RESUMEN
Mutations of the survival of motor neuron gene (SMN1) are responsible for spinal muscular atrophies (SMA), a frequent recessive autosomal motor neuron disease. SMN is involved in various processes including RNA metabolism. However, the molecular pathway linking marked deficiency of SMN to SMA phenotype remains unclear. Homozygous deletion of murine Smn exon 7 directed to neurons or skeletal muscle causes severe motor axonal or myofiber degeneration, respectively. With the use of cDNA microarrays, expression profiles of 8,400 genes were analyzed in skeletal muscle and spinal cord of muscular and neuronal mutants, respectively, and compared with age-matched controls. A high proportion of genes (20 of 429, 5%) was involved in pre-mRNA splicing, ribosomal RNA processing, or RNA decay, and 18 of them were upregulated in mutant tissues. By analyzing other neuromuscular disorders, we showed that most of them (14 of 18) were specific to the SMN defect. Quantitative PCR analysis of these transcripts showed that gene activation was an early adaptive response to the lack but not reduced amount of full-length SMN in mouse mutant tissues. In human SMA tissues, activation of this program was not observed, which could be ascribed to the reduction but not the absence of full-length SMN.
Asunto(s)
Proteína de Unión a Elemento de Respuesta al AMP Cíclico/deficiencia , Proteínas del Tejido Nervioso/deficiencia , Estabilidad del ARN/genética , ARN/metabolismo , Animales , Biomarcadores , Estudios de Casos y Controles , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/genética , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Feto/metabolismo , Regulación de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Atrofia Muscular Espinal/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , ARN/genética , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Proteínas del Complejo SMN , Proteína 1 para la Supervivencia de la Neurona Motora , Activación TranscripcionalRESUMEN
Mutations in SPG4, encoding the microtubule-severing protein spastin, are responsible for the most frequent form of hereditary spastic paraplegia (HSP), a heterogeneous group of genetic diseases characterized by degeneration of the corticospinal tracts. We previously reported that mice harboring a deletion in Spg4, generating a premature stop codon, develop progressive axonal degeneration characterized by focal axonal swellings associated with impaired axonal transport. To further characterize the molecular and cellular mechanisms underlying this mutant phenotype, we have assessed microtubule dynamics and axonal transport in primary cultures of cortical neurons from spastin-mutant mice. We show an early and marked impairment of microtubule dynamics all along the axons of spastin-deficient cortical neurons, which is likely to be responsible for the occurrence of axonal swellings and cargo stalling. Our analysis also reveals that a modulation of microtubule dynamics by microtubule-targeting drugs rescues the mutant phenotype of cortical neurons. Together, these results contribute to a better understanding of the pathogenesis of SPG4-linked HSP and ascertain the influence of microtubule-targeted drugs on the early axonal phenotype in a mouse model of the disease.
Asunto(s)
Adenosina Trifosfatasas/deficiencia , Adenosina Trifosfatasas/genética , Animales , Transporte Axonal , Axones/efectos de los fármacos , Axones/patología , Células Cultivadas , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Modelos Animales de Enfermedad , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Ratones , Ratones Noqueados , Microtúbulos/efectos de los fármacos , Microtúbulos/metabolismo , Modelos Neurológicos , Mutación , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Nocodazol/farmacología , Paclitaxel/farmacología , Paraplejía Espástica Hereditaria/tratamiento farmacológico , Paraplejía Espástica Hereditaria/genética , Paraplejía Espástica Hereditaria/metabolismo , Paraplejía Espástica Hereditaria/patología , Espastina , Vinblastina/farmacologíaRESUMEN
Bone marrow (BM) transplantation was performed on a muscular mouse model of spinal muscular atrophy that had been created by mutating the survival of motor neuron gene (Smn) in myofibers only. This model is characterized by a severe myopathy and progressive loss of muscle fibers leading to paralysis. Transplantation of wild-type BM cells following irradiation at a low dose (6 Gy) improved motor capacity (+85%). This correlated with a normalization of myofiber number associated with a higher number of regenerating myofibers (1.6-fold increase) and an activation of CD34 and Pax7 satellite cells. However, BM cells had a very limited capacity to replace or fuse to mutant myofibers (2%). These data suggest that BM transplantation was able to attenuate the myopathic phenotype through an improvement of skeletal muscle regeneration of recipient mutant mice, a process likely mediated by a biological activity of BM-derived cells. This hypothesis was further supported by the capacity of muscle protein extracts from transplanted mutant mice to promote myoblast proliferation in vitro (1.6-fold increase). In addition, a tremendous upregulation of hepatocyte growth factor (HGF), which activates quiescent satellite cells, was found in skeletal muscle of transplanted mutants compared with nontransplanted mutants. Eventually, thanks to the Cre-loxP system, we show that BM-derived muscle cells were strong candidates harboring this biological activity. Taken together, our data suggest that a biological activity is likely involved in muscle regeneration improvement mediated by BM transplantation. HGF may represent an attractive paracrine mechanism to support this activity.
