-Tooth are due to mutations in the GDAP1 gene. A striking feature of mutations in GDAP1 is that they have a variable clinical manifestation, according to disease onset and progression, histology and mode of inheritance. Studies in cellular and animal models have revealed a role of GDAP1 in mitochondrial morphology and distribution, calcium homeostasis and oxidative stress. To get a better understanding of the disease mechanism we have generated models of over-expression and RNA interference of the Drosophila GDAP1 gene. In order to get an overview about the changes that GDAP1 mutations cause in our disease model, we have combined a comprehensive determination of the metabolic profile in the flies by nuclear magnetic resonance spectroscopy with gene expression analyses and biophysical tests. Our results revealed that both up- and down-regulation of GDAP1 results in an early systemic inactivation of the insulin pathway before the onset of neuromuscular degeneration, followed by an accumulation of carbohydrates and an increase in the β-oxidation of lipids. Our findings are in line with emerging reports of energy metabolism impairments linked to different types of neural pathologies caused by defective mitochondrial function, which is not surprising given the central role of mitochondria in the control of energy metabolism. The relationship of mitochondrial dynamics with metabolism during neurodegeneration opens new avenues to understand the cause of the disease, and for the discovery of new biomarkers and treatments.
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nuevos modelos y mecanismos moleculares en la enfermedad de charcot marie tooth causada por mutaciones en GDAP1
2017Co-Authors: Victor Lopez Del AmoAbstract:Uno de los genes implicados en la enfermedad de Charcot Marie Tooth, una neuropatia periferica hereditaria, es GDAP1, que codifica para una proteina anclada a la membrana mitocondrial externa. El gen de Drosophila CG4623 es el ortologo de GDAP1 humano, y lo hemos renombrado como GDAP1. La sobreexpresion y silenciamiento de GDAP1, de manera tejido especifica, provoca una degeneracion neuronal y muscular. Ademas, se observan alteraciones en el tamano, morfologia y distribucion mitocondrial. El estudio de diferentes aspectos moleculares indica que los cambios en el estres oxidativo solo ocurren a largo plazo y no son una causa primaria. El estudio metabolomico a traves de la resonancia magnetica nuclear (RMN), revela que la sobreexpresion y silenciamiento de GDAP1 desencadenan una acumulacion de carbohidratos y un aumento en la β-oxidacion lipidica. Este cambio metabolico se explica por una atenuacion sistemica de la via de la insulina, que es probablemente causada por el contacto anormal de las mitocondrias y el reticulo endoplasmatico. Trasladamos nuestros resultados de Drosophila a cultivo neuronal de celulas de mamifero, y demostramos que la sobreexpresion de las formas mutadas de GDAP1 causa un desequilibrio de las proteinas relacionadas con la ruta de la insulina en condiciones basales. Ademas, demostramos que las proteinas mutadas de GDAP1 desencadenan un bloqueo de la respuesta a la insulina basada en la incapacidad de tomar la glucosa del medio despues del tratamiento con insulina.
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mitochondrial defects and neuromuscular degeneration caused by altered expression of drosophila GDAP1 implications for the charcot marie tooth neuropathy
Human Molecular Genetics, 2015Co-Authors: Victor Lopez Del Amo, Federico V. Pallardó, Máximo Ibo Galindo, Alexander J. Whitworth, Marta Secocervera, Jose Luis GarciagimenezAbstract:One of the genes involved in Charcot-Marie-Tooth (CMT) disease, an inherited peripheral neuropathy, is GDAP1. In this work, we show that there is a true ortholog of this gene in Drosophila, which we have named GDAP1. By up- and down-regulation of GDAP1 in a tissue-specific manner, we show that altering its levels of expression produces changes in mitochondrial size, morphology and distribution, and neuronal and muscular degeneration. Interestingly, muscular degeneration is tissue-autonomous and not dependent on innervation. Metabolic analyses of our experimental genotypes suggest that alterations in oxidative stress are not a primary cause of the neuromuscular degeneration but a long-term consequence of the underlying mitochondrial dysfunction. Our results contribute to a better understanding of the role of mitochondria in CMT disease and pave the way to generate clinically relevant disease models to study the relationship between mitochondrial dynamics and peripheral neurodegeneration.
Maribel Sanchezpiris - One of the best experts on this subject based on the ideXlab platform.
