Retrograde trophic signaling of nerve growth factor (NGF) supports neuronal survival and differentiation. Dysregulated trophic signaling could lead to various neurological disorders. Charcot-Marie-Tooth type 2B (CMT2B) is one of the most common inherited peripheral neuropathies characterized by severe terminal axonal loss. Genetic analysis of human CMT2B patients has revealed four missense point mutations in Rab7, a small GTPase that regulates late endosomal/lysosomal pathways, but the exact pathological mechanism remains poorly understood. Here, we show that these Rab7 mutants dysregulated axonal transport and diminished the retrograde signaling of NGF and its TrkA receptor. We found that all CMT2B Rab7 mutants were transported significantly faster than Rab7(wt) in the anterograde direction, accompanied with an increased percentile of anterograde Rab7-vesicles within axons of rat E15.5 dorsal root ganglion (DRG) neurons. In PC12M cells, the CMT2B Rab7 mutants drastically reduced the level of surface TrkA and NGF binding, presumably by premature degradation of TrkA. On the other hand, siRNA knock-down of endogenous Rab7 led to the appearance of large TrkA puncta in enlarged Rab5-early endosomes within the cytoplasm, suggesting delayed TrkA degradation. We also show that CMT2B Rab7 mutants markedly impaired NGF-induced Erk1/2 activation and differentiation in PC12M cells. Further analysis revealed that CMT2B Rab7 mutants caused axonal degeneration in rat E15.5 DRG neurons. We propose that Rab7 mutants induce premature degradation of retrograde NGF-TrkA trophic signaling, which may potentially contribute to the CMT2B disease.

Hereditary optic neuropathies comprise a group of clinically and genetically heterogeneous disorders. Two subgroups can be formed: isolated hereditary optic atrophies and optic neuropathy as part of complex disorders. In group 1 of hereditary optic neuropathies, optic nerve dysfunction is typically the only manifestation of the disease. This group comprises autosomal dominant, autosomal recessive and X-linked recessive optic atrophy and the maternally inherited Leber's hereditary optic neuropathy. Among the autosomal-dominant forms of optic atrophy, Kjer's disease is most frequently observed. In the second group of complex disorders, various neurologic and other systemic abnormalities are regularly observed. Most frequent in this group are mtDNA mutations, inherited peripheral neuropathies, Charcot-Marie-Tooth disorders (CMT2A2, CMTX5), hereditary sensory neuropathy type 3 (HSAN3), Friedreich's ataxia, leukodystrophies, sphingolipidoses, ceroid-lipofuscinoses and neurodegeneration with brain iron accumulation. We review current knowledge about the underlying genetic predispositions, the most urgent open questions and how this may affect our management of this heterogeneous group of disorders in the future.

Charcot-Marie-Tooth (CMT) disease represents a clinically and genetically heterogeneous group of inherited neuropathies. Here, we report a five-generation family of eight affected individuals with CMT disease type 2, CMT2. Genome-wide linkage analysis showed that the disease phenotype is closely linked to chromosomal region 10p13-14, which spans 5.41 Mb between D10S585 and D10S1477. DNA-sequencing analysis revealed a nonsense mutation, c.1455T>G (p.Tyr485(∗)), in exon 8 of dehydrogenase E1 and transketolase domain-containing 1 (DHTKD1) in all eight affected individuals, but not in other unaffected individuals in this family or in 250 unrelated normal persons. DHTKD1 mRNA expression levels in peripheral blood of affected persons were observed to be half of those in unaffected individuals. In vitro studies have shown that, compared to wild-type mRNA and DHTKD1, mutant mRNA and truncated DHTKD1 are significantly decreased by rapid mRNA decay in transfected cells. Inhibition of nonsense-mediated mRNA decay by UPF1 silencing effectively rescued the decreased levels of mutant mRNA and protein. More importantly, DHTKD1 silencing was found to lead to impaired energy production, evidenced by decreased ATP, total NAD(+) and NADH, and NADH levels. In conclusion, our data demonstrate that the heterozygous nonsense mutation in DHTKD1 is one of CMT2-causative genetic alterations, implicating an important role for DHTKD1 in mitochondrial energy production and neurological development.

