Background splicing as a predictor of aberrant splicing in genetic disease

Mutations of splice sites, auxiliary splicing elements and the splicing machinery cause a wide range of genetic disease. Here we report that many of the complex effects of splicing mutations can be predicted from background splicing information, with emphasis on BRCA1, BRCA2 and DMD. Background splicing arises from very low level splicing between rarely used background splice sites and from low-level exon skipping between intron splice sites. We show how this information can be downloaded from the Snaptron database of spliced RNA, which we then compared with databases of human splice site mutations. We report that inactivating mutations of intron splice sites typically caused the non-mutated partner splice site to splice to a known background splice site in over 90% of cases and to the strongest background splice site in the large majority of cases. Consequently, background splicing information can usefully predict the effects of splice site mutations, which include cryptic splice activation and single or multiple exon skipping. In addition, de novo splice sites and splice sites involved in pseudoexon formation, recursive splicing and aberrant splicing in cancer show a 90% match to background splice sites, so establishing that the enhancement of background splicing causes a wide range of splicing aberrations. We also discuss how background splicing information can identify cryptic splice sites that might be usefully targeted by antisense oligonucleotides (ASOs) and how it might indicate possible multiple exon skipping side effects of ASOs designed to induce single exon skipping

MYT1L mutations cause intellectual disability and variable obesity by dysregulating gene expression and development of the neuroendocrine hypothalamus

Deletions at chromosome 2p25.3 are associated with a syndrome consisting of intellectual disability and obesity. The smallest region of overlap for deletions at 2p25.3 contains PXDN and MYT1L. MYT1L is expressed only within the brain in humans. We hypothesized that single nucleotide variants (SNVs) in MYT1L would cause a phenotype resembling deletion at 2p25.3. To examine this we sought MYT1L SNVs in exome sequencing data from 4, 296 parent-child trios. Further variants were identified through a genematcher-facilitated collaboration. We report 9 patients with MYT1L SNVs (4 loss of function and 5 missense). The phenotype of SNV carriers overlapped with that of 2p25.3 deletion carriers. To identify the transcriptomic consequences of MYT1L loss of function we used CRISPR-Cas9 to create a knockout cell line. Gene Ontology analysis in knockout cells demonstrated altered expression of genes that regulate gene expression and that are localized to the nucleus. These differentially expressed genes were enriched for OMIM disease ontology terms “mental retardation”. To study the developmental effects of MYT1L loss of function we created a zebrafish knockdown using morpholinos. Knockdown zebrafish manifested loss of oxytocin expression in the preoptic neuroendocrine area. This study demonstrates that MYT1L variants are associated with syndromic obesity in humans. The mechanism is related to dysregulated expression of neurodevelopmental genes and altered development of the neuroendocrine hypothalamus

O ensino da leitura e da escrita no 1º ano de escolaridade: os resultados dos alunos em leitura