Asunto(s)
Trasplante de Médula Ósea/métodos , Atrofia Muscular Espinal/patología , Enfermedades Musculares/patología , Distrofia Muscular Animal/patología , Fenotipo , Animales , Antígenos CD34/inmunología , Células de la Médula Ósea/citología , Proliferación Celular , Regulación de la Expresión Génica , Proteínas Fluorescentes Verdes/metabolismo , Factor de Crecimiento de Hepatocito/genética , Ratones , Ratones Mutantes , Fibras Musculares Esqueléticas/citología , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/citología , Músculo Esquelético/patología , Músculo Esquelético/fisiología , Factor de Transcripción PAX7/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores Notch/genética , Regeneración , Células Satélite del Músculo Esquelético/citología , Células Satélite del Músculo Esquelético/patología , Factor A de Crecimiento Endotelial Vascular/genéticaRESUMEN
Mutations of the spastin gene (Sp) are responsible for the most frequent autosomal dominant form of spastic paraplegia, a disease characterized by the degeneration of corticospinal tracts. We show that a deletion in the mouse Sp gene, generating a premature stop codon, is responsible for progressive axonal degeneration, restricted to the central nervous system, leading to a late and mild motor defect. The degenerative process is characterized by focal axonal swellings, associated with abnormal accumulation of organelles and cytoskeletal components. In culture, mutant cortical neurons showed normal viability and neurite density. However, they develop neurite swellings associated with focal impairment of retrograde transport. These defects occur near the growth cone, in a region characterized by the transition between stable microtubules rich in detyrosinated alpha-tubulin and dynamic microtubules composed almost exclusively of tyrosinated alpha-tubulin. Here, we show that the Sp mutation has a major impact on neurite maintenance and transport both in vivo and in vitro. These results highlight the link between spastin and microtubule dynamics in axons, but not in other neuronal compartments. In addition, it is the first description of a human neurodegenerative disease which involves this specialized region of the axon.
Asunto(s)
Adenosina Trifosfatasas/genética , Axones/metabolismo , Microtúbulos/metabolismo , Mutación , Adenosina Trifosfatasas/fisiología , Animales , Axones/patología , Axones/ultraestructura , Secuencia de Bases , Conducta Animal , Transporte Biológico , Western Blotting , Células Cultivadas , Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/patología , Sistema Nervioso Central/ultraestructura , Corteza Cerebral/citología , Corteza Cerebral/metabolismo , Exones/genética , Eliminación de Gen , Heterocigoto , Homocigoto , Ratones , Ratones Noqueados , Microscopía Electrónica de Transmisión , Proteínas Asociadas a Microtúbulos/metabolismo , Neuronas Motoras/metabolismo , Neuronas Motoras/patología , Neuronas Motoras/ultraestructura , Neuritas/metabolismo , Neuritas/fisiología , Estructura Terciaria de Proteína , EspastinaRESUMEN
Spinal muscular atrophy (SMA) is characterized by degeneration of lower motor neurons caused by mutations of the survival motor neuron 1 gene (SMN1). SMN is involved in various processes including the formation of the spliceosome, pre-mRNA splicing and transcription. To know whether SMN has an essential role in all mammalian cell types or an as yet unknown specific function in the neuromuscular system, deletion of murine Smn exon 7, the most frequent mutation found among SMA patients, has been restricted to liver. Homozygous mutation results in severe impairment of liver development associated with iron overload and lack of regeneration leading to dramatic liver atrophy and late embryonic lethality of mutant mice. These data strongly suggest an ubiquitous and essential role of full-length SMN protein in various mammalian cell types. In SMA patients, the residual amount of SMN allows normal function of various organs except motor neurons. However, data from mouse and human suggest that other tissues might be involved in severe form of SMA or during prolonged disease course which reinforce the need of therapeutic approaches targeted to all tissues. In addition, liver function of patients should be carefully investigated and followed up before and during therapeutic trials.