Novel mutations in the fukutin gene in a boy with asymptomatic hyperCKemia

Mutations in the fukutin gene were first identified in Japanese patients with classic Fukuyama congenital muscular dystrophy, a severe form of congenital muscular dystrophy associated with cobblestone lissencephaly and ocular defects. Patients of different ethnicities and with milder phenotypes, including limb girdle muscular dystrophy and cardiomyopathy without brain impairment, have also been reported. The hallmark of this disorder, regardless of the clinical outcome, is moderate-to-severe hypoglycosylation of alpha-dystroglycan in muscle sections. We describe the case of a boy harboring two novel mutations in fukutin gene and presenting a five-year history of asymptomatic hyperCKemia, without overt muscle, brain or ocular involvement. Genetic investigations, guided by the presence of moderate myopathic changes on muscle biopsy with loss of immunodetectable alpha-dystroglycan, led to a definitive diagnosis. Cardiac and echocardiographic examinations at follow-up disclosed low normal left ventricular function but no active cardiovascular symptoms. We suggest that fukutin mutations should be sought in asymptomatic hyperCKemia and subclinical heart dysfunction.     

Four Caucasian patients with mutations in the fukutin gene and variable clinical phenotype

Fukuyama congenital muscular dystrophy (FCMD) is frequent in Japan, due to a founder mutation of the fukutin gene (FKTN). Outside Japan, FKTN mutations have only been reported in a few patients with a wide spectrum of phenotypes from Walker–Warburg syndrome to limb-girdle muscular dystrophy (LGMD2M). We studied four new Caucasian patients from three unrelated families. All showed raised serum CK initially isolated in one case and muscular dystrophy. Immunohistochemical studies and haplotype analysis led us to search for mutations in FKTN. Two patients (two sisters) presented with congenital muscular dystrophy, mental retardation, and posterior fossa malformation including cysts, and brain atrophy at Brain MRI. The other two patients had normal intelligence and brain MRI. Sequencing of the FKTN gene identified three previously described mutations and two novel missense mutations. Outside Japan, fukutinopathies are associated with a large spectrum of phenotypes from isolated hyperCKaemia to severe CMD, showing a clear overlap with that of FKRP.     

Fukutin gene mutations cause dilated cardiomyopathy with minimal muscle weakness

OBJECTIVE: The fukutin gene (FKTN) is the causative gene for Fukuyama-type congenital muscular dystrophy, characterized by rather homogeneous clinical features of severe muscle wasting and hypotonia from early infancy with mental retardation. In contrast with the severe dystrophic involvement of skeletal muscle, cardiac insufficiency is quite rare. Fukuyama-type congenital muscular dystrophy is one of the disorders associated with glycosylation defects of alpha-dystroglycan, an indispensable molecule for intra-extra cell membrane linkage. METHODS: Protein and functional analyses of alpha-dystroglycan and mutation screening of FKTN and other associated genes were performed. RESULTS: Surprisingly, we identified six patients in four families showing dilated cardiomyopathy with no or minimal limb girdle muscle involvement and normal intelligence, associated with a compound heterozygous FKTN mutation. One patient died by rapid progressive dilated cardiomyopathy at 12 years old, and the other patient received cardiac implantation at 18 years old. Skeletal muscles from the patients showed minimal dystrophic features but have altered glycosylation of alpha-dystroglycan and reduced laminin binding ability. One cardiac muscle that underwent biopsy showed altered glycosylation of alpha-dystroglycan similar to that observed in a Fukuyama-type congenital muscular dystrophy patient. INTERPRETATION: FKTN mutations could cause much wider spectrum of clinical features than previously perceived, including familial dilated cardiomyopathy and mildest limb girdle muscular dystrophy.     

