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Charcot-Marie-Tooth type 1B disease is a demyelinating neuropathy caused by mutations in the Myelin Protein Zero gene. It is inherited in an autosomal dominant fashion. So far only a few patients with a focally folded myelin phenotype on nerve biopsy have been shown to have mutations in the Myelin Protein Zero gene. In this report we describe a Polish patient with Charcot-Marie-Tooth type 1B disease. Sural nerve biopsy demonstrated focally folded myelin. Molecular genetic analysis of the coding region of the Myelin Protein Zero gene revealed a novel mutation, Thr65Ala, in exon 2 of the Myelin Protein Zero gene. [Copyright &y& Elsevier]

Charcot−Marie−Tooth disorders (CMT) represent a highly heterogeneous group of diseases of the peripheral nervous system in which more than 100 genes are involved. In some CMT patients, a few weak sequence variants toward other CMT genes are detected instead of one leading CMT mutation. Thus, the presence of a few variants in different CMT-associated genes raises the question concerning the pathogenic status of one of them. In this study, we aimed to analyze the pathogenic effect of c.664G>A, p.Glu222Lys variant in the GDAP1 gene, whose mutations are known to be causative for CMT type 4A (CMT4A). Due to low penetrance and a rare occurrence limited to five patients from two Polish families affected by the CMT phenotype, there is doubt as to whether we are dealing with real pathogenic mutation. Thus, we aimed to study the pathogenic effect of the c.664G>A, p.Glu222Lys variant in its natural environment, i.e., the neuronal SH-SY5Y cell line. Additionally, we have checked the pathogenic status of p.Glu222Lys in the broader context of the whole exome. We also have analyzed the impact of GDAP1 gene mutations on the morphology of the transfected cells. Despite the use of several tests to determine the pathogenicity of the p.Glu222Lys variant, we cannot point to one that would definitively solve the problem of pathogenicity.

Mutations in HNRNPA1 encoding heterogeneous nuclear ribonucleoprotein (hnRNP) A1 are a rare cause of amyotrophic lateral sclerosis (ALS) and multisystem proteinopathy (MSP). hnRNPA1 is part of the group of RNA-binding proteins (RBPs) that assemble with RNA to form RNPs. hnRNPs are concentrated in the nucleus and function in pre-mRNA splicing, mRNA stability, and the regulation of transcription and translation. During stress, hnRNPs, mRNA, and other RBPs condense in the cytoplasm to form stress granules (SGs). SGs are implicated in the pathogenesis of (neuro-)degenerative diseases, including ALS and inclusion body myopathy (IBM). Mutations in RBPs that affect SG biology, including FUS, TDP-43, hnRNPA1, hnRNPA2B1, and TIA1, underlie ALS, IBM, and other neurodegenerative diseases. Here, we characterize 4 potentially novel HNRNPA1 mutations (yielding 3 protein variants: *321Eext*6, *321Qext*6, and G304Nfs*3) and 2 known HNRNPA1 mutations (P288A and D262V), previously connected to ALS and MSP, in a broad spectrum of patients with hereditary motor neuropathy, ALS, and myopathy. We establish that the mutations can have different effects on hnRNPA1 fibrillization, liquid-liquid phase separation, and SG dynamics. P288A accelerated fibrillization and decelerated SG disassembly, whereas *321Eext*6 had no effect on fibrillization but decelerated SG disassembly. By contrast, G304Nfs*3 decelerated fibrillization and impaired liquid phase separation. Our findings suggest different underlying pathomechanisms for HNRNPA1 mutations with a possible link to clinical phenotypes.

Phylogenetic analyses based on mitochondrial 16S rDNA, nuclear 28S rDNA, and morphological and ecological traits of Aulactinia, Urticina and Cribrinopsis sea anemones inhabiting the Arctic-boreal region indicate discordances between trees derived from molecular sequences and those based on morphological traits. Nuclear genes were more informative than mitochondrial and morphological datasets. Our findings indicate that 16S rDNA has limited applicability for phylogenetic analyses at lower taxonomic levels and can only be used for distinction of families. Although 28S rDNA allowed for the classification of distinct genera, it could not confirm that species of Urticina and Cribrinopsis , which appeared to be closely related, were correctly separated into two different genera. The nuclear tree revealed inconsistencies between specimens belonging to European Urticina crassicornis and Pacific U. crassicornis ; the latter seems to be a different species. In contrast to Pacific U. crassicornis , the specimens collected from different localities in the Barents Sea are on the same tree branch. The same was observed for specimens of Aulactinia stella . Both species brood their young internally. The dispersal of sea anemones with brooding juveniles seems to be less limited than expected and might be sufficient to settle habitats more than a thousand kilometers away.

