Join us once a fornight for the online Neurogenomics seminar series

17th January: Dr Ricardo D’Oliveira Albanus & Dr Gina Finan

Reconstructing cellular crosstalk signaling networks in Alzheimer’s Disease reveals a novel role for SEMA6D in TREM2-dependent microglial activation

Disrupted cross-cellular communication signaling (cellular crosstalk) has been implicated in neurodegenerative diseases, including Alzheimer’s disease (AD). However, there is currently no systematic characterization of brain crosstalk networks in health and disease. We systematically characterized brain cellular crosstalk networks using single-nucleus transcriptomics data from a large cohort of control and AD brain donors (n=67). We found that crosstalk interactions between microglia and neurons were highly enriched to directly involve reported AD risk genes as ligands or receptors. Computational reconstruction of the co-expression networks associated with neuron-microglia crosstalk revealed they perturb additional known AD risk genes in microglia. We identified the interaction of neuronal SEMA6D (a PLXNA1 ligand) with a highly connected microglial regulatory sub-network involving TREM2, APOE, and HLA genes, which we predict is disrupted in late AD stages. Using CRISPR-modified human induced pluripotent stem cell (iPSC)-derived microglia and treatment with recombinant SEMA6D, we experimentally demonstrated that SEMA6D promotes microglial phagocytosis and cytokine (TNFα and IL-6) release in a TREM2-dependent manner. The novel discovery that the SEMA6D-PLXNA1/TREM2 signaling axis is involved in the regulation of microglia function demonstrates that our systematic characterization of cellular crosstalk networks is an important strategy for discovering specific mediators of significant cross-cellular interactions important to AD pathogenesis, gaining wider insights into the biology of this disease, and uncovering novel therapeutics

31st January: Dr Emily Brookes

Regulating cortical development through enhancer usage and genome topology

Dr Brookes' work focuses on understanding the mechanisms of gene regulation that underlie the complex development of the cortex. Enhancers are critical in determining appropriate spatiotemporal patterns of gene expression. Brain-derived neurotrophic factor (BDNF) promotes neuronal differentiation and survival during cortical development and is implicated in the pathogenesis of many neurological disorders. In a recently published study, we identified a novel intergenic enhancer located 170 kb from the Bdnf gene, which promotes the expression of Bdnf transcript variants during mouse neuronal differentiation and activity. Following Bdnf activation, enhancer-promoter contacts increase, and the region moves away from the repressive nuclear periphery. Bdnf enhancer activity is necessary for neuronal clustering and dendritogenesis in vitro, and for cortical development in vivo. Our findings provide the first evidence of a regulatory mechanism whereby the activation of a distal enhancer promotes Bdnf expression during brain development.

14th February: Dr Inge Holtman

The Netherlands Neurogenetics Database: Genetic susceptibility for brain disorders, clinical disease trajectories and end-stage neuropathology

The brain is a highly complex organ, and as such, many brain disorders show considerable overlap in clinical manifestations, genetic risk factors, and pathophysiological mechanisms. With up to 30% of brain disorders presenting conflicting clinical and neuropathological diagnoses, a better delineation of the relationships between these disorders might enable improved diagnosis, better prognosis, and personalized therapeutic interventions. The Netherlands Neurogenetics Database was created to integrate the extensive clinical, and neuropathological data of the Netherlands Brain Bank (NBB) with genetics data from a large number of donors.

Dr Holtman explains the major aims of the project - the integration of clinical, neuropathological and genetic data from the Netherlands Brain Bank to probe genetic susceptibility for brain disorders.

28th February: Dr Jeffrey Rogers

Comparative Primate Genomics: New insights into primate genetic variation and its implications for disease research

Over the past ten years, the cost of whole genome sequencing has dramatically declined, and access to DNA sequencing technology has improved. One consequence is that researchers investigating both the basic biology and various biomedical applications of nonhuman primates are generating an ever-increasing amount of fundamental data concerning genetic variation within nonhuman primate species. It is now clear that among rhesus macaques, baboons, marmosets and other nonhuman primates, there is a significant amount of within-species genetic diversity. Among rhesus macaques and baboons, levels of genetic variation exceed that observed across the broad diversity of human populations. This high level of nonhuman primate genetic variation has several implications. First, we find a substantial number of predicted damaging mutations that affect genes known to influence human disease, which provides naturally occurring primate models of human genetic disease. In addition, there is significant opportunity for the study of genotype-phenotype associations because the amount of functionally significant variation is higher than in humans. This presentation will provide an overview of recent insights into the population genomics of nonhuman primates and summarize particular examples of research progress resulting from the discovery of functionally important variation.