A novel form of recessive limb girdle muscular dystrophy with mental retardation and abnormal expression of alpha-dystroglycan

The limb girdle muscular dystrophies are a heterogeneous group of conditions characterized by proximal muscle weakness and disease onset ranging from infancy to adulthood. We report here eight patients from seven unrelated families affected by a novel and relatively mild form of autosomal recessive limb girdle muscular dystrophy (LGMD2) with onset in the first decade of life and characterized by severe mental retardation but normal brain imaging. Immunocytochemical studies revealed a significant selective reduction of alpha-dystroglycan expression in the muscle biopsies. Linkage analysis excluded known loci for both limb girdle muscular dystrophy and congenital muscular dystrophies in the consanguineous families. We consider that this represents a novel form of muscular dystrophy with associated brain involvement. The biochemical studies suggest that it may belong to the growing number of muscular dystrophies with abnormal expression of alpha-dystroglycan.     

Dystroglycan linkage and muscular dystrophy]

Dystroglycan is a key complex between basal lamina laminin, extracellularly and membrano-cytoskeleton, intracellularly. The damage of this linkage is turned out to cause muscular dystrophies. Dystroglycan knockout is lethal. Dystroglycan-associated intracellular proteins such as dystrophin, dystrobrevin, sarcoglycans, plectin and caveolin-3 are responsible for causing severe (Duchenne type) and moderate forms (Becker, LGMDs). Laminin, dystroglycan-binding extracellular protein, is deficient in the most severe form of congenital muscular dystrophy with normal intelligence and eye. Recently, a remarkable progress is made in most severe forms of congenital muscular dystrophy with anomalies of brain and eye such as Fukuyama type (Japan) and muscle-eye-brain disease (Finland). The gene product for Fukuyama type, fukutin, belongs to a family of glycosylation enzymes in bacteria and yeast. Since alpha-dystroglycan contains 14-15 o-glycans, ser/thr-mannose 2-1 GlcNAc 4-1 Gal 3-2 Sial in the middle third mucin-domain and the sial-o-glycan is essential for laminin-binding, and since alpha-dystroglycan is defective in Fukuyama type sarcolemma with anti both sugar moiety- and peptide-antidodies, defective fukutin causes incomplete o-glycosylation of alpha-dystroglycan. In '02, it is clarified that a glycosylation enzyme, POMGnT1 which modifies GlcNAc onto ser/thr-mannose, is defective in 6 MEB patients. The loss of the enzyme activity is turned out to lose alpha-dystroglycan from sarcolemma of MEB. These data strongly suggests that o-glycosylation defect of alpha-dystroglycan causes the most severe congenital muscular dystrophy such as Fukuyama type, MEB and Walker Warburg syndrome.     

Hint and luck for identification of a gene for Fukuyama muscular dystrophy, fukutin]

Fukuyama type congenital muscular dystrophy (FCMD), the second most common form of childhood muscular dystrophy in Japan, is an autosomal recessive severe muscular dystrophy, associated with brain anomalies due to neuronal overmigration. By taking advantages of the presence of a consanguineous patient with both FCMD and xeroderma pigmentosum group A, we performed homozygosity mapping using consanguineous FCMD families mainly, and localized the FCMD locus to chromosome 9q31-33. Subsequently, we have identified the gene responsible for FCMD on 9q31, which encodes a novel 461-amino-acid protein termed fukutin. Most FCMD-bearing chromosomes are derived from a single ancestral founder (87%), and a 3kb-retrotransposal insertion was found to be a founder mutation. Two independent point mutations in this gene have also been detected on chromosomes carrying the non-founder haplotype. FCMD is the first human disease to be caused by an ancient retrotransposal integration. We further identified the gene for muscle-eye-brain (MEB) disease, which encodes POMGnT1. Recent studies have revealed that posttranslational modification of alpha-dystroglycan is associated with congenital muscular dystrophy with brain malformations. Since hypoglycosylation of alpha-dystroglycan is common amongst several other disorders, a new clinical entity called alpha-dystroglycanopathy is proposed. However, only POMGnT1 (MEB) and POMT1 (WWS) are shown to have a definite enzymatic activity, and no enzymatic activity has been detected in fukutin. We show positive interactions between fukutin and POMGnT1. Fukutin may form a protein complex with POMGnT1 and modulate POMGnT1's enzymatic activity. Through cDNA microarray, we also show aberrant neuromuscular junction formation and delayed muscle fiber maturation in alpha-dystroglycanopathies, suggesting a new pathomechanism.     