Charcot-Marie-Tooth disease (CMT) is a heritable neurodegenerative disease that displays great genetic heterogeneity. The genes and mutations that underlie this heterogeneity have been extensively characterized by molecular genetics. However, the molecular pathogenesis of the vast majority of CMT subtypes remains terra incognita. Any attempts to perform experimental therapy for CMT disease are limited by a lack of understanding of the pathogenesis at a molecular level. In this study, we aim to identify the molecular pathways that are disturbed by mutations in the gene encoding GDAP1 using both yeast and human cell, based models of CMT-GDAP1 disease. We found that some mutations in GDAP1 led to a reduced expression of the GDAP1 protein and resulted in a selective disruption of the Golgi apparatus. These structural alterations are accompanied by functional disturbances within the Golgi. We screened over 1500 drugs that are available on the market using our yeast-based CMT-GDAP1 model. Drugs were identified that had both positive and negative effects on cell phenotypes. To the best of our knowledge, this study is the first report of the Golgi apparatus playing a role in the pathology of CMT disorders. The drugs we identified, using our yeast-based CMT-GDAP1 model, may be further used in translational research.

Objective: To test the hypothesis that monogenic neuropathies such as Charcot-Marie-Tooth disease (CMT) contribute to frequent but often unexplained neuropathies in the elderly, we performed genetic analysis of 230 patients with unexplained axonal neuropathies and disease onset ≥35 years., Methods: We recruited patients, collected clinical data, and conducted whole-exome sequencing (WES; n = 126) and MME single-gene sequencing (n = 104). We further queried WES repositories for MME variants and measured blood levels of the MME -encoded protein neprilysin., Results: In the WES cohort, the overall detection rate for assumed disease-causing variants in genes for CMT or other conditions associated with neuropathies was 18.3% (familial cases 26.4%, apparently sporadic cases 12.3%). MME was most frequently involved and accounted for 34.8% of genetically solved cases. The relevance of MME for late-onset neuropathies was further supported by detection of a comparable proportion of cases in an independent patient sample, preponderance of MME variants among patients compared to population frequencies, retrieval of additional late-onset neuropathy patients with MME variants from WES repositories, and low neprilysin levels in patients' blood samples. Transmission of MME variants was often consistent with an incompletely penetrant autosomal-dominant trait and less frequently with autosomal-recessive inheritance., Conclusions: A detectable fraction of unexplained late-onset axonal neuropathies is genetically determined, by variants in either CMT genes or genes involved in other conditions that affect the peripheral nerves and can mimic a CMT phenotype. MME variants can act as completely penetrant recessive alleles but also confer dominantly inherited susceptibility to axonal neuropathies in an aging population., (© 2020 American Academy of Neurology.)

Charcot-Marie-Tooth (CMT) disease encompasses a group of rare disorders that are characterized by similar clinical manifestations and a high genetic heterogeneity. Such excessive diversity presents many problems. Firstly, it makes a proper genetic diagnosis much more difficult and, even when using the most advanced tools, does not guarantee that the cause of the disease will be revealed. Secondly, the molecular mechanisms underlying the observed symptoms are extremely diverse and are probably different for most of the disease subtypes. Finally, there is no possibility of finding one efficient cure for all, or even the majority of CMT diseases. Every subtype of CMT needs an individual approach backed up by its own research field. Thus, it is little surprise that our knowledge of CMT disease as a whole is selective and therapeutic approaches are limited. There is an urgent need to develop new CMT models to fill the gaps. In this review, we discuss the advantages and disadvantages of yeast as a model system in which to study CMT diseases. We show how this single-cell organism may be used to discriminate between pathogenic variants, to uncover the mechanism of pathogenesis, and to discover new therapies for CMT disease.

The question of whether a newly identified sequence variant is truly a causative mutation is a central problem of modern clinical genetics. In the current era of massive sequencing, there is an urgent need to develop new tools for assessing the pathogenic effect of new sequence variants. In Charcot-Marie-Tooth disorders (CMT) with their extreme genetic heterogeneity and relatively homogenous clinical presentation, addressing the pathogenic effect of rare sequence variants within 80 CMT genes is extremely challenging. The presence of multiple rare sequence variants within a single CMT-affected patient makes selection for the strongest one, the truly causative mutation, a challenging issue. In the present study we propose a new yeast-based model to evaluate the pathogenic effect of rare sequence variants found within the one of the CMT-associated genes, GDAP1 . In our approach, the wild-type and pathogenic variants of human GDAP1 gene were expressed in yeast. Then, a growth rate and mitochondrial morphology and function of GDAP1 -expressing strains were studied. Also, the mutant GDAP1 proteins localization and functionality were assessed in yeast. We have shown, that GDAP1 was not only stably expressed but also functional in yeast cell, as it influenced morphology and function of mitochondria and altered the growth of a mutant yeast strain. What is more, the various GDAP1 pathogenic sequence variants caused the specific for them effect in the tests we performed. Thus, the proposed model is suitable for validating the pathogenic effect of known GDAP1 mutations and may be used for testing of unknown sequence variants found in CMT patients.