14 March: Dr Piergiorgo Salvan

Serotonin regulation of behavior via large-scale neuromodulation of serotonin receptor networks

Although we understand how serotonin receptors function at the single-cell level, what role different serotonin receptors play in regulating brain-wide activity and, in turn, human behavior, remains unknown. Here, we developed transcriptomic–neuroimaging mapping to characterize brain-wide functional signatures associated with specific serotonin receptors: serotonin receptor networks (SRNs). Probing SRNs with optogenetics–functional magnetic resonance imaging (MRI) and pharmacology in mice, we show that activation of dorsal raphe serotonin neurons differentially modulates the amplitude and functional connectivity of different SRNs, showing that receptors’ spatial distributions can confer specificity not only at the local, but also at the brain-wide, network level. In humans, using resting-state functional MRI, SRNs replicate established divisions of serotonin effects on impulsivity and negative biases. These results provide compelling evidence that heterogeneous brain-wide distributions of different serotonin receptor types may underpin behaviorally distinct modes of serotonin regulation. This suggests that serotonin neurons may regulate multiple aspects of human behavior via modulation of large-scale receptor networks.

28th March: Dr Judit Garcia-Gonzalez

PRSet: Pathway-based polygenic risk score analyses and software

Polygenic risk scores (PRSs) have been among the leading advances in biomedicine in recent years. As a proxy of genetic liability, PRSs are utilised across multiple fields and applications. While numerous statistical and machine learning methods have been developed to optimise their predictive accuracy, these typically distil genetic liability to a single number based on aggregation of an individual’s genome-wide risk alleles. This results in a key loss of information about an individual’s genetic profile, which could be critical given the functional sub-structure of the genome and the heterogeneity of complex disease. In this manuscript, we introduce a ‘pathway polygenic’ paradigm of disease risk, in which multiple genetic liabilities underlie complex diseases, rather than a single genome-wide liability.

18th April: Dr Lindsay Rizzardi

Identification of cell type-specific cis regulatory elements in Alzheimer’s disease

Cell type-specific transcriptional differences between brain tissues from donors with Alzheimer’s disease (AD) and unaffected controls have been well documented, but few studies have rigorously interrogated the regulatory mechanisms responsible for these alterations. We performed single nucleus multiomics (snRNA-seq plus snATAC-seq) on 105,332 nuclei isolated from cortical tissues from 7 AD and 8 unaffected donors to identify candidate cis-regulatory elements (CREs) involved in AD-associated transcriptional changes. We detected 319,861 significant correlations, or links, between gene expression and cell type-specific transposase accessible regions enriched for active CREs. Among these, 40,831 were unique to AD tissues. Validation experiments confirmed the activity of many regions, including several candidate regulators of APP expression. We identified ZEB1 and MAFB as candidate transcription factors playing important roles in AD-specific gene regulation in neurons and microglia, respectively. Microglia links were globally enriched for heritability of AD risk and previously identified active regulatory regions.

9th May: Brandon Le

Wnt activity reveals context-specific genetic effects on gene regulation in neural progenitors

Gene regulatory effects in bulk-post mortem brain tissues are undetected at many non-coding brain trait-associated loci. We hypothesized that context-specific genetic variant function during stimulation of a developmental signaling pathway would explain additional regulatory mechanisms. We measured chromatin accessibility and gene expression following activation of the canonical Wnt pathway in primary human neural progenitors from 82 donors. TCF/LEF motifs, brain structure-, and neuropsychiatric disorder-associated variants were enriched within Wnt-responsive regulatory elements (REs). Genetically influenced REs were enriched in genomic regions under positive selection along the human lineage. Stimulation of the Wnt pathway increased the detection of genetically influenced REs/genes by 66.2%/52.7%, and led to the identification of 397 REs primed for effects on gene expression. Context-specific molecular quantitative trait loci increased brain-trait colocalizations by up to 70%, suggesting that genetic variant effects during early neurodevelopmental patterning lead to differences in adult brain and behavioral traits.

