De novo mutations of the ATP6V1A gene cause developmental encephalopathy with epilepsy.

V-type proton (H+) ATPase (v-ATPase) is a multi-subunit proton pump that regulates pH homeostasis in all eukaryotic cells; in neurons, v-ATPase plays additional and unique roles in synapse function. Through whole exome sequencing, we identified de novo heterozygous mutations (p.Pro27Arg, p.Asp100Tyr, p.Asp349Asn, p.Asp371Gly) in ATP6V1A, encoding the A subunit of v-ATPase, in four patients with developmental encephalopathy with epilepsy. Early manifestations, observed in all patients, were developmental delay and febrile seizures, evolving to encephalopathy with profound delay, hypotonic/dyskinetic quadriparesis and intractable multiple seizure types in two patients (p.Pro27Arg, p.Asp100Tyr), and to moderate delay with milder epilepsy in the other two (p.Asp349Asn, p.Asp371Gly). Modelling performed on the available prokaryotic and eukaryotic structures of v-ATPase predicted p.Pro27Arg to perturb subunit interaction, p.Asp100Tyr to cause steric hindrance and destabilize protein folding, p.Asp349Asn to affect the catalytic function and p.Asp371Gly to impair the rotation process, necessary for proton transport. We addressed the impact of p.Asp349Asn and p.Asp100Tyr mutations on ATP6V1A expression and function by analysing ATP6V1A-overexpressing HEK293T cells and patients' lymphoblasts. The p.Asp100Tyr mutant was characterized by reduced expression due to increased degradation. Conversely, no decrease in expression and clearance was observed for p.Asp349Asn. In HEK293T cells overexpressing either pathogenic or control variants, p.Asp349Asn significantly increased LysoTracker® fluorescence with no effects on EEA1 and LAMP1 expression. Conversely, p.Asp100Tyr decreased both LysoTracker® fluorescence and LAMP1 levels, leaving EEA1 expression unaffected. Both mutations decreased v-ATPase recruitment to autophagosomes, with no major impact on autophagy. Experiments performed on patients' lymphoblasts using the LysoSensor™ probe revealed lower pH of endocytic organelles for p.Asp349Asn and a reduced expression of LAMP1 with no effect on the pH for p.Asp100Tyr. These data demonstrate gain of function for p.Asp349Asn characterized by an increased proton pumping in intracellular organelles, and loss of function for p.Asp100Tyr with decreased expression of ATP6V1A and reduced levels of lysosomal markers. We expressed p.Asp349Asn and p.Asp100Tyr in rat hippocampal neurons and confirmed significant and opposite effects in lysosomal labelling. However, both mutations caused a similar defect in neurite elongation accompanied by loss of excitatory inputs, revealing that altered lysosomal homeostasis markedly affects neurite development and synaptic connectivity. This study provides evidence that de novo heterozygous ATP6V1A mutations cause a developmental encephalopathy with a pathomechanism that involves perturbations of lysosomal homeostasis and neuronal connectivity, uncovering a novel role for v-ATPase in neuronal development.

A violência comunitária e o abuso de álcool entre adolescentes: comparação entre sexos / Community violence and alcohol abuse among adolescents: a sex comparison