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charcot marie tooth related gene GDAP1 complements cell cycle delay at g2 m phase in saccharomyces cerevisiae fis1 gene defective cells
Journal of Biological Chemistry, 2011Co-Authors: Anna Estela, Francesc Palau, David Plamartin, Maribel Sanchezpiris, Hiromi SesakiAbstract:Abstract Mutations in the GDAP1 gene are responsible of the Charcot-Marie-Tooth CMT4A, ARCMT2K, and CMT2K variants. GDAP1 is a mitochondrial outer membrane protein that has been related to the fission pathway of the mitochondrial network dynamics. As mitochondrial dynamics is a conserved process, we reasoned that expressing GDAP1 in Saccharomyces cerevisiae strains defective for genes involved in mitochondrial fission or fusion could increase our knowledge of GDAP1 function. We discovered a consistent relation between Fis1p and the cell cycle because fis1Δ cells showed G2/M delay during cell cycle progression. The fis1Δ phenotype, which includes cell cycle delay, was fully rescued by GDAP1. By contrast, clinical missense mutations rescued the fis1Δ phenotype except for the cell cycle delay. In addition, both Fis1p and human GDAP1 interacted with β-tubulins Tub2p and TUBB, respectively. A defect in the fis1 gene may induce abnormal location of mitochondria during budding mitosis, causing the cell cycle delay at G2/M due to its anomalous interaction with microtubules from the mitotic spindle. In the case of neurons harboring defects in GDAP1, the interaction between mitochondria and the microtubule cytoskeleton would be altered, which might affect mitochondrial axonal transport and movement within the cell and may explain the pathophysiology of the GDAP1-related Charcot-Marie-Tooth disease.
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the charcot marie tooth related gene GDAP1 complements cell cycle delay at g2 m in s cerevisiae fis1 defective cells
Journal of Biological Chemistry, 2011Co-Authors: Anna Estela, David Plamartin, Maribel Sanchezpiris, Hiromi Sesaki, Francesc PalauAbstract:Abstract Mutations in the GDAP1 gene are responsible of the Charcot-Marie-Tooth CMT4A, ARCMT2K, and CMT2K variants. GDAP1 is a mitochondrial outer membrane protein that has been related to the fission pathway of the mitochondrial network dynamics. As mitochondrial dynamics is a conserved process, we reasoned that expressing GDAP1 in Saccharomyces cerevisiae strains defective for genes involved in mitochondrial fission or fusion could show some knowledge on GDAP1 function. We have discovered a consistent relation between Fis1p and the cell cycle as fis1Δ cells showed G2/M delay during the cell cycle progression. fis1Δ phenotype, which includes cell cycle delay, is fully rescued by GDAP1. By contrast, clinical missense mutations rescued fis1Δ phenotype except for the cell cycle delay. In addition, both Fis1p and the human GDAP1 interact with β-tubulins Tub2p and TUBB, respectively. Defect in the fis1 gene may induce abnormal location of mitochondria during budding mitosis causing the cell cycle delay at G2/M due to its anomalous interaction with microtubules from the mitotic spindle. In the case of neurons harboring defects in GDAP1, mitochondria and microtubule cytoskeleton interaction would be altered, which might affect mitochondrial axonal transport and movement within the cell, and may explain the pathophysiology of the Charcot-Marie-Tooth disease.
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charcot marie tooth related gene GDAP1 complements cell cycle delay at g 2 m phase insaccharomyces cerevisiae fis1
2011Co-Authors: Anna Estela, David Plamartin, Maribel Sanchezpiris, Hiromi Sesaki, Francesc PalauAbstract:Mutations in the GDAP1 gene are responsible of the CharcotMarie-Tooth CMT4A, ARCMT2K, and CMT2K variants. GDAP1 is a mitochondrial outer membrane protein that has been related to the fission pathway of the mitochondrial network dynamics. As mitochondrial dynamics is a conserved process, we reasoned that expressing GDAP1 in Saccharomyces cerevisiae strains defective for genes involved in mitochondrial fission or fusion could increase our knowledge of GDAP1 function. We discovered a consistent relation between Fis1p and the cell cycle because fis1 cells showed G2/M delay during cell cycle progression. The fis1 phenotype, which includes cell cycle delay, was fully rescued by GDAP1. By contrast, clinical missense mutations rescued the fis1 phenotype except for the cell cycle delay. In addition, both Fis1p and human GDAP1 interacted with -tubulins Tub2p and TUBB, respectively. A defect in the fis1 gene may induce abnormal location of mitochondria during budding mitosis, causing the cell cycle delay at G2/M due to its anomalous interaction with microtubules from the mitotic spindle. In the case of neurons harboring defects in GDAP1, the interaction between mitochondria and the microtubule cytoskeleton would be altered, which might affect mitochondrial axonal transport and movement within the cell and may explain the pathophysiology of the GDAP1-related Charcot-Marie-Tooth disease.
