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1.
medRxiv ; 2023 Feb 08.
Article in English | MEDLINE | ID: mdl-36798226

ABSTRACT

Genome-wide association studies (GWAS) have identified many modifiers of Alzheimer disease (AD) risk enriched in microglia. Two of these modifiers are common variants in the MS4A locus (rs1582763: protective and rs6591561: risk) and serve as major regulators of CSF sTREM2 levels. To understand their functional impact on AD, we used single nucleus transcriptomics to profile brains from carriers of these variants. We discovered a "chemokine" microglial subpopulation that is altered in MS4A variant carriers and for which MS4A4A is the major regulator. The protective variant increases MS4A4A expression and shifts the chemokine microglia subpopulation to an interferon state, while the risk variant suppresses MS4A4A expression and reduces this subpopulation of microglia. Our findings provide a mechanistic explanation for the AD variants in the MS4A locus. Further, they pave the way for future mechanistic studies of AD variants and potential therapeutic strategies for enhancing microglia resilience in AD pathogenesis.

2.
Science ; 377(6608): eabi8654, 2022 08 19.
Article in English | MEDLINE | ID: mdl-35981026

ABSTRACT

Predicting the function of noncoding variation is a major challenge in modern genetics. In this study, we used massively parallel reporter assays to screen 5706 variants identified from genome-wide association studies for both Alzheimer's disease (AD) and progressive supranuclear palsy (PSP), identifying 320 functional regulatory variants (frVars) across 27 loci, including the complex 17q21.31 region. We identified and validated multiple risk loci using CRISPR interference or excision, including complement 4 (C4A) and APOC1 in AD and PLEKHM1 and KANSL1 in PSP. Functional variants disrupt transcription factor binding sites converging on enhancers with cell type-specific activity in PSP and AD, implicating a neuronal SP1-driven regulatory network in PSP pathogenesis. These analyses suggest that noncoding genetic risk is driven by common genetic variants through their aggregate activity on specific transcriptional programs.


Subject(s)
Alzheimer Disease , Chromosomes, Human, Pair 17 , Gene Regulatory Networks , Genetic Variation , Untranslated Regions , Alzheimer Disease/genetics , Chromosomes, Human, Pair 17/genetics , Genes, Reporter , Genetic Loci , Genome-Wide Association Study , Humans , Risk Factors , Supranuclear Palsy, Progressive/genetics , Untranslated Regions/genetics
3.
Nat Neurosci ; 25(9): 1149-1162, 2022 09.
Article in English | MEDLINE | ID: mdl-35953545

ABSTRACT

Microglia are emerging as key drivers of neurological diseases. However, we lack a systematic understanding of the underlying mechanisms. Here, we present a screening platform to systematically elucidate functional consequences of genetic perturbations in human induced pluripotent stem cell-derived microglia. We developed an efficient 8-day protocol for the generation of microglia-like cells based on the inducible expression of six transcription factors. We established inducible CRISPR interference and activation in this system and conducted three screens targeting the 'druggable genome'. These screens uncovered genes controlling microglia survival, activation and phagocytosis, including neurodegeneration-associated genes. A screen with single-cell RNA sequencing as the readout revealed that these microglia adopt a spectrum of states mirroring those observed in human brains and identified regulators of these states. A disease-associated state characterized by osteopontin (SPP1) expression was selectively depleted by colony-stimulating factor-1 (CSF1R) inhibition. Thus, our platform can systematically uncover regulators of microglial states, enabling their functional characterization and therapeutic targeting.


Subject(s)
Induced Pluripotent Stem Cells , Microglia , Brain/metabolism , Clustered Regularly Interspaced Short Palindromic Repeats/genetics , Humans , Induced Pluripotent Stem Cells/metabolism , Microglia/metabolism , Phagocytosis/genetics
4.
Autophagy ; 17(10): 3160-3174, 2021 10.
Article in English | MEDLINE | ID: mdl-33404278

ABSTRACT

We investigated in larval and adult Drosophila models whether loss of the mitochondrial chaperone Hsc70-5 is sufficient to cause pathological alterations commonly observed in Parkinson disease. At affected larval neuromuscular junctions, no effects on terminal size, bouton size or number, synapse size, or number were observed, suggesting that we studied an early stage of pathogenesis. At this stage, we noted a loss of synaptic vesicle proteins and active zone components, delayed synapse maturation, reduced evoked and spontaneous excitatory junctional potentials, increased synaptic fatigue, and cytoskeleton rearrangements. The adult model displayed ATP depletion, altered body posture, and susceptibility to heat-induced paralysis. Adult phenotypes could be suppressed by knockdown of dj-1ß, Lrrk, DCTN2-p50, DCTN1-p150, Atg1, Atg101, Atg5, Atg7, and Atg12. The knockdown of components of the macroautophagy/autophagy machinery or overexpression of human HSPA9 broadly rescued larval and adult phenotypes, while disease-associated HSPA9 variants did not. Overexpression of Pink1 or promotion of autophagy exacerbated defects.Abbreviations: AEL: after egg laying; AZ: active zone; brp: bruchpilot; Csp: cysteine string protein; dlg: discs large; eEJPs: evoked excitatory junctional potentials; GluR: glutamate receptor; H2O2: hydrogen peroxide; mEJP: miniature excitatory junctional potentials; MT: microtubule; NMJ: neuromuscular junction; PD: Parkinson disease; Pink1: PTEN-induced putative kinase 1; PSD: postsynaptic density; SSR: subsynaptic reticulum; SV: synaptic vesicle; VGlut: vesicular glutamate transporter.