Recent advances in genetic analysis technology have enabled a surprising progress in genetic diagnosis in the field of neurological disease research. High-throughput molecular biology techniques, such as microarrays and next-generation sequencing, are the major contributors to this progress and to new discoveries. Charcot-Marie-Tooth disease (CMT), a known hereditary motor and sensory neuropathy, is clinically and genetically heterogeneous. Genetic studies have revealed at least 35 disease causing-genes responsible for Charcot-Marie-Tooth disease. Genetic studies have revealed that abnormalities in the following factors are the cause of inherited neuropathies: myelin components, transcription factors controlling myelination, myelin maintenance system, differentiation factors related to the peripheral nerve, neurofilaments, protein transfer system, mitochondrial proteins, DNA repair, RNA/protein synthesis, ion channels, and aminoacyl-tRNA synthetase. On the other hand concomitant with the increase in the number of genes that must be screened for mutations, the labor and reagent costs for molecular genetic testing have increased significantly. Therefore, new methodology for detecting gene mutations is required. Based on the recent progress in DNA analysis methods, resequencing microarray appears to be an economical and highly sensitive method for detecting mutations. We have been screening CMT patients for mutations using originally designed microarray DNA chips since 2007, thencehaving identified disease causing mutations in MPZ, GJB1, PMP22, EGR2, MFN2, NEFL, PRX, AARS, GARS, DNM2, and SETX genes in CMT patients.

Charcot-Marie-Tooth disease is the most common inherited disorder of the peripheral nervous system. The disease is characterized by a progressive muscle weakness and atrophy, sensory loss, foot (and hand) deformities and steppage gait. While many of the genes associated with axonal CMT have been identified, to date it is unknown which mechanism(s) causes the disease. However, genetic findings indicate that the underlying mechanisms mainly converge to the axonal cytoskeleton. In this review, we will summarize the evidence for this pathogenic convergence. Furthermore, recent work with new transgenic mouse models has led to the identification of histone deacetylase 6 as a potential therapeutic target for inherited peripheral neuropathies. This enzyme deacetylates microtubules and plays a crucial role in the regulation of axonal transport. These findings offer new perspectives for a potential therapy to treat axonal Charcot-Marie-Tooth disease and other neurodegenerative disorders characterized by axonal transport defects.

Charcot-Marie-Tooth disease type 2D (CMT2D) is a dominantly inherited peripheral neuropathy caused by missense mutations in the glycyl-tRNA synthetase gene (GARS). In addition to GARS, mutations in three other tRNA synthetase genes cause similar neuropathies, although the underlying mechanisms are not fully understood. To address this, we generated transgenic mice that ubiquitously over-express wild-type GARS and crossed them to two dominant mouse models of CMT2D to distinguish loss-of-function and gain-of-function mechanisms. Over-expression of wild-type GARS does not improve the neuropathy phenotype in heterozygous Gars mutant mice, as determined by histological, functional, and behavioral tests. Transgenic GARS is able to rescue a pathological point mutation as a homozygote or in complementation tests with a Gars null allele, demonstrating the functionality of the transgene and revealing a recessive loss-of-function component of the point mutation. Missense mutations as transgene-rescued homozygotes or compound heterozygotes have a more severe neuropathy than heterozygotes, indicating that increased dosage of the disease-causing alleles results in a more severe neurological phenotype, even in the presence of a wild-type transgene. We conclude that, although missense mutations of Gars may cause some loss of function, the dominant neuropathy phenotype observed in mice is caused by a dose-dependent gain of function that is not mitigated by over-expression of functional wild-type protein.

Charcot-Marie-Tooth disease (CMT) and related peripheral neuropathies are the most commonly inherited neurological disorders in humans, characterized by clinical and genetic heterogeneity. The most prevalent clinical entities belonging to this group of disorders are CMT type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP). CMT1A and HNPP are predominantly caused by a 1.5 Mb duplication and deletion in the chromosomal region 17p11.2, respectively, and less frequently by other mutations in the peripheral myelin protein 22 (PMP22) gene. Despite being relatively common diseases, they haven't been previously studied in the Slovak population. Therefore, the aim of this study was to identify the spectrum and frequency of PMP22 mutations in the Slovak population by screening 119 families with CMT and 2 families with HNPP for causative mutations in this gene. The copy number determination of PMP22 resulted in the detection of CMT1A duplication in 40 families and the detection of HNPP deletion in 7 families, 6 of which were originally diagnosed as CMT. Consequent mutation screening of families without duplication or deletion using dHPLC and sequencing identified 6 single base changes (3 unpublished to date), from which only c.327C>A (Cys109X) present in one family was provably causative. These results confirm the leading role of PMP22 mutation analysis in the differential diagnosis of CMT and show that the spectrum and frequency of PMP22 mutations in the Slovak population is comparable to that seen in the global population.

