RNA sequencing

A practical introduction to RNA sequencing and interpretation of gene-expression data for life-science researchers. Topics include sequencing technologies, experimental design, file formats, quality control, mapping, visualisation, functional annotation, long-read analysis and Nextflow pipelines. Participants use ENA, GEO, SRA, Expression Atlas and g:Profiler with small teaching datasets; personal-data analysis is outside the course. Pre-recorded and live teaching, exercises and discussions run through Zoom, Slack and EMBL-EBI virtual machines. Basic R and Linux experience helps but is not ess

Prolonged reduction in blood flow leads to stroke, a devastating neurological insult leading cause of adult disability worldwide. One of the early features of stroke is the disruption of neuronal activity that results from the reduction of metabolic supply. Changes in oscillatory brain patterns have been observed in several neurological disorders, including stroke. Evoked neural oscillations in the gamma range have recently shown the ability to restore and maintain intrinsic homeostatic processes in the brain. However, the causal relationship between stimulation and restoration of function is

…underlying Glut1DS, we conducted an RNA-sequencing analysis on the somatosensory and motor cerebral cortex of a mouse model for Glut1DS.Our analysis revealed 195 up-regulated and 71 down-regulated genes in Glut1-deficient mice compared to wild type controls. Gene set a

Neuronal networks are at the core of the brain's vast computational capacities. Different strategies have each brought deep insights into their architecture. Yet, each method is limited in its capacity to resolve even simple subnetworks of single cells, due to the current compromise between throughput, resolution, and coverage. Viral projection tracing helps establish regional connectomes of mammalian brains. In combination with additional modification, the method can reveal cell-type-specific projection patterns. However, these maps lack single-neuron resolution. Monosynaptic connectivity tra

Joint exploration of behavior, genes and brain structure holds the promise to provide a better understanding of neural circuits than unimodal data analyses. Several consortia provide extensive data resources of different modalities. However, analyses of heteromodal data require the expertise of computer scientists for fusing the different resources into a common space. To tackle this limitation, we extended BrainTrawler, our web-based visual analytics frame work for exploration of neurobiological data of human and mice, with a selection of publicly available gene expression and connectivity da

MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression at a post-transcriptional level. Their tissue-specific expression patterns, high stability in blood, and ability to cross the blood-brain barrier make them promising biomarker candidates for diagnosing central nervous system (CNS) disorders. However, further characterization of miRNA biomarkers is needed to identify key factors for diagnosing CNS disorders. In this study, we aimed to detect and characterize a differentially expressed miRNA as a biomarker for Major Depressive Disorder (MDD). We performed RNA sequencing o

…carriers, especially those with a paternal lineage of the premutation, exhibit an increased incidence of premature ovarian failure (Hundscheid et al., 2000). Despite these findings, the impact of FMRP absence on imprinting mechanisms in the br

Modification of mRNA by methylation is involved in post-transcriptional regulation of gene expression by affecting the splicing, transport, stability and translation of mRNA. Methylation of adenosine at N6 (m6A) is the most common and most important cellular modification occurring in the mRNA of eukaryotes. Evidence that m6A mRNA methylation is involved in regulation of stress response and that its dysregulation may contribute to the pathogenesis of neuropsychiatric disorders is accumulating. We have examined the acute and subchronic (up to 18 days once per day intraperitoneally) effect of the

Neuropathic pain is an intractable disease which in case opioids provide inadequate analgesia. It has been previously reported that voluntary wheel running prevented persistent hyperalgesia in sciatic nerve ligation (PSNL) mice. However, the mechanism of the preventive effect of the voluntary wheel running on persistent hyperalgesia was unknown. Since the anterior cingulate cortex (ACC) has been known to play an important role in chronic pain, we investigated factors associated with the attenuated hypersensitivity on voluntary wheel running in the ACC of the PSNL mice.A partial sciatic nerve l

Plant homeodomain finger gene 6 (PHF6) is a highly conserved chromatin associated protein that is important for neurodevelopment and hematopoiesis. Inherited or de novo mutations in the PHF6 gene cause Börjeson–Forssman–Lehmann syndrome (BFLS), a rare X-linked neurodevelopmental and endocrine disorder. Patients with BFLS present with intellectual disability (ID), which affects cognitive functioning and adaptive behaviour. To progress our understanding of how PHF6 mutations affect the nervous system in BFLS, we studied the impact of deletion of the Phf6 gene on brain development in conditional

…Subsequent RNA sequencing analys

Among the three ApoE isoforms, ApoE4 is a major risk factor for sporadic AD due to its impact on amyloid beta (Abeta) clearance and plaque formation. Although extensive research has connected ApoE to various AD-related pathologies beyond Abeta, its association with ApoE isoforms remains an area that requires further investigation. This study aimed to elucidate the effects of ApoE isoforms on AD pathology using human ApoE Knock-In (KI) AD mouse models. First, we generated an APPNL-G-F;hApoE (ApoE Double KI) mouse model by crossing human ApoE3 or ApoE4 KI mice with APPNL-G-F KI mice. Additionall