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cell expression of GDAP1 in the nervous system and pathogenesis of charcot marie tooth type 4a disease
Journal of Cellular and Molecular Medicine, 2008Co-Authors: Laia Pedrola, Antonio Espert, Erich E. Sirkowski, Steven S. Scherer, Isabel Fariñas, Teresa Valdessanchez, Maribel Sanchezpiris, Francesc PalauAbstract:Mutations in the mitochondrial protein GDAP1 are the cause of Charcot-Marie-Tooth type 4A disease (CMT4A), a severe form of peripheral neuropathy associated with either demyelinating, axonal or intermediate pheno-types. GDAP1 is located in the outer mitochondrial membrane and it seems that may be related with the mitochondrial network dynamics. We are interested to define cell expression in the nervous system and the effect of mutations in mitochondrial morphology and pathogenesis of the disease. We investigated GDAP1 expression in the nervous system and dorsal root ganglia (DRG) neuron cultures. GDAP1 is expressed in motor and sensory neurons of the spinal cord and other large neurons such as cerebellar Purkinje neurons, hippocampal pyramidal neurons, mitral neurons of the olfactory bulb and cortical pyramidal neurons. The lack of GDAP1 staining in the white matter and nerve roots suggested that glial cells do not express GDAP1. In DRG cultures satellite cells and Schwann cells were GDAP1-negative. Overexpression of GDAP1-induced fragmentation of mitochondria suggesting a role of GDAP1 in the fission pathway of the mitochondrial dynamics. Missense mutations showed two different patterns: most of them induced mitochondrial fragmentation but the T157P mutation showed an aggregation pattern. Whereas null mutations of GDAP1 should be associated with loss of function of the protein, missense mutations may act through different pathogenic mechanisms including a dominant-negative effect, suggesting that different molecular mechanisms may underlay the pathogenesis of CMT4A.
Anna Estela - One of the best experts on this subject based on the ideXlab platform.
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Phenotypical features of a new dominant GDAP1 pathogenic variant (p.R226del) in axonal Charcot-Marie-Tooth disease.
Neuromuscular disorders : NMD, 2017Co-Authors: Francesc Palau, Anna Estela, Carmen Espinós, Tania García-sobrino, Patricia Blanco-arias, Laura Ramirez, Beatriz San Millán, Manuel Arias, María-jesús Sobrido, Julio PardoAbstract:There are few reports on axonal CMT due to dominant GDAP1 mutations. We describe two unrelated Spanish families with a dominant axonal CMT. A novel in frame GAA deletion in exon 5 of the GDAP1 gene (c.677_679del; p.R226del) was identified in both families. Disease onset varied from early childhood to adulthood. Affected family members complained of distal lower limb weakness, cramps and foot deformities with variable CMTNS score in both families. Several individuals were asymptomatic or had paraesthesia only, however neurological examination and nerve conduction studies demonstrated neuropathic signs. Transfection of HeLa cells with the p.R226del mutation led to an increased mitochondrial aggregation. We report an AD-CMT2K with large phenotypic variability due to a novel dominant GDAP1 variant. This is the second founder GDAP1 pathogenic variant reported in Spain.
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Lack of GDAP1 induces neuronal calcium and mitochondrial defects in a knockout mouse model of Charcot-Marie-tooth neuropathy
PLoS genetics, 2015Co-Authors: Manuela Barneo-muñoz, Paula Juárez, Azahara Civera-tregón, Laura Yndriago, David Pla-martin, Jennifer Zenker, Carmen Cuevas-martín, Anna Estela, María Sánchez-aragó, Jerónimo Forteza-vilaAbstract:Mutations in GDAP1, which encodes protein located in the mitochondrial outer membrane, cause axonal recessive (AR-CMT2), axonal dominant (CMT2K) and demyelinating recessive (CMT4A) forms of Charcot-Marie-Tooth (CMT) neuropathy. Loss of function recessive mutations in GDAP1 are associated with decreased mitochondrial fission activity, while dominant mutations result in impairment of mitochondrial fusion with increased production of reactive oxygen species and susceptibility to apoptotic stimuli. GDAP1 silencing in vitro reduces Ca2+ inflow through store-operated Ca2+ entry (SOCE) upon mobilization of endoplasmic reticulum (ER) Ca2+, likely in association with an abnormal distribution of the mitochondrial network. To investigate the functional consequences of lack of GDAP1 in vivo, we generated a GDAP1 knockout mouse. The affected animals presented abnormal motor behavior starting at the age of 3 months. Electrophysiological and biochemical studies confirmed the axonal nature of the neuropathy whereas histopathological studies over time showed progressive loss of motor neurons (MNs) in the anterior horn of the spinal cord and defects in neuromuscular junctions. Analyses of cultured embryonic MNs and adult dorsal root ganglia neurons from affected animals demonstrated large and defective mitochondria, changes in the ER cisternae, reduced acetylation of cytoskeletal α-tubulin and increased autophagy vesicles. Importantly, MNs showed reduced cytosolic calcium and SOCE response. The development and characterization of the GDAP1 neuropathy mice model thus revealed that some of the pathophysiological changes present in axonal recessive form of the GDAP1-related CMT might be the consequence of changes in the mitochondrial network biology and mitochondria–endoplasmic reticulum interaction leading to abnormalities in calcium homeostasis.