Tese de Doutoramento em Psicologia Aplicada, Área de especialidade Psicologia EducacionalO presente estudo teve como objetivo caracterizar as práticas de ensino da linguagem escrita em Portugal e perceber o seu impacto na aquisição da leitura no final do 1º ano de escolaridade. A primeira etapa deste estudo correspondeu a um estudo descritivo e comparativo com o realizado por Fijalkow (2003) em França. A partir da resposta a um questionário de 883 professores foi possível verificar que os professores dos dois países se aproximam no que diz respeito aos aspetos metodológicos do ensino da leitura e à avaliação da leitura e da escrita dos alunos. Contudo, os professores portugueses dizem utilizar com maior frequência do que os professores franceses, atividades a partir de livros infantis, outros materiais para além do manual e diferentes propostas de escrita. Na segunda etapa deste estudo, o questionário utilizado foi validado através da sua estrutura fatorial e foram utilizados os fatores revelados para a realização de uma análise hierárquica de clusters. Esta análise revelou três grupos de professores: a) um grupo de professores que centra as suas práticas no ensino do código da linguagem escrita (unidades curtas); b) um grupo que realça a construção de significado na interação com o material escrito (unidades longas); c) e um grupo de professores que reúne nas suas práticas características de ambos os grupos referidos anteriormente (unidades diversificadas). A terceira etapa realizada diz respeito à validação das respostas do questionário através da observação de sala de aula. Foram selecionados 5% dos professores de cada um dos três grupos constituídos (N=42). Cada professor foi observado duas vezes durante 60 minutos em situações escolhidas por si como situações privilegiadas para o ensino da linguagem escrita. Através da utilização de uma grelha de observação que divide as atividades desenvolvidas em atividades que remetem para o código ou para o significado, foi possível confirmar as respostas ao questionário para 55% dos professores participantes. No sentido de perceber o impacto das práticas dos professores na aquisição da leitura dos alunos foram selecionados os professores cujas observações foram coincidentes com as respostas ao questionário e avaliados os seus alunos com recurso a três provas de leitura: leitura de palavras isoladas, decisão lexical e compreensão. Participaram 461 alunos distribuídos pelos três grupos referidos anteriormente. Através da realização de uma MANCOVA, controlando a escolaridade das mães dos alunos, verificou-se que os alunos do grupo unidades diversificadas obtêm melhores resultados em todas as provas do que os alunos dos outros dois grupos. Foi ainda possível constatar que não existiram diferenças de desempenho entre os alunos do grupo unidades curtas e os alunos do grupo unidades longas.ABSTRACT: The present study aimed to characterize the teaching practices of written language and understand its impact on reading acquisition by the end of 1st grade. The first step of this study was a descriptive and comparative study to the one conducted by Fijalkow (2003) in France. Based on questionnaire answers of 883 teachers, we found that teachers from both countries have similar approaches regarding methodological aspects of teaching of reading and students’ assessment of reading and writing abilities. However, Portuguese teachers stated that they use more frequently activities from children's literature, other materials besides the manual and different writing proposals than French teachers. In the second step of the study, the questionnaire was validated through its factorial structure and the factors disclosed were used to perform a hierarchical cluster analysis. This analysis revealed three groups of teachers: a) a group of teachers who focus its practices in teaching the code of written language (short units), b) a group that emphasizes the construction of meaning in interaction with the written material (long units), c) and a group of teachers who gathered in their practices characteristics of both groups mentioned above (diversified units). The third step concerns the validation of the questionnaire answers through classroom observation. 5% of the teachers in each of the three groups were selected (N=42). Each teacher was observed twice during 60 minutes in situations identified by them as privileged situations for teaching written language. Using an observation grid which divides the activities conducted in code-activities or the meaning-activities it was possible to confirm questionnaire answers for 55% of the participating teachers. In order to understand the impact of teachers’ practices in students reading acquisition, teachers whose observations were coincident with the questionnaire answers were selected and their students were evaluated using three reading tests: single words reading, lexical decision task and comprehension task. Participants were 461 students distributed through the three groups previously mentioned. By conducting a MANCOVA, controlling mothers’ education level, we concluded that students in the diversified unit group obtain better results in all reading tests than students from the other groups. It was also possible to confirm that there were no differences in performance between students of the short units group and students of the long units group.Tese de Doutoramento em Psicologia Aplicada, Área de especialidade Psicologia EducacionalApoio financeiro da Fundação para a Ciência e Tecnologia (SFRH/BD/64630/2009

Glaciations pleistocènes et terrasses climatiques en Équateur

Bruet E. Glaciations pleistocènes et terrasses climatiques en Équateur. In: Bulletin de l'Association de géographes français, N°188-189, 24e année, Novembre-décembre 1947. pp. 90-99

La morphologie pliocène de la forêt de Châteauvillain et d'Arc-en-Barrois

Bruet E., Aufrère L. La morphologie pliocène de la forêt de Châteauvillain et d'Arc-en-Barrois. In: Bulletin de l'Association de géographes français, N°47, 8e année, février 1931. pp. 22-31