SYNGAP1-related encephalopathy is a rare neurodevelopmental disease characterized by diverse genetic mutations and phenotypic features. It is not known whether the blood-brain barrier (BBB) is affected in SYNGAP1 patients, therefore, this study aimed to establish a human induced pluripotent stem cell (hiPSC)-based BBB model of the SYNGAP1 disorder and identify relevant functional and molecular signatures. To establish the BBB models, SYNGAP1 knockout (KO), patient-derived hiPSC lines (Leu323Arg) and isogenic controls were differentiated into brain capillary endothelial-like cells and seeded on

…At 3 months, behavioral tests and RNA sequencing (RNAseq) of the thalamus were performed. Differential expressed genes (DEGs) and enriched categories were validated in silico via MAGMA using GWAS summary statistics of the Psych

Amyotrophic lateral sclerosis (ALS) is an incurable disease of the motoric nervous system characterized by loss of function and degeneration of upper and lower motoneurons. Models resembling clinics are required to study pathomechanisms and validate therapeutic options in ALS. Here, we reprogrammed fibroblasts from ALS patient carrying a mutation in Fused in Sarcoma (FUS) gene into induced pluripotent stem cells (iPSCs). An isogenic iPSC line was generated to serve as a control. Doxycycline-inducible NIL-cassette was introduced for motoneuron differentiation. Motoneurons were analyzed for

Persistence is a key characteristic of memory that relies on systems consolidation, a process classically defined as the gradual transfer of information from the hippocampus to the cortex for long-term memory storage. However, the underlying molecular mechanisms are unknown. DNA methylation can act as a long-term regulatory signal, therefore being a prime candidate to regulate memory duration and stabilisation within engrams– the physical substrate of a memory.Using contextual fear conditioning (CFC) and engram tagging tools in mice, we showed that reactivation of cortical engrams reflects sys

…RNA sequencing analysis revealed a signifi

…RNA sequencing and gene

Mitochondrial dysfunction has been implicated in psychiatric disorders, yet the specific cellular and molecular mechanisms remain elusive. Previous research has identified a connection between high trait anxiety and low levels of mitofusin 2 (Mfn2), a mitochondrial fusion protein, in the nucleus accumbens (NAc), a brain region crucial for motivation and goal-directed behavior. To assess the implications of Mfn2 in a cell-type specific manner, in this study we selectively manipulated Mfn2 in NAc dopamine receptor 1-expressing medium spiny neurons (D1-MSNs) using a viral vector (AAV1-Dyn-CRE) in

…its molecular basis, we integrated RNA-sequencing and micro-electrode array (MEA) recordings in hiPSCs-derived neurons carrying NDD-related mutations. Clustering analysis of 345 recordings from 35 NDD-related cell lines at 28 days in-vitro (DIVs) revealed seven discrete activity phenotypes. While a continuous phenotypic landscape of network alterations emerged, specific clusters were identified for certain disorders. At DIV49, stronger associations between activity p

Anorexia Nervosa (AN) is a multifactorial mental disorder characterized by voluntary food restriction and excessive physical activity leading to dramatic weight loss. The Brain-Derived Neurotropic Factor (BNDF) gene has been associated with AN. In rodents, both central and peripheral BDNF have been found to induce weight loss and anorexic effects. Thus, besides its function in neuronal development and mood, BDNF also acts as a metabolic regulator, making it a candidate biomarker for AN diagnosis. We used C57BL/6J female mice under 2-week 50% food restriction with access to running wheels to mi

…The mRNA containing the extended repeat is described to be toxic and forms foci with RNA-binding proteins like MBNL1-3. Here, our objective is to employ Cas13b-mediated cleavage to knock down the DMPK in DM1 patient muscle cells. Primary myoblasts from controls and DM1 patients were immortalized using lentiviral transduction of CDK4 and hTERT to generate comparable cell lines. Immortalized myoblasts were transduced with adeno-associated virus (AAV) vectors containing Cas

Dysregulated lipid homeostasis is a characteristic of brain aging and age-related neurodegenerative diseases. Perilipins, a family of proteins decorating the surface of lipid droplets (LDs), are key regulators of lipid metabolism in peripheral organs. Research indicates that the expression of perilipin2 increases in the brain with neurodegenerative diseases, such as Alzheimer’s disease, and preliminary data from our group suggest a reduced LD size in hippocampal neurons in response to the deletion of perilipin2. This project aims to investigate the connection between aging and the accumulation

Trait anxiety is a major risk factor for anxiety and stress-induced disorders in humans. However, animal models accounting for the inter-individual variability in stress vulnerability are largely lacking. Moreover, the pervasive bias of using mostly male animals in preclinical studies poorly reflects the increased prevalence of psychiatric disorders in women. Based on the threat imminence continuum theory, we designed and validated an auditory aversive conditioning-based pipeline in both female and male inbred mice. We operationalized trait anxiety by harnessing the naturally occurring variabi