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O.2 Fukutin mutations in congenital muscular dystrophies with defective glycosylation of dystroglycan in Korea
Abstracts / Neuromuscular Disorders 20 (2010) 596–680
retardation and a neuronal migration defect on brain magnetic resonance imaging are candidates for a carbohydrate deficient glycoprotein syndrome in the form of O-mannosylation defect, called ‘‘the dystroglycanopathies”. Muscle biopsy clearly shows reduction or absence of a-dystroglycan. Fukuyama CMD, Walker Warburg syndrome, Muscle Eye Brain disease, MDC1C and MDC1D, are primarily due to mutations in FCMD, POMT1, POMGnT1, FKRP and LARGE, respectively. There is more to that now. Almost all of these six genes can be responsible from a spectrum of severe to mild cases. We have reported a novel form of limb girdle muscular dystrophy with mild mental retardation due to mutations in the POMT1 gene. As the third group, mutations in SEPN1, encoding an endoplasmic reticulum resident selenoprotein of unknown function, result in CMD with spinal rigidity (RSMD1). CMD may be a component in some cases with the Marinescu–Sjogren syndrome. Recent identification of SIL1 mutations implicates this disorder as a disease of endoplasmic reticulum. Apart form the above pretty well described conditions there are others such as cases with mental retardation, ichthyosis and bizarre looking mitochondria in the muscle biopsies. It may be a logical estimation that there will be new clinical descriptions within CMD. doi:10.1016/j.nmd.2010.07.008
CONGENITAL MUSCULAR DYSTROPHIES (CELEBRATING THE 50TH ANNIVERSARY OF FUKUYAMA CONGENITAL MUSCULAR DYSTROPHY) 2; INVITED LECTURES, ORAL PRESENTATIONS C.I.4 Molecular therapeutic approaches to the extracellular matrixrelated congenital muscular dystrophies C.G. Bönnemann Neurogenetics Branch, NIH/NINDS, Bethesda, United States I will discuss molecular therapeutic approaches and their clinical potential in the major forms congenital muscular dystrophies (CMD) that affect the interaction of the muscle fibers with the extracellular matrix: ,erosin deficient CMD caused by mutations in LAMA2, the alpha-dystroglycanopathies caused by mutations in genes involved in O-mannosyl linked glycosylation of alpha-dystroglycan, and the collagen VI related CMDs, caused by mutations in COL6A1, A2, and A3. In merosin deficient CMD genetic as well as pharmacological approaches have been developed to counteract myofiber apoptosis, while the introduction of a mini-agrin construct to bridge the defect has been proposed as a gene-therapeutic approach. Protein therapy and stem cell transplantation are also under preclinical investigation. In the alpha-dystroglycanopathies it was recognized that up-regulation of LARGE as a ‘‘universal” glycosylation facilitator carries the potential for ameliorating the majority of the various alpha dystroglycanopathies – so far using gene transfer, but pharmacological means of LARGE up-regulation seem feasible. Up-regulation of other glycosyltransferases and direct gene replacement therapy are being developed also. For the collagen VI related myopathies approaches have included pharmacological counteraction of myofiber apoptosis mediated by dysfunction of the mitochondrial permeability transition pore. Since the majority of mutations in the collagen VI genes (COL6A1, A2, A3) are dominantly acting, efforts are directed at eliminating the dominantly acting transcript that interferes with the wildtype allele. Some approaches are independent of the primary genetic defect and may therefore have broader applicability, such as those directed at promoting muscle growth and regeneration and the prevention of fibrosis. doi:10.1016/j.nmd.2010.07.009
597
O.1 Novel O-mannosyl phosphorylation of alpha-dystroglycan is required for laminin binding: implications for congenital muscular dystrophy T. Moriguchi 1, L. Yu 1, H. Schachter 2, L. Wells 3, K. Campbell 1 1