Hereditary motor and sensory neuropathies (HMSN) also called as Charcot-Marie-Tooth disorders (CMT) are extremely heterogeneous group of disorders of peripheral nervous system. Over 80 genes have been reported in different types of CMT. In all CMT affected patients the main symptoms are slowly progressive wasting of the distal muscles of the lower and upper limbs. To date no efficient therapeutic approach basing upon molecular pathology of CMT has been proposed. This review presents the current state of knowledge concerning clinical, molecular pathogenesis and experimental therapy aspects in CMT disorders. Additionally the possibilities resulting from the use of the yeast model to the identification of new therapeutic substances as well as of neurotoxins are also discussed.

To date, only two splice-site mutations within the TPM2 gene have been shown to be causative for congenital myopathies. While the majority of TPM2 gene mutations are causative for nemaline myopathy, cap disease or distal arthrogryposis, some mutations in this gene have been found to be associated with non-specific congenital myopathy. We report on a patient with such an unspecified congenital myopathy associated with distinctive facial dysmorphic features and distal arthrogryposis. Using the whole exome sequencing (WES) approach we were able to identify a novel heterozygous splice-site mutation within the TPM2 gene, showing the utility of WES in molecular diagnostics of congenital myopathies without recognizable morphological hallmarks.

Charcot-Marie-Tooth disease type 2A (CMT2A) is an autosomal dominant axonal peripheral neuropathy caused by mutations in the mitofusin 2 gene (MFN2). Mitofusin 2 is a GTPase protein present in the outer mitochondrial membrane and responsible for regulation of mitochondrial network architecture via the fusion of mitochondria. As that fusion process is known to be strongly dependent on the GTPase activity of mitofusin 2, it is postulated that the MFN2 mutation within the GTPase domain may lead to impaired GTPase activity, and in turn to mitochondrial dysfunction. The work described here has therefore sought to verify the effects of MFN2 mutation within its GTPase domain on mitochondrial and endoplasmic reticulum morphology, as well as the mtDNA content in a cultured primary fibroblast obtained from a CMT2A patient harboring a de novo Arg274Trp mutation. In fact, all the parameters studied were affected significantly by the presence of the mutant MFN2 protein. However, using the stable model for mitofusin 2 obtained by us, we were next able to determine that the Arg274Trp mutation does not impact directly upon GTP binding. Such results were also confirmed for GTP-hydrolysis activity of MFN2 protein in patient fibroblast. We therefore suggest that the biological malfunctions observable with the disease are not consequences of impaired GTPase activity, but rather reflect an impaired contribution of the GTPase domain to other MFN2 activities involving that region, for example protein-protein interactions., Competing Interests: The authors have declared that no competing interests exist.

The Warburg micro syndrome (WARBM) is a genetically heterogeneous syndrome linked to at least 4 loci. At the clinical level, WARBM is characterized by microcephaly, microphthalmia, microcornea, congenital cataracts, corpus callosum hypoplasia, severe mental retardation, and hypogonadism. In some families additional clinical features have been reported. The presence of uncommon clinical features (peripheral neuropathy, cardiomyopathy) may result in misdirected molecular diagnostics. Using the next generation sequencing approach (NGS), we were able to diagnose WARBM1 syndrome by detection of a new mutation within the RAB3GAP1 gene. We have detected some DNA variants which may be responsible for cardiomyopathy. We did not find any obvious pathogenic mutation within a set of genes known to be responsible for hereditary motor and sensory neuropathy (HMSN). We conclude that: (i) in clinically delineated syndromes, a classical single-gene oriented approach may be not conclusive especially in the presence of rare clinical features, (ii) peripheral neuropathy and cardiomyopathy are rare additional symptoms coexisting with WARBM1, (iii) a pleiotropic effect of a single point mutation is sufficient to be causative for WARBM1 and (iv) more WARBM-affected patients should be reported to delineate a complete phenotype.

Background: Limb girdle muscular dystrophy 2A (LGMD2A) is the most frequent LGMD variant in the European population, representing about 40% of LGMD. The c.550delA mutation in the CANP3 (calcium activated neutral protease 3) gene is the most commonly reported mutation in LGMD2A. Prevalence of this mutation in the Polish population has not been previously investigated. The aim of this study was to identify and estimate the frequency of the c.550delA mutation in Polish LGMD2A patients., Methods: Polymerase chain reaction-sequencing analysis, restriction fragment length polymorphism polymerase chain reaction method., Results: We analyzed 76 families affected with LGMD and identified 62 probands with mutations in the CANP3 gene. C.550delA was the most common mutation identified, being found in 78% of the LGMD2A families. The remaining mutations observed multiple times were as follows: c.598-612del15ntd; c.2242C>T; c.418dupC; c.1356insT, listed in terms of decreasing frequency. Two novel variants in the CANP3 gene, that is, c.700G>A Gly234Arg and c.661G>A Gly221Ser were also characterized. Overall, mutations in the LGMD2A gene were estimated to be present in 81% of patients with the LGMD phenotype who were without sarcoglycans and dysferlin deficiency on immunocytochemical analysis. The frequency of the heterozygous c.550delA mutation in the healthy Polish population was estimated at 1/124., Conclusions: The c.550delA is the most frequent CANP3 mutation in the Polish population, thus sequencing of exon 4 of this gene could identify the majority of LGMD2A patients in Poland.