Previous studies in our laboratory have shown that in models for three distinct forms of the inherited and incurable nerve disorder, Charcot-Marie-Tooth neuropathy, low-grade inflammation implicating phagocytosing macrophages mediates demyelination and perturbation of axons. In the present study, we focus on colony-stimulating factor-1, a cytokine implicated in macrophage differentiation, activation and proliferation and fostering neural damage in a model for Charcot-Marie-Tooth neuropathy 1B. By crossbreeding a model for the X-linked form of Charcot-Marie-Tooth neuropathy with osteopetrotic mice, a spontaneous null mutant for colony-stimulating factor-1, we demonstrate a robust and persistent amelioration of demyelination and axon perturbation. Furthermore, functionally important domains of the peripheral nervous system, such as juxtaparanodes and presynaptic terminals, were preserved in the absence of colony-stimulating factor-1-dependent macrophage activation. As opposed to other Schwann cell-derived cytokines, colony-stimulating factor-1 is expressed by endoneurial fibroblasts, as revealed by in situ hybridization, immunocytochemistry and detection of β-galactosidase expression driven by the colony-stimulating factor-1 promoter. By both light and electron microscopic studies, we detected extended cell-cell contacts between the colony-stimulating factor-1-expressing fibroblasts and endoneurial macrophages as a putative prerequisite for the effective and constant activation of macrophages by fibroblasts in the chronically diseased nerve. Interestingly, in human biopsies from patients with Charcot-Marie-Tooth type 1, we also found frequent cell-cell contacts between macrophages and endoneurial fibroblasts and identified the latter as main source for colony-stimulating factor-1. Therefore, our study provides strong evidence for a similarly pathogenic role of colony-stimulating factor-1 in genetically mediated demyelination in mice and Charcot-Marie-Tooth type 1 disease in humans. Thus, colony-stimulating factor-1 or its cognate receptor are promising target molecules for treating the detrimental, low-grade inflammation of several inherited neuropathies in humans.

Inherited neuropathies are clinically and genetically heterogeneous. At least 30 genes have been associated with Charcot-Marie-Tooth disease (CMT) and related inherited neuropathies. Genetic studies have revealed that abnormalities in the following factors are the cause of inherited neuropathies: myelin components, transcription factors controlling myelination, myelin maintenance system, differentiation factors related to the peripheral nerve, neurofilaments, protein transfer system, mitochondrial proteins, DNA repair, RNA/protein synthesis, ion channels, and aminoacyl-tRNA synthetase. On the other hand, a precise molecular diagnosis is often needed to confirm a clinical diagnosis, offer genetic counseling to the patient and family, and provide prognostic information to the patient. Unfortunately, along with the increase in the number of genes that must be screened for mutations, the labor and reagent costs of molecular genetic testing have increased significantly. On the basis of the recent progress of DNA analysis methods, the use of resequencing microarray seems to be an economical and highly sensitive method to detect mutations. In this study, we attempted to screen for CMT patients mutations using these methods.

Mutations in genes expressed in Schwann cells and the axons they ensheathe cause the hereditary motor and sensory neuropathies, also known as Charcot-Marie-Tooth disease (CMT). More than 40 different genes have been shown to cause inherited neuropathies; chromosomal localizations of many other distinct inherited neuropathies have been mapped, and new genetic causes for inherited neuropathies continue to be discovered. How to keep track of all of these disorders, when to pursue genetic testing, and what tests to order for specific patients are difficult challenges for any neurologist. This review addresses these issues and provides illustrative cases to help in dealing with them. CMT serves as a living system to identify molecules necessary for normal peripheral nervous system (PNS) function. Understanding how these various molecules interact will provide a better understanding of the pathogenesis of peripheral neuropathies in general as well as other neurodegenerative disorders involving the PNS.