University of Iowa, Iowa City, United States, 2 The Hospital for Sick Children, Toronto, Canada, 3 University of Georgia, Athens, United States Alpha-dystroglycan (alpha-DG) is a ubiquitously expressed cellsurface glycoprotein that serves as a high affinity laminin receptor. Anchoring of the basal lamina to the sarcolemma via alpha-DG is essential for maintaining the sarcolemmal integrity and protecting muscle from contraction-induced injury. A growing number of muscular dystrophies are associated with perturbations of the lamininbinding ability of alpha-DG. Collectively, these muscular dystrophies are called dystroglycanopathies. In these disorders, mutations in known or putative glycosyltransferase genes cause aberrant posttranslational modification of alpha-DG, which leads to the loss of its laminin receptor ability. Nevertheless, the modification pathway, which enables alpha-DG to bind laminin, is largely unknown. Here we show that the alpha-DG laminin receptor function requires phosphorylation of an O-mannosyl glycan, and that laminin-binding moiety is assembled onto the phosphoryl residue. Using mass spectrometry and NMR-based structural analyses, we identified a novel phosphorylated O-glycan [GalNAcb1-3GlcNAcb1-4(PO4-6)Man] on the mucin-like domain of recombinant alpha-DG. Additional studies showed that alpha-DG is phosphorylated within the Golgi complex and that this phosphorylation occurs independently of the mannose-6-phosphate synthetic pathway that is required for lysosomal protein modification. We also demonstrated that patients with muscle–eye–brain disease and Fukuyama congenital muscular dystrophy, and myodystrophy mice (Largemyd) are defective in a postphosphoryl modification of this phosphorylated O-linked mannose, and that this modification is mediated by the like-acetylglucosaminyltransferase (LARGE) protein. These convergent mechanisms to pathology explain how the forced expression of LARGE circumvents defects in alpha-DG modification in cells from patients with these congenital muscular dystrophies, as reported previously (Barresi R. et al., Nat Med. 10:696–703, 2000).
doi:10.1016/j.nmd.2010.07.010
O.2 Fukutin mutations in congenital muscular dystrophies with defective glycosylation of dystroglycan in Korea B.C.L. Lim 1, J.H.C. Chae 1, A.C. Cho 1, C.S.K. Ki 2 1
Seoul National University Hospital, Department of pediatrics, Seoul, Republic of Korea, 2 Samsung Medical Center, Seoul, Republic of Korea Background: The aim of this study was to identify Fukutin (FKTN)related congenital muscular dystrophies (CMD) with defective adystroglycan (a-DG) glycosylation in Korea and to discuss their genotype–phenotype spectrum focusing on detailed brain magnetic resonance imaging (MRI) findings. Methods: Twelve patients were analyzed. Eleven patients were confirmed as having hypoglycosylation of a-DG on muscle biopsy and one patient was highly suggestive of CMD with defective glycosylation, based on clinical features alone. Results: FKTN mutations were found in nine of the 12 patients (75%). Two patients were homozygous for the Japanese founder
598
Abstracts / Neuromuscular Disorders 20 (2010) 596–680
retrotransposal (RT) insertion mutation. Seven patients were heterozygous for the RT insertion mutation, five of whom carried a novel intronic mutation that activates a pseudoexon between exons 5 and 6 (c.647 + 2084G > T). Compared with individuals that were homozygous for the RT insertion mutation, the seven heterozygotes for the RT insertion mutation, including five patients with the novel pseudoexon mutation, exhibited a more severe clinical phenotype in terms of motor abilities and more extensive brain MRI abnormalities (i.e., a wider distribution of cortical malformation, pons and cerebellar hypoplasia, and more frequent diffuse white matter changes and ventricular dilatation). Conclusions FKTN mutations are the most common genetic cause of CMD with defective a-DG glycosylation in Korea. Compound heterozygosity of the RT insertion and the novel pseudoexon mutation is the most prevalent genotype in Korea and is associated with a more severe clinical phenotype and a wider extent of brain MRI abnormalities compared with homozygosity for the RT insertion mutation. doi:10.1016/j.nmd.2010.07.011
O.3 Satellite cell loss is the pathomechanism leading to muscle atrophy in selenoprotein N deficiency P. Castets 1, A.T. Bertrand 1, M. Beuvin 1, A. Ferry 1, F. Le Grand 2, M. Castets 3, G. Chazot 3, M. Rederstorff 4, A. Krol 4, A. Lescure 4, N.B. Romero 1, P. Guicheney 5, V. Allamand 1 1
UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS, UMR7215, and Inserm, U974, Paris, France, 2 Institut Cochin, Université Paris Descartes, CNRS, UMR8104, and Inserm, U1016, Paris, France, 3 CNRS, UMR5238, Université de Lyon, Centre Léon Bérard, Lyon, France, 4 Architecture et Réactivité de l’ARN, Université de Strasbourg, CNRS, IBMC, Strasbourg, France, 5 UPMC Univ Paris 06, IFR14, and Inserm, U956, Paris, France Selenoprotein N (SelN) deficiency is responsible for a group of inherited neuromuscular disorders termed SEPN1-Related Myopathies. These congenital diseases are characterized by an early onset generalized muscle atrophy and weakness leading to spinal rigidity, severe scoliosis and respiratory insufficiency. Although the function of SelN remains unknown, recent data demonstrated that it is dispensable for mouse embryogenesis and suggested its involvement in the regulation of ryanodine receptors and/or cellular redox homeostasis. Here, we investigate the role of SelN in satellite cell dynamics taking advantage of the Sepn1 knock-out mouse model. We show that SelN deficiency results in a basal defect in the satellite cell population in adult skeletal muscle that does not impede muscle regeneration following a single cardiotoxin-induced injury. Subsequently, however, an almost complete depletion of the satellite cell pool was detected and prevented further regeneration of Sepn1 / muscles. Using isolated single fibres, we demonstrate that SelN deficiency alters the satellite cell fate choice, leading to a reduced number of self-renewing cells. Moreover, an increased proliferation of Sepn1 / muscle precursors was detected both in vivo and in vitro. Most importantly, exhaustion of the satellite cell population was also uncovered in muscle biopsies from SelN-deficient patients, emphasizing the essential role of SelN for satellite cell homeostasis also in human. In conclusion, we describe for the first time a major physiological function of SelN in skeletal muscle, as a key regulator of satellite cell maintenance. We propose that this defect is responsible for the characteristic early onset muscle atrophy in SEPN1-Related Myopathies, by reducing the pool of muscle progenitors participating in muscle growth. doi:10.1016/j.nmd.2010.07.012
O.4 Mouse model of LMNA-congenital muscular dystrophy shows severe skeletal and cardiac muscle maturation defects associated with major metabolic defects leading to early death A.T. Bertrand 1, L. Renou 1, M. Beuvin 1, A. Angelini 1, E. Lacène 1, T. Arimura 1, Y. Gruenbaum 2, G. Bonne 3 1
UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS UMR7215, Inserm U974, Paris, France, 2 Department of Genetics, The Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel, 3 UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS UMR7215, Inserm U974, APHP Pitié-Salpêtrière, Paris, France LMNA gene encodes for lamins A/C, ubiquitous proteins of the nuclear envelope. Lamins A/C are thought to have structural but also essential regulatory roles in various signalization pathways by interaction with transcription factors such as Sterol Regulatory Element Binding Protein 1 (SREBP1). LMNA mutations are responsible for more than 10 different disorders affecting various tissues in isolated or systemic fashion. Among those the LMNA-related congenital muscular dystrophy (L-CMD) is characterized by early onset of muscle weakness and joint contractures. To get insight on the disease pathophysiology, we reproduced in mice the Lmna Lys32 deletion found in L-CMD patients. Homozygous mice were born with expected Mendelian ratios but shortly after birth developed general growth retardation and died within the 2nd week of life with severe hypoglycemia. This is associated with 80% decrease of mutated lamin A/C expression despite normal mRNA levels. Analysis of lamin A/C filaments in vitro in C. elegans showed altered polymerization of mutant proteins. Analysis of mouse organs revealed a maturation defect of skeletal and cardiac muscle but also of organs involved in lipid and glucose storage and release (pancreas, white adipose tissue). Interestingly, mutant liver retains high glycogen levels. Considering the involvement of SREBP1 in the maturation of white adipose tissue and pancreas and its role in glucose release from glycogen storage in the liver, we checked the activation of this pathway and the expression of its target genes in affected tissues. Preliminary data showed an upregulation of active SREBP1 in skeletal muscle and liver. Thus the deletion of Lys32 localized in the dimerization domain, destabilizes the mutated proteins that are partly degraded. Decreased protein level plus abnormally polymerized lamin A/C may modify the multiple lamin A/C interactions with as consequence abnormal SREBP1 signaling inducing major metabolic defects leading to early mouse death.
