X-linked myopathy with excessive autophagy (XMEA), caused by mutations of the VMA21 gene, is a strictly skeletal muscle disease. Extensive studies in yeast established VMA21 as the master assembly chaperone of V-ATPase, the complex multisubunit proton pump that acidifies organelles and that is vital to all mammalian tissues. As such, skeletal muscle disease exclusivity in XMEA is highly surprising. We now show that the severest VMA21 mutation, c.164-6t>g, does result in XMEA-typical pathology with autophagic vacuolar changes outside skeletal muscle, namely in the heart. However, even patients with this mutation do not exhibit clinical extramuscular disease, including cardiac disease, despite extreme skeletal muscle wasting to the extent of ventilation dependence. Uncovering the unique skeletal muscle vulnerability to defective organellar acidification, and resultant tissue-destructive excessive autophagy, will be informative to the understanding of muscle physiology. Alternatively, understanding extramuscular resistance to VMA21 mutation might disclose heretofore unknown mammalian V-ATPase assembly chaperones other than VMA21.

In X-linked myopathy with excessive autophagy (XMEA) progressive sarcoplasmic accumulation of autolysosomes filled with undegraded debris leads to atrophy and weakness of skeletal muscles. XMEA is caused by compromised acidification of lysosomes resulting from hypofunction of the proton pump vacuolar ATPase (V-ATPase), due to hypomorphic mutations in VMA21, whose protein product assembles V-ATPase. To what extent the cardiac muscle is affected is unknown. Therefore we performed a comprehensive cardiac evaluation in four male XMEA patients, and also examined pathology of one deceased patient's cardiac and skeletal muscle. None of the symptomatic men (aged 25-48 years) had history or symptoms of cardiomyopathy. Resting electrocardiograms and echocardiographies were normal. MRI showed normal left ventricle ejection fraction and myocardial mass. Myocardial late-gadolinium enhancement was not detected. The deceased patient's skeletal but not cardiac muscle showed characteristic accumulation of autophagic vacuoles. In conclusion, in classic XMEA the myocardium is structurally, electrically and clinically spared.

BACKGROUND:Congenital myopathies are inherited primary disorders of the muscle caused by mutations affecting structural, contractile, or regulatory proteins. In the more than 20 genes associated to these conditions, ryanodine receptor type 1 gene (RYR1) is responsible for the most common forms and is associated with a wide range of clinical phenotypes and pathological findings. Magnetic resonance imaging of muscle has been used increasingly to direct genetic testing in myopathies. PATIENT DESCRIPTION:We describe a consanguineous family affected by cystinuria type B, a metabolic condition linked to chromosome 19q13.2, and a different muscle phenotype that, although related to a congenital myopathy, does not have the striking histological features helping in direct genetic tests. RESULTS:The assessment of the selective involvement on muscle magnetic resonance imaging allowed the suspicion of RYR1 as the most likely gene responsible for this myopathy. The diagnosis was subsequently confirmed by the finding of a recessive RYR1 mutation. CONCLUSIONS:The occurrence of congenital myopathy together with cystinuria type B is reported for the first time. The use of muscle magnetic resonance imaging and the homozygosity by descent in SLC7A9, a gene flanking RYR1, allowed us to discover a new mutation in the RYR1 gene.

X-linked myopathy with excessive autophagy (XMEA) is characterized by weakness and wasting primarily of the proximal muscles of the lower extremities. Onset is usually after age 5 and progression is extremely slow with ambulation maintained well into the 50s. The heart, CNS, peripheral nervous system, and other organs are clinically spared. Pathology reveals large autophagic vacuoles enclosing incompletely degraded cytoplasmic components, which translocate to the myofiber surface and extrude their contents, forming a field of cell debris between multiplied layers of basal lamina.1 XMEA is caused by mutations of the VMA21 gene, which reduce, but do not eliminate, expression of the chief assembly chaperone (VMA21) of the main proton pump (V-ATPase [vacuolar-type H+–adenosine triphosphatase]) of all mammalian cells.2

In skeletal muscle, excitation-contraction (EC) coupling is the process whereby the voltage-gated dihydropyridine receptor (DHPR) located on the transverse tubules activates calcium release from the sarcoplasmic reticulum by activating ryanodine receptor (RyR1) Ca(2+) channels located on the terminal cisternae. This subcellular membrane specialization is necessary for proper intracellular signaling and any alterations in its architecture may lead to neuromuscular disorders. In this study, we present evidence that patients with recessive RYR1-related congenital myopathies due to primary RyR1 deficiency also exhibit downregulation of the alfa 1 subunit of the DHPR and show disruption of the spatial organization of the EC coupling machinery. We created a cellular RyR1 knockdown model using immortalized human myoblasts transfected with RyR1 siRNA and confirm that knocking down RyR1 concomitantly downregulates not only the DHPR but also the expression of other proteins involved in EC coupling. Unexpectedly, this was paralleled by the upregulation of inositol-1,4,5-triphosphate receptors; functionally however, upregulation of the latter Ca(2+) channels did not compensate for the lack of RyR1-mediated Ca(2+) release. These results indicate that in some patients, RyR1 deficiency concomitantly alters the expression pattern of several proteins involved in calcium homeostasis and that this may influence the manifestation of these diseases.