The dystroglycanopathies: the new disorders of O-linked glycosylation

It has become clear in the past half decade that a number of forms of congenital muscular dystrophy are in fact congenital disorders of glycosylation. Genes for Walker Warburg syndrome, muscle-eye-brain disease, Fukuyama congenital muscular dystrophy, congenital muscular dystrophy 1C and 1D, and limb girdle muscular dystrophy 21 have been identified, and gene mutations resulting in these diseases all cause the underglycosylation of alpha dystroglycan with O-linked carbohydrates. Unlike congenital disorders of glycosylation involving the N-linked pathway, these O-linked disorders possess distinctive muscle, eye, and brain phenotypes. Studies using mice and patient tissues strongly suggest that underglycosylation of dystroglycan inhibits the binding extracellular matrix proteins, effectively divorcing this important cell adhesion molecule from its extracellular environment. Moreover, defects in dystroglycan alone can account for most, if not all, cellular pathology. Thus, these disorders are now collectively referred to as dystroglycanopathies.     

Coexisting muscular dystrophies and epilepsy in children

Muscular dystrophies are composed of a variety of genetic muscle disorders linked to different chromosomes and loci and associated with different gene mutations that lead to progressive muscle atrophy and weakness. Fukuyama congenital muscular dystrophy is frequently associated with partial and generalized epilepsy and congenital brain anomalies, including cobblestone complex and other neuronal migration defects. We report generalized convulsive epilepsy in a boy with normal brain magnetic resonance imaging and Duchenne muscular dystrophy with deletion of dystrophin gene, and we report absence epilepsy with normal brain magnetic resonance imaging in another boy with limb girdle muscular dystrophy with partial calpain deficiency. We, therefore, review coexisting muscular dystrophies and epilepsy in children. In addition to Fukuyama congenital muscular dystrophy, partial or generalized epilepsy has also been reported in the following types of muscular dystrophies, including Duchenne/Becker dystrophy, facioscapulohumeral dystrophy, congenital muscular dystrophy with partial and complete deficiency of laminin alpha2 (merosin) chain, and limb girdle muscular dystrophy with partial calpain deficiency.     

A novel FKRP mutation in congenital muscular dystrophy disrupts the dystrophin glycoprotein complex

Mutations in the gene encoding fukutin related protein (FKRP) produce a spectrum of disease including congenital muscular dystrophy and limb girdle muscular dystrophy. FKRP is one member of a class of molecules thought to be glycosyltransferases that mediate O-linked glycosylation. The primary target of these glycosyltransferases is thought to be dystroglycan. We now report two unrelated Mexican children with congenital muscular dystrophy who each have the identical, novel 1387A>G, N463D mutation. Muscle biopsies from these children show a reduction of alpha-dystroglycan and also show reduction of beta-dystroglycan, and alpha-, beta-, and gamma-sarcoglycan, suggesting that FKRP mutations can perturb membrane associated proteins beyond dystroglycan.     