Facioscapulohumeral muscular dystrophy cases with facial weakness before the age of 5 and signs of shoulder weakness by the age of 10 are defined as early onset. Contraction of the D4Z4 repeat on chromosome 4q35 is causally related to facioscapulohumeral muscular dystrophy type 1, and the residual size of the D4Z4 repeat shows a roughly inverse correlation with the severity of the disease. Contraction of the D4Z4 repeat on chromosome 4q35 is believed to induce a local change in chromatin structure and consequent transcriptional deregulation of 4qter genes. We present early-onset cases in the Polish population that amounted to 21% of our total population with facioscapulohumeral muscular dystrophy. More than 27% of them presented with severe phenotypes (wheelchair dependency). The residual D4Z4 repeat sizes ranged from 1 to 4 units. In addition, even within early-onset facioscapulohumeral muscular dystrophy type 1 phenotypes, some cases had uncommon features (head drop, early disabling contractures, progressive ptosis, and respiratory insufficiency and cardiomyopathy)., (© The Author(s) 2014.)

Hereditary sensory and autonomic neuropathies (HSANs) represent a group of heritable peripheral nerve disorders usually taking a severe clinical course. HSAN-affected patients manifest with deep, poorly-healing ulcerations of the feet and hands. To date no definitive cure for HSANs has been developed and the molecular pathology of these disorders is complex. The aim of this review is therefore to present recent findings in terms of HSAN molecular pathogenesis. So far, mutations in 12 genes coding for different proteins have been reported in association with HSAN and the molecular pathogenesis has been elucidated in HSAN1a, HSAN4 and HSAN5. The genes involved in molecular pathogenesis of HSAN code for a wide spectrum of proteins from enzymes to specific nerve growth factors. As far as HSAN1a is concerned, the enhanced understanding has given rise to achievements in experimental therapy particularly in respect to disease models. Despite a rapid progress in studies on the molecular background of HSAN, numerous loci and genes remain still to be discovered.

At the time of its first description in 2004, MFN2 was considered the most frequently mutated gene in hereditary motor and sensory neuropathy type 2 (HMSN 2). However recent studies have shown that the frequency of MFN2 gene mutations in HMSN II patients is surprisingly low. To date, no systematic studies devoted to HMSN IIa in Poland have been carried out. In this study, we searched for MFN2 gene mutations in Polish patients representing the population of nearly 40 million. We decided to include a wide spectrum of clinical phenotypes in the study, proving able to detect, in a group of 67 affected patients: 1) 3 pathogenic mutations; 2) 3 sequence variants of unknown pathogenic status; 3) 9 rare MFN2 gene sequence variants; 4) 6 common polymorphisms. The frequency of MFN2 gene mutations in the whole group of patients is 4.5%. Due to the high frequency of MFN2 gene sequence variants within single patients we could not definitely exclude the cumulative effect of these contributing to the HMSN II phenotype. The MFN2 gene should therefore be considered in Polish HMSN II patients, though it is still not possible to determine its position in HMSN II molecular diagnostics.

Charcot-Marie-Tooth disease type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP) represent the most common heritable neuromuscular disorders. Molecular diagnostics of CMT1A/HNPP diseases confirm clinical diagnosis, but their value is limited to the clinical course and prognosis. However, no biomarkers of CMT1A/HNPP have been identified. We decided to explore if the LITAF/SIMPLE gene shared a functional link to the PMP22 gene, whose duplication or deletion results in CMT1A and HNPP, respectively. By studying a large cohort of CMT1A/HNPP-affected patients, we found that the LITAF I92V sequence variant predisposes patients to an earlier age of onset of both the CMT1A and HNPP diseases. Using cell transfection experiments, we showed that the LITAF I92V sequence variant partially mislocalizes to the mitochondria in contrast to wild-type LITAF which localizes to the late endosome/lysosomes and is associated with a tendency for PMP22 to accumulate in the cells. Overall, this study shows that the I92V LITAF sequence variant would be a good candidate for a biomarker in the case of the CMT1A/HNPP disorders.

Introduction: The first episode of hereditary neuropathy with liability to pressure palsy (HNPP) in childhood is rare., Methods: We analyzed retrospectively the data of 7 patients with a deletion in PMP22 and onset of symptoms before age 18 years. Direct sequencing of the LITAF (lipopolysaccharide-induced tumor necrosis factor) gene was performed in patients and family members., Results: Clinical presentations varied from mononeuropathies to brachial plexopathy, with recurrent episodes in 4 patients. Electrophysiological abnormalities characteristic for HNNP were found in most subjects. Analysis of the LITAF gene revealed an Ile92Val polymorphism in 6 of 7 (86%) probands and 5 of 7 (83%) family members, over 4 times greater frequency than in the general population., Conclusions: Clinical suspicion of HNPP even when nerve conduction study results do not fulfill HNPP criteria should indicate genetic testing. In our patients, early-onset HNPP was associated frequently with isoleucine92valine LITAF polymorphism., (© 2014 Wiley Periodicals, Inc.)