23rd May: Dr Dervis Salih

Genetic variation associated with human longevity and Alzheimer’s disease risk act through microglia and oligodendrocyte cross-talk

The objective of this study was to gain new insights into the biological processes which connect ageing with AD and lifespan using an unbiased approach by integrating common human genetic variation associated with human lifespan or AD from Genome-Wide Association Studies (GWAS) with co-expression transcriptome networks altered with age in the mouse and human central nervous system. Our work revealed that genetic variation associated with AD was enriched in both microglial and oligodendrocytic gene networks, which show the strongest increases in expression with ageing in the hippocampus. Compellingly, longevity-associated genetic variation was enriched in a single-cell genetic network expressed by homeostatic microglia, which may drive “inflammageing.” Thus, we observed that variants contributing to ageing and AD balance different aspects of microglial function. In conclusion, this work provides new insights into cellular processes associated with ageing in the brain, and how these may contribute to the resilience of the brain against ageing or AD-risk. Our findings have important implications for developing markers indicating the physiological age of the brain and new targets for therapeutic intervention.

27th September: Dr Xin Jin

In vivo Perturb-seq: scaled investigation of gene functions in the brain

The thousands of disease risk genes and loci identified through human genetic studies far outstrip our current capacity to systematically study their functions. I will discuss our attempt to develop a scalable genetic screen approach, in vivo Perturb-seq, and apply this method to the functional evaluation of a panel of autism spectrum disorder (ASD) de novo loss-of-function risk genes. We identified recurrent and cell type-specific gene signatures from both neuronal and glial cell classes that are affected by genetic perturbations and pointed at elements of both convergent and divergent cellular effects across many ASD risk genes. Our lab, newly founded in July 2021, will use these systematic approaches, connecting genomic technology development with rigorous dissection of molecular mechanisms, to learn new insight about how complex inputs are integrated into the developing brain.

11th October: Dr Susanne Kooistra

Epigenetic regulation of innate immune memory in microglia

Microglia are the tissue-resident macrophages of the CNS. They originate in the yolk sac, colonize the CNS during embryonic development and form a self-sustaining population with limited turnover. A consequence of their relative slow turnover is that microglia can serve as a long-term memory for inflammatory or neurodegenerative events. We characterized the epigenomes and transcriptomes of microglia exposed to different stimuli; an endotoxin challenge (LPS) and genotoxic stress (DNA repair deficiency-induced accelerated aging). Whereas the enrichment of permissive epigenetic marks at enhancer regions explains training (hyperresponsiveness) of primed microglia to LPS challenge, the tolerized response of microglia seems to be regulated by loss of permissive epigenetic marks. Here, we identify that inflammatory stimuli and accelerated aging because of genotoxic stress activate distinct gene networks. These gene networks and associated biological processes are partially overlapping, which is likely driven by specific transcription factor networks, resulting in altered epigenetic signatures and distinct functional (desensitized vs. primed) microglia phenotypes.

