The central Blue Straggler population in four outer-halo globular clusters

Using HST/WFPC2 data, we have performed a comparative study of the Blue Straggler Star (BSS) populations in the central regions of the globular clusters AM 1, Eridanus, Palomar 3, and Palomar 4. Located at distances RGC > 50 kpc from the Galactic Centre, these are (together with Palomar 14 and NGC 2419) the most distant clusters in the Halo. We determine their colour-magnitude diagrams and centres of gravity. The four clusters turn out to have similar ages (10.5-11 Gyr), significantly smaller than those of the inner-Halo globulars, and similar metallicities. By exploiting wide field ground based data, we build the most extended radial density profiles from resolved star counts ever published for these systems. These are well reproduced by isotropic King models of relatively low concentration. BSSs appear to be significantly more centrally segregated than red giants in all globular clusters, in agreement with the estimated core and half-mass relaxation times which are smaller than the cluster ages. Assuming that this is a signature of mass segregation, we conclude that AM 1 and Eridanus are slightly dynamically more evolved than Pal 3 and Pal 4.Comment: Accepted for publication in ApJ, references update

Allelome.PRO, a pipeline to define allele-specific genomic features from high-throughput sequencing data

Detecting allelic biases from high-throughput sequencing data requires an approach that maximises sensitivity while minimizing false positives. Here, we present Allelome.PRO, an automated user-friendly bioinformatics pipeline, which uses high-throughput sequencing data from reciprocal crosses of two genetically distinct mouse strains to detect allele-specific expression and chromatin modifications. Allelome.PRO extends approaches used in previous studies that exclusively analyzed imprinted expression to give a complete picture of the ‘allelome’ by automatically categorising the allelic expression of all genes in a given cell type into imprinted, strain-biased, biallelic or non-informative. Allelome.PRO offers increased sensitivity to analyze lowly expressed transcripts, together with a robust false discovery rate empirically calculated from variation in the sequencing data. We used RNA-seq data from mouse embryonic fibroblasts from F1 reciprocal crosses to determine a biologically relevant allelic ratio cutoff, and define for the first time an entire allelome. Furthermore, we show that Allelome.PRO detects differential enrichment of H3K4me3 over promoters from ChIP-seq data validating the RNA-seq results. This approach can be easily extended to analyze histone marks of active enhancers, or transcription factor binding sites and therefore provides a powerful tool to identify candidate cis regulatory elements genome wide

Long non-coding RNAs display higher natural expression variation than protein-coding genes in healthy humans

Background: Long non-coding RNAs (lncRNAs) are increasingly implicated as gene regulators and may ultimately be more numerous than protein-coding genes in the human genome. Despite large numbers of reported lncRNAs, reference annotations are likely incomplete due to their lower and tighter tissue-specific expression compared to mRNAs. An unexplored factor potentially confounding lncRNA identification is inter-individual expression variability. Here, we characterize lncRNA natural expression variability in human primary granulocytes. Results: We annotate granulocyte lncRNAs and mRNAs in RNA-seq data from 10 healthy individuals, identifying multiple lncRNAs absent from reference annotations, and use this to investigate three known features (higher tissue-specificity, lower expression, and reduced splicing efficiency) of lncRNAs relative to mRNAs. Expression variability was examined in seven individuals sampled three times at 1- or more than 1-month intervals. We show that lncRNAs display significantly more inter-individual expression variability compared to mRNAs. We confirm this finding in two independent human datasets by analyzing multiple tissues from the GTEx project and lymphoblastoid cell lines from the GEUVADIS project. Using the latter dataset we also show that including more human donors into the transcriptome annotation pipeline allows identification of an increasing number of lncRNAs, but minimally affects mRNA gene number. Conclusions: A comprehensive annotation of lncRNAs is known to require an approach that is sensitive to low and tight tissue-specific expression. Here we show that increased inter-individual expression variability is an additional general lncRNA feature to consider when creating a comprehensive annotation of human lncRNAs or proposing their use as prognostic or disease markers