Using a combination of exome sequencing and linkage analysis, we investigated an English family with two affected siblings in their 40s with recessive Charcot-Marie Tooth disease type 2 (CMT2). Compound heterozygous mutations in the immunoglobulin-helicase-μ-binding protein 2 (IGHMBP2) gene were identified. Further sequencing revealed a total of 11 CMT2 families with recessively inherited IGHMBP2 gene mutations. IGHMBP2 mutations usually lead to spinal muscular atrophy with respiratory distress type 1 (SMARD1), where most infants die before 1 year of age. The individuals with CMT2 described here, have slowly progressive weakness, wasting and sensory loss, with an axonal neuropathy typical of CMT2, but no significant respiratory compromise. Segregating IGHMBP2 mutations in CMT2 were mainly loss-of-function nonsense in the 5' region of the gene in combination with a truncating frameshift, missense, or homozygous frameshift mutations in the last exon. Mutations in CMT2 were predicted to be less aggressive as compared to those in SMARD1, and fibroblast and lymphoblast studies indicate that the IGHMBP2 protein levels are significantly higher in CMT2 than SMARD1, but lower than controls, suggesting that the clinical phenotype differences are related to the IGHMBP2 protein levels., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Charcot-Marie-Tooth (CMT) disease caused by mutations in the GDAP1 gene has been shown to be inherited via traits that may be either autosomal recessive (in the majority of cases) [CMT4A] or autosomal dominant [CMT2K]. CMT4A disease is characterized by an early onset, and a severe clinical course often leading to a loss of ambulation, whereas CMT2K is characterized by a mild clinical course of benign axonal neuropathy beginning even in the 6th decade of life. Clinical data from a GDAP1 mutated patient suggests that the presence of a particular mutation is associated with a certain trait of inheritance. The association of a particular GDAP1 gene mutation and a dominant or recessive trait of inheritance is of special importance for genetic counseling and the prenatal diagnostics as regards severe forms of CMT. In the present study we report on two CMT families in which a newly identified Glu222Lys mutation within the GDAP1 gene segregates both in autosomal dominant and recessive traits. Our study shows that at least some GDAP1 gene mutations may segregate with the CMT phenotype as both dominant and recessive traits. Thus, genetic counseling for CMT4A/CMT2K families requires more extensive data on GDAP1 phenotype-genotype correlations.

Mitofusin 2 (Mfn2), a protein of the mitochondrial outer membrane, is essential for mitochondrial fusion and contributes to the maintenance and operation of the mitochondrial network. Mutations in the mitofusin 2 gene cause axonal Charcot-Marie-Tooth type 2A (CMT2A), an inherited disease affecting peripheral nerve axons. The precise mechanism by which mutations in MFN2 selectively cause the degeneration of long peripheral axons is not known. There is a hypothesis suggesting the involvement of reduced expression of a homologous protein, mitofusin 1 (Mfn1), in the peripheral nervous system, and less effective compensation of defective mitofusin 2 by mitofusin 1. We therefore aimed to perform an analysis of the mitofusin 1 and mitofusin 2 mRNA and protein expression profiles in different mouse tissues, with special attention paid to dorsal root ganglia (DRGs), as parts of the peripheral nervous system. Quantitative measurement relating to mRNA revealed that expression of the Mfn2 gene dominates over Mfn1 mainly in mouse DRG, as opposed to other nervous system samples and other tissues studied. This result was further supported by Western blot evaluation. Both these sets of data confirm the hypothesis that the cellular consequences of mutations in the mitofusin 2 gene can mostly be manifested in the peripheral nervous system.

Charcot-Marie-Tooth type 2A disease (CMT2A) caused by mutations in the Mitofusin 2 gene (Mfn2) has been shown to be an early-onset axonal neuropathy with severe clinical course in the majority of the patients. In this study we present a unique phenotype of CMT2A disease characterized by late-onset polyneuropathy with a very mild clinical course. This rare form of CMT2A disease is caused by a new splice-site (c.311+1G>T) mutation within the MFN2 gene. Due to disturbance of the MFN2 splicing process, this mutation generates a short transcript which encodes a very short fragment of MFN2 protein. The c.311+1G>T mutation within the MFN2 gene results in the late -onset CMT2 disease.

Hereditary sensory and autonomic neuropathy type 2 is a rare disorder caused by recessive mutations in the WNK1/HSN2 gene located on chromosome 12p13.33. Phenotype of the patients is characterized by severe sensory loss affecting all sensory modalities. We report a novel homozygous Lys179fsX182 (HSN2); Lys965fsX968 (WNK1/HSN2) mutation causing an early childhood onset hereditary sensory and autonomic neuropathy type 2, with acromutilations in upper and lower limbs, and autonomic dysfunction. To the best of our knowledge this is the first genetically proven case of hereditary sensory and autonomic neuropathy type 2 originating from East Europe.