Fukutin mutations in non-Japanese patients with congenital muscular dystrophy: less severe mutations predominate in patients with a non-Walker-Warburg phenotype

Six genes including POMT1, POMT2, POMGNT1, FKRP, Fukutin (FKTN) and LARGE encode proteins involved in the glycosylation of α-dystroglycan (α-DG). Abnormal glycosylation of α-DG is a common finding in Walker-Warburg syndrome (WWS), muscle-eye-brain disease (MEB), Fukuyama congenital muscular dystrophy (FCMD), congenital muscular dystrophy types 1C and 1D and some forms of autosomal recessive limb-girdle muscular dystrophy (LGMD2I, LGMD2K, LGMD2M), and is associated with mutations in the above genes. FCMD, caused by mutations in Fukutin (FKTN), is most frequent in Japan, but an increasing number of FKTN mutations are being reported outside of Japan. We describe four new patients with FKTN mutations and phenotypes ranging from: severe WWS in a Greek-Croatian patient, to congenital muscular dystrophy and cobblestone lissencephaly resembling MEB-FCMD in two Turkish patients, and limb-girdle muscular dystrophy and no mental retardation in a German patient. Four of the five different FKTN mutations have not been previously described.     

Alpha-dystroglycanopathy (FCMD, MEB, etc): abnormal glycosylation and muscular dystrophy]

Fukuyama congenital muscular dystrophy (FCMD), Walker-Warburg syndrome (WWS), and muscle-eye-brain (MEB) disease are similar disorders characterized by congenital muscular dystrophy, brain and eye anomalies. We previously identified the genes for FCMD and MEB, which encode fukutin and POMGnT1. Recent studies have revealed that posttranslational modification of alpha-dystroglycan is associated with congenital muscular dystrophy with brain malformations. Since hypoglycosylation of alpha-dystroglycan is common amongst several other disorders, a new clinical entity called alpha-dystroglycanopathy is proposed. However, only POMGnT1 (MEB) and POMT1 (WWS) are shown to have a definite enzymatic activity, and no enzymatic activity has been detected in fukutin. We show positive interactions between fukutin and POMGnT1. Fukutin may form a protein complex with POMGnT1 and modulate POMGnT1's enzymatic activity. Through cDNA microarray, we also show aberrant neuromuscular junction formation and delayed muscle fiber maturation in alpha-dystroglycanopathies, suggesting a new pathomechanism.     

Expression of genes related to muscular dystrophy with lissencephaly

There is a group of congenital muscular dystrophies accompanying the brain lesions termed cobblestone lissencephaly. Abnormal glia limitans could be considered the major pathogenesis of cobblestone lissencephaly. In this group, protein-O-linked mannose-beta1,2-N-acetylglucosaminyltransferase and protein-O-mannosyltransferase 1 are considered to be responsible for muscle-eye-brain disease and Walker-Warburg syndrome, respectively, by glycosylation of alpha-dystroglycan. However, the functions of fukutin, a gene responsible for Fukuyama type congenital muscular dystrophy, are still unclear. In this study, expression of the three aforementioned genes was compared by in situ hybridization in control cases to elucidate the functions of fukutin.Immunohistochemistry of fukutin and alpha-dystroglycan was also performed. In the central nervous system, all three genes were expressed in astrocytes and in immature neurons. A few mature neurons expressed fukutin, but many expressed the other two genes. All genes were expressed in various non-nervous tissues including tissues relating to secretion. Fukutin and alpha-dystroglycan were generally colocalized, but localization was not always the same, especially in the liver. Fukutin may be associated with the glycosylation of alpha-dystroglycan, and expression in astrocytes may indicate a relation to glia limitans. The roles of fukutin in mature neurons may be less critical compared with the other two genes. Additional functions of fukutin, especially in the liver, are suspected.     

