VMA21 deficiency causes an autophagic myopathy by compromising V-ATPase activity and lysosomal acidification.

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.

Brain dopamine-serotonin vesicular transport disease and its treatment.

We describe a disease encompassing infantile-onset movement disorder (including severe parkinsonism and nonambulation), mood disturbance, autonomic instability, and developmental delay, and we describe evidence supporting its causation by a mutation in SLC18A2 (which encodes vesicular monoamine transporter 2 [VMAT2]). VMAT2 translocates dopamine and serotonin into synaptic vesicles and is essential for motor control, stable mood, and autonomic function. Treatment with levodopa was associated with worsening, whereas treatment with direct dopamine agonists was followed by immediate ambulation, near-complete correction of the movement disorder, and resumption of development.

LGI2 truncation causes a remitting focal epilepsy in dogs.

One quadrillion synapses are laid in the first two years of postnatal construction of the human brain, which are then pruned until age 10 to 500 trillion synapses composing the final network. Genetic epilepsies are the most common neurological diseases with onset during pruning, affecting 0.5% of 2-10-year-old children, and these epilepsies are often characterized by spontaneous remission. We previously described a remitting epilepsy in the Lagotto romagnolo canine breed. Here, we identify the gene defect and affected neurochemical pathway. We reconstructed a large Lagotto pedigree of around 34 affected animals. Using genome-wide association in 11 discordant sib-pairs from this pedigree, we mapped the disease locus to a 1.7 Mb region of homozygosity in chromosome 3 where we identified a protein-truncating mutation in the Lgi2 gene, a homologue of the human epilepsy gene LGI1. We show that LGI2, like LGI1, is neuronally secreted and acts on metalloproteinase-lacking members of the ADAM family of neuronal receptors, which function in synapse remodeling, and that LGI2 truncation, like LGI1 truncations, prevents secretion and ADAM interaction. The resulting epilepsy onsets at around seven weeks (equivalent to human two years), and remits by four months (human eight years), versus onset after age eight in the majority of human patients with LGI1 mutations. Finally, we show that Lgi2 is expressed highly in the immediate post-natal period until halfway through pruning, unlike Lgi1, which is expressed in the latter part of pruning and beyond. LGI2 acts at least in part through the same ADAM receptors as LGI1, but earlier, ensuring electrical stability (absence of epilepsy) during pruning years, preceding this same function performed by LGI1 in later years. LGI2 should be considered a candidate gene for common remitting childhood epilepsies, and LGI2-to-LGI1 transition for mechanisms of childhood epilepsy remission.

VMA21 deficiency prevents vacuolar ATPase assembly and causes autophagic vacuolar myopathy.

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.

Dravet syndrome: seizure control and gait in adults with different SCN1A mutations.

PURPOSE: Dravet syndrome (DS) is an aggressive epileptic encephalopathy. Pharmacoresistant seizures of several types plague most patients with DS throughout their lives. Gait difficulties are a common, but inconsistent finding. The majority of cases are caused by mutations in the SCN1A gene, but little information is available about how particular mutations influence the adult phenotype. The purpose of this study is to correlate different types of SCN1A mutations and (1) seizure control, (2) occurrence of convulsive status epilepticus (cSE), and (3) the presence of crouch gait in adult patients. METHODS: In a cohort of 10 adult patients with DS caused by SCN1A mutations, we investigated seizure frequency, history of cSE, and gait. All patients were identified in the epilepsy clinic between 2009 and 2011. SCN1A mutations were divided into four different groups based on location or effect of the mutation. Retrospective chart review and recent physical examination were completed in all cases. KEY FINDINGS: All patients had a pathogenic mutation in the SCN1A gene. Four SCN1A mutations have not been described previously. Greater than 90% seizure reduction was observed (compared to childhood frequency) in six of seven patients with missense mutations in the pore-forming region (PFR) of the Na(v) 1.1 protein (group A) and nonsense mutations (group B). One patient with a splice-site mutation (group C) and another with a mutation outside the PFR (group D) became free of all types of seizures. cSE after the age of 19 years was observed in only one patient. Crouch gait, without spasticity, is identified as an element of the adult DS phenotype. However, only one half of our adult DS cohort demonstrated crouch gait. This feature was observed in five of seven patients from groups A and B. SIGNIFICANCE: This study shows that seizure control improves and cSE become less frequent in DS as patients age, independent of their SCN1A mutation type. Complete seizure freedom was seen in two patients (groups C and D). Finally, this study shows that in DS, crouch gait can be observed in up to 50% of adults with SCN1A mutation. Although no definite statistical correlations could be made due to the small number of patients, it is interesting to note that crouch gait was observed only in those patients with nonsense mutations or mutations in the PFR. Future studies with larger cohorts will be required to formally assess an association of gait abnormalities with particular SCN1A mutations.

Global landscape of protein complexes in the yeast Saccharomyces cerevisiae.

Identification of protein-protein interactions often provides insight into protein function, and many cellular processes are performed by stable protein complexes. We used tandem affinity purification to process 4,562 different tagged proteins of the yeast Saccharomyces cerevisiae. Each preparation was analysed by both matrix-assisted laser desorption/ionization-time of flight mass spectrometry and liquid chromatography tandem mass spectrometry to increase coverage and accuracy. Machine learning was used to integrate the mass spectrometry scores and assign probabilities to the protein-protein interactions. Among 4,087 different proteins identified with high confidence by mass spectrometry from 2,357 successful purifications, our core data set (median precision of 0.69) comprises 7,123 protein-protein interactions involving 2,708 proteins. A Markov clustering algorithm organized these interactions into 547 protein complexes averaging 4.9 subunits per complex, about half of them absent from the MIPS database, as well as 429 additional interactions between pairs of complexes. The data (all of which are available online) will help future studies on individual proteins as well as functional genomics and systems biology.