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Generation of GDAP1-/- mice.
2015Co-Authors: Manuela Barneo-muñoz, Paula Juárez, Azahara Civera-tregón, Laura Yndriago, David Pla-martin, Jennifer Zenker, Carmen Cuevas-martín, Anna Estela, María Sánchez-aragó, Jerónimo Forteza-vilaAbstract:(A) Schematic representation of GDAP1-/- targeting strategy. Diagram is not to scale. Hatched rectangles represent GDAP1 exons 1 to 6, solid line represents mouse chromosome 1. FRT sites are represented by double triangles and loxP sites are right-faced triangles. (B) GDAP1 protein expression was assessed by immunoblotting of selected tissue homogenates prepared from 2 months-old wild-type (WT), GDAP1+/- (+/-) and GDAP1-/- (-/-) mice.
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Lack of GDAP1 leads to loss of motor neurons and abnormal neuromuscular junctions.
2015Co-Authors: Manuela Barneo-muñoz, Paula Juárez, Azahara Civera-tregón, Laura Yndriago, David Pla-martin, Jennifer Zenker, Carmen Cuevas-martín, Anna Estela, María Sánchez-aragó, Jerónimo Forteza-vilaAbstract:(A) Anterior horns from lumbar spinal cord of 5 and 12-months-old mice were stained by Nissl staining. Reduced number of MNs and evidence of chromatolysis are visible in GDAP1-/- mice. (B) Progressive graphics representing the number of healthy motor neurons in anterior horns per section in WT (black line) and GDAP1-/- (gray line) mice at several ages (n = 3). (C) Representative confocal stack images of NMJs from the gastrocnemius muscle. Axons were immunostained with anti-β-III tubulin (β-III tub, green) and the postsynaptic acetyl-choline receptor was stained with AF488-coupled α-bungarotoxin (BTX, red). (D) Histograms show the percentage of NMJ occupancy by terminal axons in WT (black bars) and GDAP1-/- (gray bars) mice. (E) Magnification of tangle-like abnormal structures (white arrows) at the terminal axons closed to the NMJ observed in GDAP1-/- mice muscles. * represents significant differences between WT and GDAP1-/- mice; & indicates differences between ages of WT mice; and # indicates differences between ages of GDAP1-/- animals (Student’s t test, data are presented as means ±S.E.M.).
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Behavioural testing and electrophysiological measurements on GDAP1-/- mice.
2015Co-Authors: Manuela Barneo-muñoz, Paula Juárez, Azahara Civera-tregón, Laura Yndriago, David Pla-martin, Jennifer Zenker, Carmen Cuevas-martín, Anna Estela, María Sánchez-aragó, Jerónimo Forteza-vilaAbstract:(A) Upper panel shows photographs of 3 months-old mice suspended by its tail. WT mice show a characteristic response trying to escape by splaying its hind limbs away from the trunk of its body. In contrast, hind limbs of GDAP1-/- mice are held tonically against its trunk in an abnormal dystonic posture. Lower panels display a low body position and a dragging tail present in GDAP1-/- mice as compared to age-matched WT mice. (B) Motor coordination was assessed by rotarod test, (n = 10 for each genotype and at each age group). (C) Representative hind limb walking patterns of 5 months-old WT and GDAP1-/- mice where the stride length (SL) and stride angle (SA) have been depicted. Footprints revealed that GDAP1-/- mice walk with an abnormal gait. The scheme of a hindpaw footprint indicating measured parameters (PL: plantar length; TS: toe spreading) has been included. (D) Quantification of various parameters obtained from the gait analysis of WT (black columns) and GDAP1-/- (grey columns) animals at 5 and 12 months of age. Upper graphs show stride length (left) and stride angle (right). Lower graphs show the quantitative analysis of the hindpaw footprint parameters toe spreading (left) and plantar length (right). Analysis was conducted on 10 clearly visible footprints at 5 animals per genotype. Determination of sciatic nerve compound muscle action potential (CMAP) amplitudes at both distal and proximal (E) as well as motor nerve conduction velocities (MNCV) (F) measured in WT and GDAP1-/- mice at 2 and 5 months of age (n = 4). Error bars indicate standard error of the mean (S.E.M.). p values were calculated using Student's t test,*p