A new mutation of the fukutin gene causing late-onset limb girdle muscular dystrophy

Defects in glycosylations of α-dystroglycan are associated with mutations in several genes, including the fukutin gene (FKTN). Hypoglycosylation of α-dystroglycan results in several forms of muscular dystrophy with variable phenotype. Outside Japan, the prevalence of muscular dystrophies related to aberrations of FKTN is rare, with only eight reported cases of limb girdle phenotype (LGMD2M). We describe the mildest affected patient outside Japan with genetically confirmed LGMD2M and onset of symptoms at age 14. She was brought to medical attention at age 12, not because of muscle weakness, but due to episodes of tachycardia caused by Wolff–Parkinson–White syndrome. On examination, she had rigid spine syndrome, a typical limb girdle dystrophy pattern of muscle weakness, cardiomyopathy, and serum CK levels >2000 IU/L (normal <150 IU/L). A homozygous, novel c.917A>G; p.Y306C mutation in the FKTN gene was found. The case confirms FKTN mutations as a cause of LGMD2M without mental retardation and expands the phenotypic spectrum for LGMD2M to include cardiomyopathy and rigid spine syndrome in the mildest affected non-Japanese patient reported so far.     

An autosomal recessive limb girdle muscular dystrophy (LGMD2) with mild mental retardation is allelic to Walker-Warburg syndrome (WWS) caused by a mutation in the POMT1 gene

Mutations of the protein O-mannosyltransferase (POMT1) gene affect glycosylation of alpha-dystroglycan, leading to Walker-Warburg syndrome, a lethal disorder in early life with severe congenital muscular dystrophy, and brain and eye malformations. Recently, we described a novel form of recessive limb girdle muscular dystrophy with mild mental retardation, associated with an abnormal alpha-dystroglycan pattern in the muscle, suggesting a glycosylation defect. Here, we present evidence that this distinct phenotype results from a common mutation (A200P) in the POMT1 gene. Our findings further expand the phenotype of glycosylation disorders linked to POMT1 mutations. Furthermore, the A200P mutation is part of a conserved core haplotype, indicating an ancestral founder mutation.     

Basement membrane fragility underlies embryonic lethality in fukutin-null mice

Fukuyama-type congenital muscular dystrophy (FCMD), associated with brain malformation due to defects in neuronal migration, is caused by mutations in fukutin. Several lines of evidence suggest that the fukutin protein plays a pivotal role in synthesis of O-mannosyl sugar moieties of alpha-dystroglycan, a cell surface laminin receptor. Here, through targeted disruption of the orthologous mouse fukutin gene, we show that the fukutin protein is essential, as homozygous-null embryos die by E9.5 of gestation. Fukutin-null embryos show phenotypic diversity, features of which include growth retardation, folding of the egg cylinder, leakage of maternal red blood cells into the yolk sac cavity, and an increased number of apoptotic cells in the ectoderm. Loss of immunoreactivity against sugar moieties in alpha-dystroglycan suggests a reduced laminin-binding capacity. Ultrastructural analysis shows thin and breached basement membranes (BMs). BM fragility may underlie all of these abnormal phenotypes, and maintenance of BM function may require fukutin-mediated glycosylation of alpha-dystroglycan early in embryonic development.     

Further evidence of Fukutin mutations as a cause of childhood onset limb-girdle muscular dystrophy without mental retardation

The dystroglycanopathies comprise a clinically and genetically heterogeneous group of muscular dystrophies characterized by deficient glycosylation of α-dystroglycan. Mutations in the fukutin (FKTN) gene have primarily been identified among patients with classic Fukuyama congenital muscular dystrophy (FCMD), a severe form of dystroglycanopathy characterized by CMD, cobblestone lissencephaly and ocular defects. We describe two brothers of Caucasian and Japanese ancestry with normal intelligence and limb-girdle muscular dystrophy (LGMD) due to compound heterozygous FKTN mutations. Muscle biopsy showed a dystrophy with selectively reduced α-dystroglycan glycoepitope immunostaining. Immunoblots revealed hypoglycosylation of α-dystroglycan and loss of laminin binding. FKTN gene sequencing identified two variants: c.340G>A and c.527T>C, predicting missense mutations p.A114T and p.F176S, respectively. Our results provide further evidence for ethnic and allelic heterogeneity and the presence of milder phenotypes in FKTN-dystroglycanopathy despite a substantial degree of α-dystroglycan hypoglycosylation in skeletal muscle.     