doi:10.1016/j.nmd.2010.07.013
O.5 Skeletal muscle-specific calpain, p94/calpain 3, dynamically distributes in skeletal muscle cells to adapt to physical stress, defects of which cause muscular dystrophy K. Ojima 1, Y. Ono 1, N. Doi 1, F. Kitamura 1, S. Hata 1, Y. Kawabata 2, K. Suzuki 2, T. Maeda 2, K. Abe 2, H. Nakao 2, A. Aiba 2, K. Nakao 3, H. Suzuki 4, H. Kawahara 5, C. Witt 6, S. Labeit 6, C. Ottenheijm 7, H. Granzier 7, N. Toyama-Sorimachi 8, M. Sorimachi 9, H. Sorimachi 1 1 The Tokyo Metropolitan Institute of Medical Science (Rinshoken), Calpain Project, Tokyo, Japan, 2 The University of Tokyo, Tokyo, Japan, 3 Riken, Kobe, Japan, 4 The Tokyo Metropolitan Institute of Medical Science (Rinshoken), EM Lab, Tokyo, Japan, 5 Tokyo Metropolitan University, Tokyo, Japan, 6 Universitätsklinikum Mannheim, Mannheim, Germany, 7 University of Arizona, Tucson, United States, 8 IMCJ, Research Institute, Tokyo, Japan, 9 Polytechnic University, The Institute of Research and Development, Sagamihara, Japan
retardation and a neuronal migration defect on brain magnetic resonance imaging are candidates for a carbohydrate deficient glycoprotein syndrome in the form of O-mannosylation defect, called ‘‘the dystroglycanopathies”. Muscle biopsy clearly shows reduction or absence of a-dystroglycan. Fukuyama CMD, Walker Warburg syndrome, Muscle Eye Brain disease, MDC1C and MDC1D, are primarily due to mutations in FCMD, POMT1, POMGnT1, FKRP and LARGE, respectively. There is more to that now. Almost all of these six genes can be responsible from a spectrum of severe to mild cases. We have reported a novel form of limb girdle muscular dystrophy with mild mental retardation due to mutations in the POMT1 gene. As the third group, mutations in SEPN1, encoding an endoplasmic reticulum resident selenoprotein of unknown function, result in CMD with spinal rigidity (RSMD1). CMD may be a component in some cases with the Marinescu–Sjogren syndrome. Recent identification of SIL1 mutations implicates this disorder as a disease of endoplasmic reticulum. Apart form the above pretty well described conditions there are others such as cases with mental retardation, ichthyosis and bizarre looking mitochondria in the muscle biopsies. It may be a logical estimation that there will be new clinical descriptions within CMD. doi:10.1016/j.nmd.2010.07.008
CONGENITAL MUSCULAR DYSTROPHIES (CELEBRATING THE 50TH ANNIVERSARY OF FUKUYAMA CONGENITAL MUSCULAR DYSTROPHY) 2; INVITED LECTURES, ORAL PRESENTATIONS C.I.4 Molecular therapeutic approaches to the extracellular matrixrelated congenital muscular dystrophies C.G. Bönnemann Neurogenetics Branch, NIH/NINDS, Bethesda, United States I will discuss molecular therapeutic approaches and their clinical potential in the major forms congenital muscular dystrophies (CMD) that affect the interaction of the muscle fibers with the extracellular matrix: ,erosin deficient CMD caused by mutations in LAMA2, the alpha-dystroglycanopathies caused by mutations in genes involved in O-mannosyl linked glycosylation of alpha-dystroglycan, and the collagen VI related CMDs, caused by mutations in COL6A1, A2, and A3. In merosin deficient CMD genetic as well as pharmacological approaches have been developed to counteract myofiber apoptosis, while the introduction of a mini-agrin construct to bridge the defect has been proposed as a gene-therapeutic approach. Protein therapy and stem cell transplantation