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Nogo-A has been extensively studied as a myelin-associated neurite outgrowth inhibitor in the lesioned adult central nervous system. However, its role in the intact central nervous system has not yet been clarified. Analysis of the intact adult nervous system of C57BL/6 Nogo-A knock-out (KO) versus wild-type (WT) mice by a combined two-dimensional gel electrophoresis and isotope-coded affinity tagging approach revealed regulation of cytoskeleton-, transport-, and signaling growth-related proteins, pointing to regulation of the actin cytoskeleton, the neuronal growth machinery, and in particular the Rho-GTPase/LIMK1/cofilin pathway. Nogo-A KO adult neurons showed enlarged, more motile growth cones compared with WT neurons. The phenotype was reproduced by acute in vitro neutralization of neuronal Nogo-A. LIMK1 phosphorylation was increased in Nogo-A KO growth cones, and its reduction caused the decrease of KO growth cone motility to WT levels. Our study suggests that in the unlesioned adult nervous system, neuronal Nogo-A can restrict neuronal growth through negative modulation of growth cone motility.

Ullrich congenital muscular dystrophy (UCMD) is a severe disorder caused, in most cases, by a deficiency in collagen VI microfibrils. Recessive mutations in two of the three collagen VI genes, COL6A2 and COL6A3, have been identified in eight of the nine UCMD patients reported thus far. A heterozygous COL6A1 gene deletion, resulting in a mutant protein that exerts a dominant negative effect, has recently been described in a severely affected UCMD patient. Here we describe a patient in whom reverse transcription-PCR analysis of fibroblast RNA suggested a heterozygous in-frame deletion of exon 13 in the triple-helical domain of COL6A2, which is predicted to be dominantly acting. However, a homozygous A --> G mutation at -10 of intron 12 was found in the genomic DNA. The intron mutation activated numerous cryptic splice acceptor sites, generating normal and exon 13-deleted COL6A2 mRNA, and multiple aberrant transcripts containing frameshifts that were degraded through a nonsense-mediated decay mechanism. Northern analysis indicated diminished COL6A2 mRNA expression as the primary pathogenic mechanism in this UCMD patient. Our results underscore the importance of multifaceted analyses in the accurate molecular diagnosis and interpretation of genotype-phenotype correlations of UCMD.

Motor and sensory nerve conduction velocities (NCVs) and needle electromyography (EMG) results were reviewed in 26 children with different types of congenital muscular dystrophy (CMD), including patients with mutations in the genes LAMA2, FKRP, and COL6A2. In every patient, at least one EMG examination detected myopathic changes that were predominant in proximal muscles, although EMG performed at birth was normal in two patients. Brief bursts of high-frequency repetitive discharges were electrically elicited in four patients. Uniformly slowed motor NCVs without signs of denervation were observed in seven patients: five merosin-deficient, one merosin-positive, and one with unavailable merosin status. The merosin-deficient neuropathy also involved sensory nerves in three patients and worsened with age in two. In conclusion, myopathic EMG changes were typical and early findings in all types of CMD. An associated neuropathy was detected in most patients with merosin-deficient CMD, and also in a child with normal merosin expression.

Ullrich syndrome is a recessive congenital muscular dystrophy affecting connective tissue and muscle. The molecular basis is unknown. Reverse transcription-PCR amplification performed on RNA extracted from fibroblasts or muscle of three Ullrich patients followed by heteroduplex analysis displayed heteroduplexes in one of the three genes coding for collagen type VI (COL6). In patient A, we detected a homozygous insertion of a C leading to a premature termination codon in the triple-helical domain of COL6A2 mRNA. Both healthy consanguineous parents were carriers. In patient B, we found a deletion of 28 nucleotides because of an A --> G substitution at nucleotide -2 of intron 17 causing the activation of a cryptic acceptor site inside exon 18. The second mutation was an exon skipping because of a G -->> A substitution at nucleotide -1 of intron 23. Both mutations are present in an affected brother. The first mutation is also present in the healthy mother, whereas the second mutation is carried by their healthy father. In patient C, we found only one mutation so far-the same deletion of 28 nucleotides found in patient B. In this case, it was a de novo mutation, as it is absent in her parents. mRNA and protein analysis of patient B showed very low amounts of COL6A2 mRNA and of COL6. A near total absence of COL6 was demonstrated by immunofluorescence in fibroblasts and muscle. Our results demonstrate that Ullrich syndrome is caused by recessive mutations leading to a severe reduction of COL6.