OBJETIVO: Verificar a existência de associação entre vitimização e uso de álcool entre meninos e meninas de Porto Alegre (RS). MÉTODOS: Foi realizado um estudo transversal. Participaram estudantes de escolas públicas com idade entre 10 e 19 anos, da 5ª série do ensino fundamental até a 3ª série do ensino médio, compondo amostra por conglomerados em dois estágios, definidas pelas características socioeconômicas do bairro da escola e pela turma. A coleta de dados foi realizada por questionário padronizado de forma anônima e voluntária. RESULTADOS: O álcool foi utilizado por 54 por cento dos meninos adolescentes (14-19) e 17 por cento dos adolescentes (10-13), por 58 por cento das meninas adolescentes mais velhas e 19 por cento das adolescentes mais novas. Cinqüenta e sete por cento dos estudantes sofreram vitimização severa, e 53 por cento sofreram vitimização moderada. Quando as amostras separadas por sexos foram avaliadas, observou-se que meninos e meninas que faziam uso de álcool relataram 2,6 e 1,8, respectivamente, mais vitimização grave, ao passo que o uso de álcool entre meninos e meninas se associou a 3,1 e 2,5 mais prevalência de vitimização moderada, respectivamente. Para os episódios de embriaguez, observou-se que adolescentes se embriagaram mais que pré-adolescentes e que as exposições à violência mostraram associação aumentada para embriaguez. Um percentual de 32 por cento de meninos (razão de prevalência, RP = 4,4; IC95 por cento 2,6-7,3) e 22 por cento de meninas (RP = 2,2; IC95 por cento 1,2-4,1) vítimas de violência severa relatou embriaguez pelo menos uma vez. CONCLUSÕES: Adolescentes de ambos os sexos que consomem mais bebidas alcoólicas têm maior risco de sofrerem violência comunitária.

OBJECTIVE: To verify whether there is an association between victimization and alcohol use among boys and girls in Porto Alegre, RS, Brazil. METHODS: This was a cross-sectional study of students from public schools, aged 10 to 19 years, from the fifth grade of primary education to the third grade of secondary education, selected by two-stage cluster sampling, defined by the socioeconomic characteristics of the neighborhood in which each school is located and by school class. Data collection was by means of a questionnaire which was completed anonymously and voluntarily. RESULTS: Fifty-four percent of the older adolescent boys (aged 14-19) drank alcohol, compared to 17 percent of the younger adolescent boys (10-13), as did 58 percent of the older adolescent girls and 19 percent of the younger adolescent girls. Fifty-seven percent of the students had suffered severe victimization, and 53 percent had suffered moderate victimization. When the samples were separated by sex, it was observed that boys and girls who drank alcohol reported 2.6 and 1.8 times more severe victimization respectively, while alcohol was associated with 3.1 and 2.5 times greater prevalence of moderate victimization, among boys and girls respectively. When episodes of drunkenness were analyzed, it was observed that adolescents got drunk more than pre-adolescents and that exposure to violence exhibited an increased association with drunkenness. Thirty-two percent of the boys (prevalence ratio, PR = 4.4; 95 percentCI 2.6-7.3) and 22 percent of the girls (PR = 2.2; 95 percentCI 1.2-4.1) who had been the victims of severe violence reported being drunk at least once. CONCLUSIONS: Adolescents of both sexes who consume more alcohol are at greater risk of suffering community violence.

Community violence and alcohol abuse among adolescents: a sex comparison.

OBJECTIVE: To verify whether there is an association between victimization and alcohol use among boys and girls in Porto Alegre, RS, Brazil. METHODS: This was a cross-sectional study of students from public schools, aged 10 to 19 years, from the fifth grade of primary education to the third grade of secondary education, selected by two-stage cluster sampling, defined by the socioeconomic characteristics of the neighborhood in which each school is located and by school class. Data collection was by means of a questionnaire which was completed anonymously and voluntarily. RESULTS: Fifty-four percent of the older adolescent boys (aged 14-19) drank alcohol, compared to 17% of the younger adolescent boys (10-13), as did 58% of the older adolescent girls and 19% of the younger adolescent girls. Fifty-seven percent of the students had suffered severe victimization, and 53% had suffered moderate victimization. When the samples were separated by sex, it was observed that boys and girls who drank alcohol reported 2.6 and 1.8 times more severe victimization respectively, while alcohol was associated with 3.1 and 2.5 times greater prevalence of moderate victimization, among boys and girls respectively. When episodes of drunkenness were analyzed, it was observed that adolescents got drunk more than pre-adolescents and that exposure to violence exhibited an increased association with drunkenness. Thirty-two percent of the boys (prevalence ratio, PR = 4.4; 95%CI 2.6-7.3) and 22% of the girls (PR = 2.2; 95%CI 1.2-4.1) who had been the victims of severe violence reported being drunk at least once. CONCLUSIONS: Adolescents of both sexes who consume more alcohol are at greater risk of suffering community violence.