are also under preclinical investigation. In the alpha-dystroglycanopathies it was recognized that up-regulation of LARGE as a ‘‘universal” glycosylation facilitator carries the potential for ameliorating the majority of the various alpha dystroglycanopathies – so far using gene transfer, but pharmacological means of LARGE up-regulation seem feasible. Up-regulation of other glycosyltransferases and direct gene replacement therapy are being developed also. For the collagen VI related myopathies approaches have included pharmacological counteraction of myofiber apoptosis mediated by dysfunction of the mitochondrial permeability transition pore. Since the majority of mutations in the collagen VI genes (COL6A1, A2, A3) are dominantly acting, efforts are directed at eliminating the dominantly acting transcript that interferes with the wildtype allele. Some approaches are independent of the primary genetic defect and may therefore have broader applicability, such as those directed at promoting muscle growth and regeneration and the prevention of fibrosis. doi:10.1016/j.nmd.2010.07.009
597
O.1 Novel O-mannosyl phosphorylation of alpha-dystroglycan is required for laminin binding: implications for congenital muscular dystrophy T. Moriguchi 1, L. Yu 1, H. Schachter 2, L. Wells 3, K. Campbell 1 1
University of Iowa, Iowa City, United States, 2 The Hospital for Sick Children, Toronto, Canada, 3 University of Georgia, Athens, United States Alpha-dystroglycan (alpha-DG) is a ubiquitously expressed cellsurface glycoprotein that serves as a high affinity laminin receptor. Anchoring of the basal lamina to the sarcolemma via alpha-DG is essential for maintaining the sarcolemmal integrity and protecting muscle from contraction-induced injury. A growing number of muscular dystrophies are associated with perturbations of the lamininbinding ability of alpha-DG. Collectively, these muscular dystrophies are called dystroglycanopathies. In these disorders, mutations in known or putative glycosyltransferase genes cause aberrant posttranslational modification of alpha-DG, which leads to the loss of its laminin receptor ability. Nevertheless, the modification pathway, which enables alpha-DG to bind laminin, is largely unknown. Here we show that the alpha-DG laminin receptor function requires phosphorylation of an O-mannosyl glycan, and that laminin-binding moiety is assembled onto the phosphoryl residue. Using mass spectrometry and NMR-based structural analyses, we identified a novel phosphorylated O-glycan [GalNAcb1-3GlcNAcb1-4(PO4-6)Man] on the mucin-like domain of recombinant alpha-DG. Additional studies showed that alpha-DG is phosphorylated within the Golgi complex and that this phosphorylation occurs independently of the mannose-6-phosphate synthetic pathway that is required for lysosomal protein modification. We also demonstrated that patients with muscle–eye–brain disease and Fukuyama congenital muscular dystrophy, and myodystrophy mice (Largemyd) are defective in a postphosphoryl modification of this phosphorylated O-linked mannose, and that this modification is mediated by the like-acetylglucosaminyltransferase (LARGE) protein. These convergent mechanisms to pathology explain how the forced expression of LARGE circumvents defects in alpha-DG modification in cells from patients with these congenital muscular dystrophies, as reported previously (Barresi R. et al., Nat Med. 10:696–703, 2000).