The Fukuyama congenital muscular dystrophy story

Fukuyama congenital muscular dystrophy is one of the most common autosomal recessive disorders in the Japanese population, characterized by congenital muscular dystrophy in combination with cortical dysgenesis (micropolygyria). Recently, we have identified the gene responsible for fukuyama congenital muscular dystrophy on 9q31, which encodes a novel 461-amino-acid protein termed fukutin. Most Fukuyama congenital muscular dystrophy-bearing chromosomes are derived from a single ancestral founder (87%), and a 3 kb-retrotransposal insertion into the 3' untranslated region of this gene was found to be a founder mutation. Two independent point mutations causing premature termination confirmed that that this gene is responsible for Fukuyama congenital muscular dystrophy. Fukuyama congenital muscular dystrophy is the first human disease to be caused by an ancient retrotransposal integration. Fukutin contains an amino-terminal signal sequence, which together with results from transfection experiments suggests that it is an extracellular protein. Discovery of the Fukuyama congenital muscular dystrophy gene represents an important step toward greater understanding of the pathogenesis of muscular dystrophies and also of normal brain development.     

A new mutation of the fukutin gene in a non-Japanese patient

Fukuyama-type congenital muscular dystrophy (FCMD), Walker-Warburg syndrome, and muscle-eye-brain disease are clinically similar autosomal recessive disorders characterized by congenital muscular dystrophy, cobblestone lissencephaly, and eye anomalies. FCMD is frequent in Japan, but no FCMD patient with confirmed fukutin gene mutations has been identified in a non-Japanese population. Here, we describe a Turkish CMD patient with severe brain and eye anomalies. Sequence analysis of the patient's DNA identified a homozygous 1bp insertion mutation in exon 5 of the fukutin gene. To our knowledge, this is the first case worldwide in which a fukutin mutation has been found outside the Japanese population. This report emphasizes the importance of considering fukutin mutations for diagnostic purposes outside of Japan.     

Episodes of exercise-induced dark urine and myalgia in LGMD 2I

Lindberg C, Sixt C, Oldfors A. Episodes of exercise‐induced dark urine and myalgia in LGMD 2I.
Acta Neurol Scand: 2012: 125: 285–287.
© 2011 John Wiley & Sons A/S. Background – Mutations in the fukutin‐related protein gene FKRP (MIM *606596) cause a form of congenital muscular dystrophy (MDC1C) and also limb girdle muscular dystrophy type 2I (LGMD2I). Exercise‐induced myoglobinuria, frequently occurring in metabolic myopathies, has been described in Becker muscular dystrophy and in a few cases of LGMD. Objectives – To describe that episodes with myoglobinuria, often associated with exercise‐induced myalgia, may be common and a presenting symptom in patients with LGMD2I. Methods – Data on episodes of suspected myoglobinuria and myalgia were collected from the patient records on 14 patients with a diagnosis of LGMDI. Results – Five LGMD2I patients reported recurrent episodes of dark urine and myalgia after exercise, and in three of them, this was the only symptom for several years. Conclusions – We conclude that episodes compatible with exercise‐induced myoglobinuria may be frequent in LGMD2I.     

Walker-Warburg Syndrome with POMT1 mutations can be associated with cleft lip and cleft palate

Walker-Warburg Syndrome (WWS) is an alpha-dystroglycan deficient congenital muscular dystrophy that is associated with brain and eye abnormalities. Patients present with hypotonia, weakness, developmental delay, mental retardation and occasional seizures. Other abnormalities were also described including cleft lip and palate. Mutations in POMT1, POMT2, fukutin, FKRP and LARGE genes are found in 20-30% of children with WWS. We report a novel mutation in POMT1 gene and provide further evidence that WWS with cleft lip and palate is associated with POMT1 mutations. We recommend POMT1 analysis in WWS cases associated with cleft lip and palate when considering which gene to sequence first.