Haploinsufficiency of the intellectual disability gene SETD5 disturbs developmental gene expression and cognition

SETD5 gene mutations have been identified as a frequent cause of idiopathic intellectual disability. Here we show that Setd5-haploinsufficient mice present developmental defects such as abnormal brain-to-body weight ratios and neural crest defect-associated phenotypes. Furthermore, Setd5-mutant mice show impairments in cognitive tasks, enhanced long-term potentiation, delayed ontogenetic profile of ultrasonic vocalization, and behavioral inflexibility. Behavioral issues are accompanied by abnormal expression of postsynaptic density proteins previously associated with cognition. Our data additionally indicate that Setd5 regulates RNA polymerase II dynamics and gene transcription via its interaction with the Hdac3 and Paf1 complexes, findings potentially explaining the gene expression defects observed in Setd5-haploinsufficient mice. Our results emphasize the decisive role of Setd5 in a biological pathway found to be disrupted in humans with intellectual disability and autism spectrum disorder

Cul3 regulates cytoskeleton protein homeostasis and cell migration during a critical window of brain development

De novo loss of function mutations in the ubiquitin ligase-encoding gene Cullin3 lead to autism spectrum disorder (ASD). In mouse, constitutive haploinsufficiency leads to motor coordination deficits as well as ASD-relevant social and cognitive impairments. However, induction of Cul3 haploinsufficiency later in life does not lead to ASD-relevant behaviors, pointing to an important role of Cul3 during a critical developmental window. Here we show that Cul3 is essential to regulate neuronal migration and, therefore, constitutive Cul3 heterozygous mutant mice display cortical lamination abnormalities. At the molecular level, we found that Cul3 controls neuronal migration by tightly regulating the amount of Plastin3 (Pls3), a previously unrecognized player of neural migration. Furthermore, we found that Pls3 cell-autonomously regulates cell migration by regulating actin cytoskeleton organization, and its levels are inversely proportional to neural migration speed. Finally, we provide evidence that cellular phenotypes associated with autism-linked gene haploinsufficiency can be rescued by transcriptional activation of the intact allele in vitro, offering a proof of concept for a potential therapeutic approach for ASDs

Impaired amino acid transport at the blood brain barrier is a cause of autism spectrum disorder

Autism spectrum disorders (ASD) are a group of genetic disorders often overlapping with other neurological conditions. We previously described abnormalities in the branched-chain amino acid (BCAA) catabolic pathway as a cause of ASD. Here, we show that the solute carrier transporter 7a5 (SLC7A5), a large neutral amino acid transporter localized at the blood brain barrier (BBB), has an essential role in maintaining normal levels of brain BCAAs. In mice, deletion of Slc7a5 from the endothelial cells of the BBB leads to atypical brain amino acid profile, abnormal mRNA translation, and severe neurological abnormalities. Furthermore, we identified several patients with autistic traits and motor delay carrying deleterious homozygous mutations in the SLC7A5 gene. Finally, we demonstrate that BCAA intracerebroventricular administration ameliorates abnormal behaviors in adult mutant mice. Our data elucidate a neurological syndrome defined by SLC7A5 mutations and support an essential role for the BCAA in human brain function