Charcot-Marie-Tooth type 1C disease (CMT1C) is a rare form of hereditary demyelinating neuropathy caused by mutations in the LITAF (lipopolysaccharide-induced tumor necrosis factor-) gene. CMT1C disease was mapped to chromosome 16p12-p 13.3. To date only a few mutations in the LITAF gene have been reported. Due to a small group of CMT1C reported patients, the phenotype of CMT1C is poorly characterized. CMT1C disease is a pure demyelinating neuropathy limited to the peripheral nervous system with a mild clinical course, manifesting without any additional symptoms. To the best of our knowledge, in this study, for the first time we present a three generational CMT1C family in which in the proband, CMT1C disease coexists with central demyelination fulfilling criteria of primary progressive multiple sclerosis (PPMS). The coexistence of PPMS and CMT1C in one family may not result from a common pathogenetic trait, however only in the proband with central demyelination and CMT1C we have detected a -308G>A sequence variant in the promoter of the TNF-α gene.

Early onset hereditary motor and sensory neuropathies are rare disorders encompassing congenital hypomyelinating neuropathy with disease onset in the direct post-natal period and Dejerine-Sottas neuropathy starting in infancy. The clinical spectrum, however, reaches beyond the boundaries of these two historically defined disease entities. De novo dominant mutations in PMP22, MPZ and EGR2 are known to be a typical cause of very early onset hereditary neuropathies. In addition, mutations in several other dominant and recessive genes for Charcot-Marie-Tooth disease may lead to similar phenotypes. To estimate mutation frequencies and to gain detailed insights into the genetic and phenotypic heterogeneity of early onset hereditary neuropathies, we selected a heterogeneous cohort of 77 unrelated patients who presented with symptoms of peripheral neuropathy within the first year of life. The majority of these patients were isolated in their family. We performed systematic mutation screening by means of direct sequencing of the coding regions of 11 genes: MFN2, PMP22, MPZ, EGR2, GDAP1, NEFL, FGD4, MTMR2, PRX, SBF2 and SH3TC2. In addition, screening for the Charcot-Marie-Tooth type 1A duplication on chromosome 17p11.2-12 was performed. In 35 patients (45%), mutations were identified. Mutations in MPZ, PMP22 and EGR2 were found most frequently in patients presenting with early hypotonia and breathing difficulties. The recessive genes FGD4, PRX, MTMR2, SBF2, SH3TC2 and GDAP1 were mutated in patients presenting with early foot deformities and variable delay in motor milestones after an uneventful neonatal period. Several patients displaying congenital foot deformities but an otherwise normal early development carried the Charcot-Marie-Tooth type 1A duplication. This study clearly illustrates the genetic heterogeneity underlying hereditary neuropathies with infantile onset.

Charcot-Marie-Tooth disease (CMT) caused by mutations in the ganglioside-induced differentiation-associated protein 1 (GDAP1) gene is characterized by a spectrum of phenotypes. Recurrent nonsense mutations (Q163X and S194X) showing regional distribution segregate with an early onset, severe course of recessive CMT disease with early loss of ambulancy. Missense mutations in GDAP1 have been reported in sporadic CMT cases with variable course of disease, among them the recurrent L239F missense GDAP1 mutation occurring in the European population. Finally, some GDAP1 mutations are associated with a mild form of CMT inherited as an autosomal dominant trait. In this study, we characterize the CMT phenotype in one Polish family with recessive trait of inheritance at the clinical, electrophysiological, morphological, cellular, and genetic level associated with a new Gly327Asp mutation in the GDAP1 gene. In spite of the nature of Gly327Asp mutation (missense), the CMT phenotype associated with this variant may be characterized as an early onset, severe axonal neuropathy, with severe skeletal deformities. The mutation lies within the transmembrane domain of GDAP1 and interferes with the mitochondrial targeting of the protein, similar to the loss of the domain in the previously reported Q163X and S194X mutations. We conclude that the loss of mitochondrial targeting is associated with a severe course of disease. Our study shows that clinical outcome of CMT disease caused by mutations in the GDAP1 gene cannot be predicted solely on the basis of genetic results (missense/nonsense mutations).

Mutations in the myelin protein zero (MPZ) gene are the third most frequent cause of hereditary motor and sensory neuropathies (HMSN), also called Charcot-Marie-Tooth disorders (CMT). Only in case of recurrent mutations occurring in the MPZ gene is it possible to draw phenotype-genotype correlations essential for establishing the prognosis and outcomes of CMT1. We have surveyed a cohort of 67 Polish patients from CMT families with demyelinating neuropathy for mutations in the MPZ gene. In this study, we report two CMT families in which the Ile135Thr and Pro132Leu mutations have been identified for the MPZ gene. These MPZ gene mutations had not been identified hitherto in the Polish population. The Pro132Leu mutation segregates with a severe early-onset dysmyelinating-hypomyelinating neuropathy, whereas the Ile135Thr substitution is associated with the classical phenotype of CMT1. To the best of our knowledge, we present here, for the first time, morphological data obtained in two sural nerve biopsies pointing to a hypomyelination-dysmyelination process in a family harboring the Pro132Leu mutation in the MPZ gene.