doi:10.1016/j.nmd.2010.07.010
O.2 Fukutin mutations in congenital muscular dystrophies with defective glycosylation of dystroglycan in Korea B.C.L. Lim 1, J.H.C. Chae 1, A.C. Cho 1, C.S.K. Ki 2 1
Seoul National University Hospital, Department of pediatrics, Seoul, Republic of Korea, 2 Samsung Medical Center, Seoul, Republic of Korea Background: The aim of this study was to identify Fukutin (FKTN)related congenital muscular dystrophies (CMD) with defective adystroglycan (a-DG) glycosylation in Korea and to discuss their genotype–phenotype spectrum focusing on detailed brain magnetic resonance imaging (MRI) findings. Methods: Twelve patients were analyzed. Eleven patients were confirmed as having hypoglycosylation of a-DG on muscle biopsy and one patient was highly suggestive of CMD with defective glycosylation, based on clinical features alone. Results: FKTN mutations were found in nine of the 12 patients (75%). Two patients were homozygous for the Japanese founder
598
Abstracts / Neuromuscular Disorders 20 (2010) 596–680
retrotransposal (RT) insertion mutation. Seven patients were heterozygous for the RT insertion mutation, five of whom carried a novel intronic mutation that activates a pseudoexon between exons 5 and 6 (c.647 + 2084G > T). Compared with individuals that were homozygous for the RT insertion mutation, the seven heterozygotes for the RT insertion mutation, including five patients with the novel pseudoexon mutation, exhibited a more severe clinical phenotype in terms of motor abilities and more extensive brain MRI abnormalities (i.e., a wider distribution of cortical malformation, pons and cerebellar hypoplasia, and more frequent diffuse white matter changes and ventricular dilatation). Conclusions FKTN mutations are the most common genetic cause of CMD with defective a-DG glycosylation in Korea. Compound heterozygosity of the RT insertion and the novel pseudoexon mutation is the most prevalent genotype in Korea and is associated with a more severe clinical phenotype and a wider extent of brain MRI abnormalities compared with homozygosity for the RT insertion mutation. doi:10.1016/j.nmd.2010.07.011
O.3 Satellite cell loss is the pathomechanism leading to muscle atrophy in selenoprotein N deficiency P. Castets 1, A.T. Bertrand 1, M. Beuvin 1, A. Ferry 1, F. Le Grand 2, M. Castets 3, G. Chazot 3, M. Rederstorff 4, A. Krol 4, A. Lescure 4, N.B. Romero 1, P. Guicheney 5, V. Allamand 1 1
UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS, UMR7215, and Inserm, U974, Paris, France, 2 Institut Cochin, Université Paris Descartes, CNRS, UMR8104, and Inserm, U1016, Paris, France, 3 CNRS, UMR5238, Université de Lyon, Centre Léon Bérard, Lyon, France, 4 Architecture et Réactivité de l’ARN, Université de Strasbourg, CNRS, IBMC, Strasbourg, France, 5 UPMC Univ Paris 06, IFR14, and Inserm, U956, Paris, France Selenoprotein N (SelN) deficiency is responsible for a group of inherited neuromuscular disorders termed SEPN1-Related Myopathies. These congenital diseases are characterized by an early onset generalized muscle atrophy and weakness leading to spinal rigidity, severe scoliosis and respiratory insufficiency. Although the function of SelN remains unknown, recent data demonstrated that it is dispensable for mouse embryogenesis and suggested its involvement in the regulation of ryanodine receptors and/or cellular redox homeostasis. Here, we investigate the role of SelN in satellite cell dynamics taking advantage of the Sepn1 knock-out mouse model. We show that SelN deficiency results in a basal defect in the satellite cell population in adult skeletal muscle that does not impede muscle regeneration following a single cardiotoxin-induced injury. Subsequently, however, an almost complete depletion of the satellite cell pool was detected and prevented further regeneration of Sepn1 / muscles. Using isolated single fibres, we demonstrate that SelN deficiency alters the satellite cell fate choice, leading to a reduced number of self-renewing cells. Moreover, an increased proliferation of Sepn1 / muscle precursors was detected both in vivo and in vitro. Most importantly, exhaustion of the satellite cell population was also uncovered in muscle biopsies from SelN-deficient patients, emphasizing the essential role of SelN for satellite cell homeostasis also in human. In conclusion, we describe for the first time a major physiological function of SelN in skeletal muscle, as a key regulator of satellite cell maintenance. We propose that this defect is responsible for the characteristic early onset muscle atrophy in SEPN1-Related Myopathies, by reducing the pool of muscle progenitors participating in muscle growth. doi:10.1016/j.nmd.2010.07.012