ISTA Thesis

Autism spectrum disorders (ASDs) are a group of neurodevelopmental disorders characterized by behavioral symptoms such as problems in social communication and interaction, as well as repetitive, restricted behaviors and interests. These disorders show a high degree of heritability and hundreds of risk genes have been identifed using high throughput sequencing technologies. This genetic heterogeneity has hampered eforts in understanding the pathogenesis of ASD but at the same time given rise to the concept of convergent mechanisms. Previous studies have identifed that risk genes for ASD broadly converge onto specifc functional categories with transcriptional regulation being one of the biggest groups. In this thesis, I focus on this subgroup of genes and investigate the gene regulatory consequences of some of them in the context of neurodevelopment. First, we showed that mutations in the ASD and intellectual disability risk gene Setd5 lead to perturbations of gene regulatory programs in early cell fate specifcation. In addition, adult animals display abnormal learning behavior which is mirrored at the transcriptional level by altered activity dependent regulation of postsynaptic gene expression. Lastly, we link the regulatory function of Setd5 to its interaction with the Paf1 and the NCoR complex. Second, by modeling the heterozygous loss of the top ASD gene CHD8 in human cerebral organoids we demonstrate profound changes in the developmental trajectories of both inhibitory and excitatory neurons using single cell RNA-sequencing. While the former were generated earlier in CHD8+/- organoids, the generation of the latter was shifted to later times in favor of a prolonged progenitor expansion phase and ultimately increased organoid size. Finally, by modeling heterozygous mutations for four ASD associated chromatin modifers, ASH1L, KDM6B, KMT5B, and SETD5 in human cortical spheroids we show evidence of regulatory convergence across three of those genes. We observe a shift from dorsal cortical excitatory neuron fates towards partially ventralized cell types resembling cells from the lateral ganglionic eminence. As this project is still ongoing at the time of writing, future experiments will aim at elucidating the regulatory mechanisms underlying this shift with the aim of linking these three ASD risk genes through biological convergence

Supplementary data for the research paper "Haploinsufficiency of the intellectual disability gene SETD5 disturbs developmental gene expression and cognition"

This dataset contains the supplementary data for the research paper "Haploinsufficiency of the intellectual disability gene SETD5 disturbs developmental gene expression and cognition". The contained files have the following content: 'Supplementary Figures.pdf' Additional figures (as referenced in the paper). 'Supplementary Table 1. Statistics.xlsx' Details on statistical tests performed in the paper. 'Supplementary Table 2. Differentially expressed gene analysis.xlsx' Results for the differential gene expression analysis for embryonic (E9.5; analysis with edgeR) and in vitro (ESCs, EBs, NPCs; analysis with DESeq2) samples. 'Supplementary Table 3. Gene Ontology (GO) term enrichment analysis.xlsx' Results for the GO term enrichment analysis for differentially expressed genes in embryonic (GO E9.5) and in vitro (GO ESC, GO EBs, GO NPCs) samples. Differentially expressed genes for in vitro samples were split into upregulated and downregulated genes (up/down) and the analysis was performed on each subset (e.g. GO ESC up / GO ESC down). 'Supplementary Table 4. Differentially expressed gene analysis for CFC samples.xlsx' Results for the differential gene expression analysis for samples from adult mice before (HC - Homecage) and 1h and 3h after contextual fear conditioning (1h and 3h, respectively). Each sheet shows the results for a different comparison. Sheets 1-3 show results for comparisons between timepoints for wild type (WT) samples only and sheets 4-6 for the same comparisons in mutant (Het) samples. Sheets 7-9 show results for comparisons between genotypes at each time point and sheet 10 contains the results for the analysis of differential expression trajectories between wild type and mutant. 'Supplementary Table 5. Cluster identification.xlsx' Results for k-means clustering of genes by expression. Sheet 1 shows clustering of just the genes with significantly different expression trajectories between genotypes. Sheet 2 shows clustering of all genes that are significantly differentially expressed in any of the comparisons (includes also genes with same trajectories). 'Supplementary Table 6. GO term cluster analysis.xlsx' Results for the GO term enrichment analysis and EWCE analysis for enrichment of cell type specific genes for each cluster identified by clustering genes with different expression trajectories (see Table S5, sheet 1). 'Supplementary Table 7. Setd5 mass spectrometry results.xlsx' Results showing proteins interacting with Setd5 as identified by mass spectrometry. Sheet 1 shows protein protein interaction data generated from these results (combined with data from the STRING database. Sheet 2 shows the results of the statistical analysis with limma. 'Supplementary Table 8. PolII ChIP-seq analysis.xlsx' Results for the Chip-Seq analysis for binding of RNA polymerase II (PolII). Sheet 1 shows results for differential binding of PolII at the transcription start site (TSS) between genotypes and sheets 2+3 show the corresponding GO enrichment analysis for these differentially bound genes. Sheet 4 shows RNAseq counts for genes with increased binding of PolII at the TSS