In contrast to mutations in the coding sequences of a genes involved in the pathogenesis of Charcot-Marie-Tooth disease (CMT), little is known about CMT phenotypes resulting from a DNA variants located in regulatory sequences of a given " CMT gene". Charcot-Marie-Tooth type X1 disease (CMTX1) is caused by mutations in the GJB1 gene coding for an ion channel known as connexin, with a molecular mass of 32 kDa (Cx32). Only 0.01% of the GJB1 gene mutations have been reported in its 5' regulatory sequence. Pathogenic mutations occured in the internal ribosome entry site (IRES) are extremely rarely reported in human genetic disorders. To the best of our knowledge, in this study we report for the first time in an Eastern European population, two CMTX1 families in which two pathogenic mutations in the 5' regulatory sequence of the GJB1 gene (c.-529T>C and -459C>T) have been found. The two mutations identified in our study disturb the 5' UTR sequence in two different ways, by affecting the transcription factor SOX10 binding site (c.-529T>C) and by the disrupting IRES element of GJB1 gene (c.-459C>T). These regions are responsible for transcription (SOX10) and initiation of translation (IRES), respectively. On the basis of our findings that, in contrast to the most DNA sequence variants reported in untranslated regulatory regions of genes, the c.-459C>T and c.-529T>C mutations remain pathogenic in the context of different ethnic background.

Charcot-Marie-Tooth disease 2 is an inherited axonal motor and sensory neuropathy. It is very heterogenous, both clinically and genetically. Till present, 15 types of CMT2, 14 loci and 13 genes are known to be causative of CMT2. Studying mechanisms of molecular pathogenesis is very important for finding a therapy for patients but the diversity of proteins involved in pathogenesis makes this very difficult. Proteins involved in molecular pathogenesis are e.g. proteins of the mitochondrial outer membrane with opposite functions (mitofusin 2 and GDAP1) responsible for fusion and fission of the mitochondrial network. Mutations also occur in genes encoding tRNA-synthetases, neuronal cytoskeletal protein, cation channel protein and molecular chaperones. This review presents knowledge of CMT2 and possible pathogenetic mechanisms responsible for the disease.

Over 40 mutations in the GDAP1 gene have been shown to segregate with Charcot-Marie-Tooth disease (CMT). Among these, only two mutations, i.e., S194X and Q163X have been reported in a sufficient number of CMT families to allow for the construction of reliable phenotype-genotype correlations. Both the S194X and Q163X mutations have been shown to segregate with an early-onset and severe neuropathy resulting in loss of ambulance at the beginning of the second decade of life. In this study, we identified the L239F mutation in the GDAP1 gene in one Bulgarian and five Polish families. We hypothesized that the L239F mutation may result from a founder effect in the European population since this mutation has previously been reported in Belgian, Czech, and Polish patients. In fact, we detected a common disease-associated haplotype within the 8q13-q21 region in the Polish, German, Italian, Czech, and Bulgarian CMT families. Like the previously detected "regional" S194X and Q163X mutations, respectively present in Maghreb countries and in patients of Spanish descent, the L239F mutation seems to be the most common GDAP1 pathogenic variant in the Central and Eastern European population. Given the likely presence of a common ancestor harboring the L239F mutation, we decided to compare the phenotypes of the CMT (L239F) patients collected in this study with those of previously reported cases. In contrast to CMT4A caused by the S194X and Q163X mutations, the CMT phenotype resulting from the L239F substitution represents a milder clinical entity with a long-preserved period of ambulance at least until the end of the second decade of life.

Little is known about the molecular background of clinical variability of Charcot-Marie-Tooth type 1A (CMT1A) disease and hereditary neuropathy with liability to pressure palsies (HNPP). The CMT1A and HNPP disorders result from duplication and deletion of the PMP22 gene respectively. In a series of studies performed on affected animal transgenic models of CMT1A disease, expression of the PMP22 gene (gene dosage) was shown to correlete with severity of CMT course (gene dosage effect). In this study we hypothesized that single nucleotide polymorphisms (SNPs) located within the 5' regulatory sequence of PMP22 gene may be responsible for the CMT1A/HNPP clinical variability. We have sequenced the PMP22 5' upstream regulatory sequence in a group of 45 CMT1A/HNPP patients harboring the PMP22 duplication (37) /deletion (8). We have identified five SNPs in the regulatory sequence of the PMP22 gene. Three of them i.e. -819C>T, -4785G>T, -4800C>T were detected both in the patients and in the control group. Thus, their pathogenic role in the regulation of the expression of the PMP22 gene seems not to be significant. Two SNPs i.e. -4210T>C and -4759T>A were found only in the CMT patients. Their role in the regulation of the PMP22 gene expression can not be excluded. Additionally we have detected the Thr118Met variant in exon 4 of the PMP22 gene, which was previously reported by other authors, in one patient. We conclude that the 5' regulatory sequence of the PMP22 gene is conserved at the nucleotiode level, however rarely occurring SNPs variant in the PMP22 regulatory sequence may be associated with the gene dosage effect.