Subject(s)
Drosophila Proteins , Drosophila , Animals , Autophagy/genetics , Autophagy-Related Protein 7/metabolism , Autophagy-Related Proteins/metabolism , Drosophila/metabolism , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Hydrogen Peroxide , Nerve Tissue Proteins/metabolism , Neuromuscular Junction/metabolism , Protein Deglycase DJ-1/metabolism , Protein Serine-Threonine Kinases
5.
EMBO Rep ; 18(11): 2051-2066, 2017 11.
Article in English | MEDLINE | ID: mdl-28893863

ABSTRACT

Endocytic processes are facilitated by both curvature-generating BAR-domain proteins and the coordinated polymerization of actin filaments. Under physiological conditions, the N-BAR protein Bin1 has been shown to sense and curve membranes in a variety of cellular processes. Recent studies have identified Bin1 as a risk factor for Alzheimer's disease, although its possible pathological function in neurodegeneration is currently unknown. Here, we report that Bin1 not only shapes membranes, but is also directly involved in actin binding through its BAR domain. We observed a moderate actin bundling activity by human Bin1 and describe its ability to stabilize actin filaments against depolymerization. Moreover, Bin1 is also involved in stabilizing tau-induced actin bundles, which are neuropathological hallmarks of Alzheimer's disease. We also provide evidence for this effect in vivo, where we observed that downregulation of Bin1 in a Drosophila model of tauopathy significantly reduces the appearance of tau-induced actin inclusions. Together, these findings reveal the ability of Bin1 to modify actin dynamics and provide a possible mechanistic connection between Bin1 and tau-induced pathobiological changes of the actin cytoskeleton.


Subject(s)
Actins/genetics , Adaptor Proteins, Signal Transducing/genetics , Carrier Proteins/genetics , Drosophila Proteins/genetics , Drosophila melanogaster/genetics , Nuclear Proteins/genetics , Tauopathies/genetics , Transcription Factors/genetics , Tumor Suppressor Proteins/genetics , tau Proteins/genetics , Actin Cytoskeleton/genetics , Actin Cytoskeleton/metabolism , Actins/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Animals , Binding Sites , Carrier Proteins/metabolism , Cloning, Molecular , Disease Models, Animal , Drosophila Proteins/metabolism , Drosophila melanogaster/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Gene Expression Regulation , Genetic Vectors/chemistry , Genetic Vectors/metabolism , Humans , Nuclear Proteins/metabolism , Protein Binding , Protein Interaction Domains and Motifs , Protein Isoforms/genetics , Protein Isoforms/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Tauopathies/metabolism , Tauopathies/pathology , Transcription Factors/metabolism , Tumor Suppressor Proteins/metabolism , tau Proteins/metabolism
6.
J Neurochem ; 137(1): 12-25, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26756400

ABSTRACT

Abnormal tau accumulations were observed and documented in post-mortem brains of patients affected by Alzheimer's disease (AD) long before the identification of mutations in the Microtubule-associated protein tau (MAPT) gene, encoding the tau protein, in a different neurodegenerative disease called Frontotemporal dementia and Parkinsonism linked to chromosome 17 (FTDP-17). The discovery of mutations in the MAPT gene associated with FTDP-17 highlighted that dysfunctions in tau alone are sufficient to cause neurodegeneration. Invertebrate models have been diligently utilized in investigating tauopathies, contributing to the understanding of cellular and molecular pathways involved in disease etiology. An important discovery came with the demonstration that over-expression of human tau in Drosophila leads to premature mortality and neuronal dysfunction including neurodegeneration, recapitulating some key neuropathological features of the human disease. The simplicity of handling invertebrate models combined with the availability of a diverse range of experimental resources make these models, in particular Drosophila a powerful invertebrate screening tool. Consequently, several large-scale screens have been performed using Drosophila, to identify modifiers of tau toxicity. The screens have revealed not only common cellular and molecular pathways, but in some instances the same modifier has been independently identified in two or more screens suggesting a possible role for these modifiers in regulating tau toxicity. The purpose of this review is to discuss the genetic modifier screens on tauopathies performed in Drosophila and C. elegans models, and to highlight the common cellular and molecular pathways that have emerged from these studies. Here, we summarize results of tau toxicity screens providing mechanistic insights into pathological alterations in tauopathies. Key pathways or modifiers that have been identified are associated with a broad range of processes including, but not limited to, phosphorylation, cytoskeleton organization, axonal transport, regulation of cellular proteostasis, transcription, RNA metabolism, cell cycle regulation, and apoptosis. We discuss the utility and application of invertebrate models in elucidating the cellular and molecular functions of novel and uncharacterized disease modifiers identified in large-scale screens as well as for investigating the function of genes identified as risk factors in genome-wide association studies from human patients in the post-genomic era. In this review, we combined and summarized several large-scale modifier screens performed in invertebrate models to identify modifiers of tau toxicity. A summary of the screens show that diverse cellular processes are implicated in the modification of tau toxicity. Kinases and phosphatases are the most predominant class of modifiers followed by components required for cellular proteostasis and axonal transport and cytoskeleton elements.


Subject(s)
Invertebrates/metabolism , Tauopathies/metabolism , Animals , Animals, Genetically Modified , Apoptosis , Axonal Transport , Caenorhabditis elegans/metabolism , Cell Cycle , Cytoskeleton/metabolism , Cytoskeleton/ultrastructure , Disease Models, Animal , Drosophila melanogaster/metabolism , Gene Expression Regulation , Humans , Longevity/genetics , Metabolic Networks and Pathways , Mice , Mice, Knockout , Mutation , Nerve Degeneration/genetics , Phosphorylation , Protein Processing, Post-Translational , Recombinant Fusion Proteins/metabolism , Recombinant Fusion Proteins/toxicity , Zebrafish , tau Proteins/genetics , tau Proteins/metabolism , tau Proteins/toxicity
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