25th October: Dr Marco Trizzino

Impact of Transposable Elements in neurodevelopment and neurodegeneration

Transposable Elements (TEs) account for ~50% of the human genome, and several lines of evidence now suggest an extensive role for these parasitic elements as a critical source of gene-regulatory novelty in mammals.  In this context, my laboratory aims at unveiling the mechanisms that TEs adopt to regulate and rewire mammalian gene regulatory networks, with strong focus to neurodevelopment and neurodegeneration.  In particular, the fundamental problem that we aim to solve is how a very recent genomic invasion by young mobile elements (SINE-VNTR-Alus = SVAs) has set the ground for human-specific traits.  Among the 5 main TE classes, SVAs are the youngest, and include 7 subfamilies being either hominid-specific (SVA-A, -B, -C, and -D) or human-specific (SVA-E, -F, and F1).  Importantly, SVAs are still replication competent in humans, and over 60% of the existing human SVA copies are located within 10 kb of a coding gene. Given their young age, SVAs are particularly relevant for understanding human evolution.  Importantly, genes proximal to human-specific SVA insertions are enriched for biological processes associated with brain development, craniofacial morphology, and cognitive behavior. Consistent with this evidence, we were able to demonstrate that this young family of transposons have provided the human genome with thousands of novel binding sites for TBR2, a transcription factor critical for human hippocampal neurogenesis, where it specifically regulates a population of progenitors called hippocampal intermediate progenitors. Using human and chimpanzee iPSC differentiation, comparative genomics, and CRISPR-interference we were able to demonstrate that cis-regulatory elements derived from human-specific SVAs account for a very significant fraction of the transcriptomic differences between human and chimpanzee intermediate progenitors. 

Additionally, my lab has been working to investigate the causes and consequences of aberrant mobilization of normally repressed transposable elements in the brains of Alzheimer’s patients.  We discovered that cDNA-RNA hybrids derived from the aberrantly mobilized transposons accumulate in the cytoplasm of the Alzheimer’s neurons, and that this leads to an innate immune response which ultimately elicits apoptosis.

15th November: Dr Jiang-An Yin

Robust and Versatile Arrayed Libraries for Human Genome-Wide CRISPR Activation, Deletion and Silencing

Genome-wide CRISPR phenotypic screens are clarifying many fundamental biological phenomena. While pooled screens can be used to study selectable features, arrayed CRISPR libraries extend the screening territory to cell-nonautonomous, biochemical and morphological phenotypes. Using a novel high-fidelity liquid-phase plasmid cloning technology, we generated two human genome-wide arrayed libraries termed T.spiezzo (gene ablation, 19,936 plasmids) and T.gonfio (gene activation and epigenetic silencing, 22,442 plasmids). Each plasmid encodes four non-overlapping single-guide RNAs (sgRNAs), each driven by a unique housekeeping promoter, as well as lentiviral and transposable vector sequences. The sgRNAs were designed to tolerate most DNA polymorphisms identified in 10,000 human genomes, thereby maximizing their versatility. Sequencing confirmed that ∼90% of each plasmid population contained ≥3 intact sgRNAs. Deletion, activation and epigenetic silencing experiments showed efficacy of 75-99%, up to 10,000x and 76-92%, respectively; lentiviral titers were ∼107/ml. As a proof of concept, we investigated the effect of individual activation of each human transcription factor (n=1,634) on the expression of the cellular prion protein PrPC. We identified 24 upregulators and 12 downregulators of PrPC expression. Hence, the T.spiezzo and T.gonfio libraries represent a powerful resource for the individual perturbation of human protein-coding genes.

22nd November: Sam Morabito

hdWGCNA: High dimensional co-expression networks enable discovery of transcriptomic drivers in complex biological systems

Biological systems are immensely complex, organized into a multi-scale hierarchy of functional units based on tightly-regulated interactions between distinct molecules, cells, organs, and organisms. While experimental methods enable transcriptome-wide measurements across millions of cells, the most ubiquitous bioinformatic tools do not support systems-level analysis. Here we present hdWGCNA, a comprehensive framework for analyzing co-expression networks in high dimensional transcriptomics data such as single-cell and spatial RNA-seq. hdWGCNA provides built-in functions for network inference, gene module identification, functional gene enrichment analysis, statistical tests for network reproducibility, and data visualization. In addition to conventional single-cell RNA-seq, hdWGCNA is capable of performing isoform-level network analysis using long-read single-cell data. We showcase hdWGCNA using publicly available single-cell datasets from Autism spectrum disorder and Alzheimer's disease brain samples, identifying disease-relevant co-expression network modules in specific cell populations. hdWGCNA is directly compatible with Seurat, a widely-used R package for single-cell and spatial transcriptomics analysis, and we demonstrate the scalability of hdWGCNA by analyzing a dataset containing nearly one million cells.