Mitochondrial Ca uptake plays key roles in cell life and death. Physiological Ca signalling regulates aerobic metabolism, while pathological Ca overload triggers cell death. The process is mediated by the Ca uniporter complex in the inner mitochondrial membrane, which includes MCU, a Ca-selective ion channel and its regulator, MICU1. Here we report mutations of MICU1 in patients with a novel disease phenotype characterised by proximal myopathy, learning difficulties and a progressive extrapyramidal movement disorder. In fibroblasts from patients with MICU1 mutations, agonist-induced mitochondrial Ca uptake at low cytosolic [Ca] was increased and cytosolic Ca signals were reduced. Whilst resting mitochondrial membrane potential was unchanged in MICU1-deficient cells, the mitochondrial network was severely fragmented. While the pathophysiology of muscular dystrophy and the core myopathies involves abnormal mitochondrial Ca handling, the MICU1 deficiency phenotype is caused by a primary defect in mitochondrial Ca signaling, demonstrating the crucial role of mitochondrial Ca uptake in man.

X-linked Myopathy with Excessive Autophagy (XMEA) is a childhood onset disease characterized by progressive vacuolation and atrophy of skeletal muscle. We show that XMEA is caused by hypomorphic alleles of the VMA21 gene, that VMA21 is the diverged human ortholog of the yeast Vma21p protein, and that like Vma21p, VMA21 is an essential assembly chaperone of the vacuolar ATPase (V-ATPase), the principal mammalian proton pump complex. Decreased VMA21 raises lysosomal pH which reduces lysosomal degradative ability and blocks autophagy. This reduces cellular free amino acids which leads to downregulation of the mTORC1 pathway, and consequent increased macroautophagy resulting in proliferation of large and ineffective autolysosomes that engulf sections of cytoplasm, merge, and vacuolate the cell. Our results uncover a novel mechanism of disease, namely macroautophagic overcompensation leading to cell vacuolation and tissue atrophy.

Michael Duchen, Francesco Muntoni, Eamonn Sheridan and colleagues show that loss-of-function mutations in MICU1 cause a recessive disorder characterized by proximal myopathy, learning difficulties and progressive extrapyramidal motor deficits. The mutations alter mitochondrial calcium homeostasis, leading to mitochondrial damage and dysfunction. Mitochondrial Ca2+ uptake has key roles in cell life and death. Physiological Ca2+ signaling regulates aerobic metabolism, whereas pathological Ca2+ overload triggers cell death. Mitochondrial Ca2+ uptake is mediated by the Ca2+ uniporter complex in the inner mitochondrial membrane1,2, which comprises MCU, a Ca2+-selective ion channel, and its regulator, MICU1. Here we report mutations of MICU1 in individuals with a disease phenotype characterized by proximal myopathy, learning difficulties and a progressive extrapyramidal movement disorder. In fibroblasts from subjects with MICU1 mutations, agonist-induced mitochondrial Ca2+ uptake at low cytosolic Ca2+ concentrations was increased, and cytosolic Ca2+ signals were reduced. Although resting mitochondrial membrane potential was unchanged in MICU1-deficient cells, the mitochondrial network was severely fragmented. Whereas the pathophysiology of muscular dystrophy3 and the core myopathies4 involves abnormal mitochondrial Ca2+ handling, the phenotype associated with MICU1 deficiency is caused by a primary defect in mitochondrial Ca2+ signaling, demonstrating the crucial role of mitochondrial Ca2+ uptake in humans.

Ryanodine receptor 1 (RYR1) mutations are a common cause of congenital myopathies associated with both dominant and recessive inheritance. Histopathological findings frequently feature central cores or multiminicores, more rarely, type 1 predominance/uniformity, fiber-type disproportion, increased internal nucleation, and fatty and connective tissue. We describe 71 families, 35 associated with dominant RYR1 mutations and 36 with recessive inheritance. Five of the dominant mutations and 35 of the 55 recessive mutations have not been previously reported. Dominant mutations, typically missense, were frequently located in recognized mutational hotspot regions, while recessive mutations were distributed throughout the entire coding sequence. Recessive mutations included nonsense and splice mutations expected to result in reduced RyR1 protein. There was wide clinical variability. As a group, dominant mutations were associated with milder phenotypes; patients with recessive inheritance had earlier onset, more weakness, and functional limitations. Extraocular and bulbar muscle involvement was almost exclusively observed in the recessive group. In conclusion, our study reports a large number of novel RYR1 mutations and indicates that recessive variants are at least as frequent as the dominant ones. Assigning pathogenicity to novel mutations is often difficult, and interpretation of genetic results in the context of clinical, histological, and muscle magnetic resonance imaging findings is essential. Hum Mutat 33: 981-988, 2012. (C) 2012 Wiley Periodicals, Inc.

X-linked myopathy with excessive autophagy (XMEA) is a childhood-onset disease characterized by progressive vacuolation and atrophy of skeletal muscle. We show that XMEA is caused by hypomorphic alleles of the VMA21 gene, that VMA21 is the diverged human ortholog of the yeast Vma21p protein, and that like Vma21p it is an essential assembly chaperone of the V-ATPase, the principal mammalian proton pump complex. Decreased VMA21 raises lysosomal pH, which reduces lysosomal degradative ability and blocks autophagy. This reduces cellular free amino acids, which upregulates the mTOR pathway and mTOR-dependent macroautophagy, resulting in proliferation of large and ineffective autolysosomes that engulf sections of cytoplasm, merge together, and vacuolate the cell. Our results uncover macroautophagic overcompensation leading to cell vacuolation and tissue atrophy as a mechanism of disease.

Lafora progressive myoclonus epilepsy is characterized by pathognomonic endoplasmic reticulum (ER)-associated polyglucosan accumulations. We previously discovered that mutations in EPM2A cause Lafora disease. Here, we identify a second gene associated with this disease, NHLRC1 (also called EPM2B ), which encodes malin, a putative E3 ubiquitin ligase with a RING finger domain and six NHL motifs. Laforin and malin colocalize to the ER, suggesting they operate in a related pathway protecting against polyglucosan accumulation and epilepsy.