O.4 Mouse model of LMNA-congenital muscular dystrophy shows severe skeletal and cardiac muscle maturation defects associated with major metabolic defects leading to early death A.T. Bertrand 1, L. Renou 1, M. Beuvin 1, A. Angelini 1, E. Lacène 1, T. Arimura 1, Y. Gruenbaum 2, G. Bonne 3 1
UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS UMR7215, Inserm U974, Paris, France, 2 Department of Genetics, The Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel, 3 UPMC Univ Paris 06, IFR14, Institut de Myologie, CNRS UMR7215, Inserm U974, APHP Pitié-Salpêtrière, Paris, France LMNA gene encodes for lamins A/C, ubiquitous proteins of the nuclear envelope. Lamins A/C are thought to have structural but also essential regulatory roles in various signalization pathways by interaction with transcription factors such as Sterol Regulatory Element Binding Protein 1 (SREBP1). LMNA mutations are responsible for more than 10 different disorders affecting various tissues in isolated or systemic fashion. Among those the LMNA-related congenital muscular dystrophy (L-CMD) is characterized by early onset of muscle weakness and joint contractures. To get insight on the disease pathophysiology, we reproduced in mice the Lmna Lys32 deletion found in L-CMD patients. Homozygous mice were born with expected Mendelian ratios but shortly after birth developed general growth retardation and died within the 2nd week of life with severe hypoglycemia. This is associated with 80% decrease of mutated lamin A/C expression despite normal mRNA levels. Analysis of lamin A/C filaments in vitro in C. elegans showed altered polymerization of mutant proteins. Analysis of mouse organs revealed a maturation defect of skeletal and cardiac muscle but also of organs involved in lipid and glucose storage and release (pancreas, white adipose tissue). Interestingly, mutant liver retains high glycogen levels. Considering the involvement of SREBP1 in the maturation of white adipose tissue and pancreas and its role in glucose release from glycogen storage in the liver, we checked the activation of this pathway and the expression of its target genes in affected tissues. Preliminary data showed an upregulation of active SREBP1 in skeletal muscle and liver. Thus the deletion of Lys32 localized in the dimerization domain, destabilizes the mutated proteins that are partly degraded. Decreased protein level plus abnormally polymerized lamin A/C may modify the multiple lamin A/C interactions with as consequence abnormal SREBP1 signaling inducing major metabolic defects leading to early mouse death.
doi:10.1016/j.nmd.2010.07.013
O.5 Skeletal muscle-specific calpain, p94/calpain 3, dynamically distributes in skeletal muscle cells to adapt to physical stress, defects of which cause muscular dystrophy K. Ojima 1, Y. Ono 1, N. Doi 1, F. Kitamura 1, S. Hata 1, Y. Kawabata 2, K. Suzuki 2, T. Maeda 2, K. Abe 2, H. Nakao 2, A. Aiba 2, K. Nakao 3, H. Suzuki 4, H. Kawahara 5, C. Witt 6, S. Labeit 6, C. Ottenheijm 7, H. Granzier 7, N. Toyama-Sorimachi 8, M. Sorimachi 9, H. Sorimachi 1 1 The Tokyo Metropolitan Institute of Medical Science (Rinshoken), Calpain Project, Tokyo, Japan, 2 The University of Tokyo, Tokyo, Japan, 3 Riken, Kobe, Japan, 4 The Tokyo Metropolitan Institute of Medical Science (Rinshoken), EM Lab, Tokyo, Japan, 5 Tokyo Metropolitan University, Tokyo, Japan, 6 Universitätsklinikum Mannheim, Mannheim, Germany, 7 University of Arizona, Tucson, United States, 8 IMCJ, Research Institute, Tokyo, Japan, 9 Polytechnic University, The Institute of Research and Development, Sagamihara, Japan