Manav Kapoor, United States of America
Icahn School of Medicine at Mount Sinai NeuroscienceAuthor Of 3 Presentations
GENETIC STUDIES IMPLICATE MICROGLIAL FUNCTION IN ALZHEIMER’S DISEASE RISK
Abstract
Abstract Body
Genome-wide association studies (GWAS) have identified more than 40 loci associated with Alzheimer’s disease (AD), but the causal variants, regulatory elements, genes and pathways remain largely unknown, impeding a mechanistic understanding of AD pathogenesis. Previously, we showed that AD risk alleles are enriched in myeloid-specific epigenomic annotations. Here, we show that they are specifically enriched in active enhancers of monocytes, macrophages and microglia. We integrated AD GWAS with myeloid epigenomic and transcriptomic datasets using novel analytical approaches to link myeloid enhancer activity to target gene expression regulation and AD risk modification. We identify AD risk enhancers and nominate candidate causal genes among their likely targets (including AP4E1, AP4M1, APBB3, BIN1, MS4A4A, MS4A6A, PILRA, RABEP1, SPI1, TP53INP1, and ZYX) in twenty loci. Fine-mapping of these enhancers nominates candidate functional variants that likely modify AD risk by regulating gene expression in myeloid cells. In the MS4A locus we identified a single candidate functional variant and validated it in human induced pluripotent stem cell (hiPSC)-derived microglia and brain. Taken together, this study integrates AD GWAS with multiple myeloid genomic datasets to investigate the mechanisms of AD risk alleles and nominates candidate functional variants, regulatory elements and genes that likely modulate disease susceptibility.
INTEGRATION OF ALZHEIMER'S DISEASE GENETICS, MYELOID CELL GENOMICS AND GENE REGULATORY NETWORKS REVEALS NOVEL DISEASE RISK MECHANISMS
Abstract
Aims
We aim to identify myeloid regulatory elements that are enriched in Alzheimer’s disease (AD) risk alleles, identify their target genes and nominate candidate causal variants in disease risk loci. We also want to study the likely downstream effects of AD risk genes and functionally validate these findings in microglia.
Methods
Integration of myeloid epigenomic, chromatin interactions and quantitative trait loci (QTL) datasets as well as a Mendelian Randomization framework were used to link AD enhancers to their target genes. Fine-mapping analyses were used to nominate candidate causal variants and generate the mechanism of action hypotheses. Gene regulatory networks were used to study the regulons of AD risk genes. Knockdown and overexpression studies were utilized to validate these findings.
Results
We found that myeloid active enhancers are enriched in AD risk alleles. We linked these enhancers to their likely target genes, nominating AD risk genes in twenty loci. Fine-mapping of these enhancers nominates candidate functional variants in these loci. In the MS4A locus we identified a candidate functional variant and validated it in microglia and the brain. We highlight the coalescence of candidate causal genes in the myeloid endolysosomal system. We constructed myeloid single-cell gene regulatory networks and found that the predicted targets of SPI1, an AD risk gene, were enriched in the endolysosomal compartment. We validate these findings in Spi1 knockdown and overexpression experiments in microglia.
Conclusions
This study explores the links between AD risk variants, myeloid enhancer activity, gene expression and subsequent network-level dysregulations that likely contribute to AD risk modification.
CHOLESTEROL AND MATRISOME PATHWAYS DYSREGULATED IN HUMAN APOE Ε4 GLIA
Abstract
Aims
The association of Apolipoprotein E (APOE) ɛ4 with Alzheimer's disease risk is well-established however, the impact of APOE ɛ4 on human brain cell function remains unclear. We hypothesized that human APOE ε4/ε4 genotype contributes to disease risk through cell autonomous and non-cell autonomous mechanisms in a human specific manner.
Methods
Global transcriptomic analysis (pathway, network and cell type deconvolution) were performed on brain cell types, astrocytes, microglia, mixed cortical cultures (neurons and astrocytes) and brain microvascular endothelial cells, derived from human induced pluripotent stem cells (population and isogenic cells), post-mortem brains and APOE targeted replacement mice. For in vitro mechanistic study, filipin staining, Gas Chromatography/Mass Spectrometry (GC/MS), live cell imaging, immunocytochemistry, and Luminex multiplex immunoassays were utilized for lipid trafficking, the measurement of intracellular cholesterol level, localization, and cytokine/chemokine secretion.
Results
Global transcriptomic analyses identified human specific APOE ε4-driven lipid metabolic dysregulation in astrocytes and microglia. Decoupled cholesterol metabolism in APOE ε4 glia is due to lysosomal cholesterol sequestration; despite increased intracellular cholesterol leads to elevated de novo synthesis and decreased efflux. The most significant upregulated signal in post-mortem APOE ε4 AD brains is matrisome associated with chemotaxis, glial activation and lipid biosynthesis, which derives from astrocytes, paralleled pathways uncovered in astrocytes when co-cultured with neurons. Further, APOE ε4 astrocytes show enhanced chemokine/proinflammatory factors production when communicating with neurons.
Conclusions
Human APOE ε4 initiates human glia-specific cell autonomous and non-cell autonomous dysregulation that may increase AD risk.