Charcot-Marie-Tooth type 1X (CMT1X) disease is inherited as an X-linked dominant trait. Female CMT1X patients are usually mildly affected or even asymptomatic carriers of mutations in the GJB1 gene coding for a gap junction protein called connexin-32 (Cx32). In this report, a five-generation CMT1X family is described from which the new mutation in the GJB1 gene Cys179Gly was identified. The Cys179Gly mutation is located in the highly conservative domain of the Cx32 protein. Previous functional studies performed in the oocyte system have shown that point mutations in the highly conserved Cx32 cysteine residues result in a complete loss of function of the gap junction. However, despite severe biochemical defects, the Cys179Gly mutation segregates with a mild CMT1X phenotype. This study further documents a discrepancy between biochemical effects of GJB1 mutations and the CMT1X phenotype.

Unlabelled: THE AIM was focused on molecular analysis of the selected genes associated with autosomal recessive Charcot-Marie-Tooth neuropathies (AR-CMT) and construction of a molecular diagnostic algorithm in this group of disorders in the Polish population., Material and Methods: We analyzed a group of 138 subjects from 62 families with probably autosomal recessive inheritance and the control group of 52 individuals. The studies covered molecular genetic analysis of PMP22 gene dosage (real-time polymerase chain reaction and polymerase chain reaction restriction fragments length polymorphisms), analysis of coding regions of the GDAP1, PRX, EGR2 and CTDP1 genes using: mutation screening (single strand conformation polymorphism and heteroduplex analysis), sequencing and bioinformatics approach to the gene sequence variants., Results: Thirty sequence variants have been found in the analysed genes, 5 pathogenic mutations in the GDAP1 gene and 2 pathogenic mutations in the PRX gene. On the basis of bioinformatic analysis other nucleotide changes have been categorized as harmless polymorphisms and variants of unknown pathogenic effect., Conclusions: This is the first study focused on the autosomal recessive Charcot-Marie-Tooth disease in the Polish population. Our results show the difficulties in the interpretation of the pathogenic effect of the sequence variants (pathogenic mutation or polymorphism) which is essential for molecular diagnostics in CMT disease.

Hereditary neuropathy with liability to pressure palsies (HNPP) is manifested by a spectrum of phenotypes, from the classical HNPP course associated with intermittent nerve palsies to a neuropathy resembling Charcot-Marie-Tooth type 1 (CMT1) disease. The majority of HNPP cases are associated with submicroscopical deletions in the 17p11.2-p12 region containing the PMP22 gene, while PMP22 point mutations are rare, representing about 15% of HNPP cases. In this study, we present a patient manifesting with atypical HNPP phenotype associated with a new Thr99fsX110 mutation in the PMP22 gene. We conclude that all patients who fulfill the electrophysiological criteria of HNPP, even if they lack the typical HNPP phenotype, should be tested for point mutations in the PMP22 gene.

In the present study, we report a single Polish SMA family in which the 17p11.2-p12 duplication causative for the Charcot-Marie-Tooth type 1A disease (CMT1A) was found in addition to a deletion of exons 7 and 8 of the SMN1 gene. A patient harboring both SMA and CMT1A mutations manifested with SMA3 phenotype and foot deformity. Her electrophysiological testing showed chronic neurogenic changes in proximal muscles that are typical for SMA, but also slowed conduction velocity in motor and sensory fibers that is typical for demyelinating neuropathy.

Charcot-Marie-Tooth type 4C4 disease (CMT4C4) is an early onset, autosomal recessive neuropathy with hoarseness caused by mutations in the GDAP1 gene which maps to the 8q13 region. To date, only 24 mutations in the GDAP1 gene have been reported. Neuropathological findings of sural nerve biopsies have been published for a limited number of CMT4C4 patients. Herein, a novel Pro153Leu mutation in the GDAP1 gene identified in a consanguineous Polish family is described and longitudinal clinical and electrophysiological studies as well as morphological findings are presented.

In contrast to Charcot-Marie-Tooth type 1 disease (CMT1), which is most commonly caused by 17p11.2-p12 duplication (in 70% of CMT1 cases), the axonal form of hereditary motor and sensory neuropathy (CMT2) seemed to be a genetically heterogeneous disease group, with no single gene playing a major pathogenetic role. In 2004, 10 mutations were identified in CMT2A families in the MFN2 gene coding for the mitochondrial protein mitofusin-2, previously mapped to the 1p35-36 locus. In the last two years, MFN2 gene mutations were shown to be the most common cause of autosomal dominant hereditary axonopathy. In addition, MFN2 gene mutations were also identified in CMT type 6 (axonal neuropathy with optic nerve atrophy). Recent reports indicate that some MFN2 gene mutations may by inherited as autosomal recessive traits. As MFN2 gene mutations are the most common cause of autosomal dominant CMT2 disease (33% of cases), MFN2 gene testing may be considered a diagnostic test for CMT2.