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1.
Nat Genet ; 56(3): 408-419, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38424460

ABSTRACT

Humans display remarkable interindividual variation in their immune response to identical challenges. Yet, our understanding of the genetic and epigenetic factors contributing to such variation remains limited. Here we performed in-depth genetic, epigenetic and transcriptional profiling on primary macrophages derived from individuals of European and African ancestry before and after infection with influenza A virus. We show that baseline epigenetic profiles are strongly predictive of the transcriptional response to influenza A virus across individuals. Quantitative trait locus (QTL) mapping revealed highly coordinated genetic effects on gene regulation, with many cis-acting genetic variants impacting concomitantly gene expression and multiple epigenetic marks. These data reveal that ancestry-associated differences in the epigenetic landscape can be genetically controlled, even more than gene expression. Lastly, among QTL variants that colocalized with immune-disease loci, only 7% were gene expression QTL, while the remaining genetic variants impact epigenetic marks, stressing the importance of considering molecular phenotypes beyond gene expression in disease-focused studies.


Subject(s)
Influenza, Human , Humans , Influenza, Human/genetics , Individuality , Quantitative Trait Loci/genetics , Chromosome Mapping , Epigenesis, Genetic
2.
Cell Genom ; 3(5): 100294, 2023 May 10.
Article in English | MEDLINE | ID: mdl-37228750

ABSTRACT

Genetic variants, including mobile element insertions (MEIs), are known to impact the epigenome. We hypothesized that genome graphs, which encapsulate genetic diversity, could reveal missing epigenomic signals. To test this, we sequenced the epigenome of monocyte-derived macrophages from 35 ancestrally diverse individuals before and after influenza infection, allowing us to investigate the role of MEIs in immunity. We characterized genetic variants and MEIs using linked reads and built a genome graph. Mapping epigenetic data revealed 2.3%-3% novel peaks for H3K4me1, H3K27ac chromatin immunoprecipitation sequencing (ChIP-seq), and ATAC-seq. Additionally, the use of a genome graph modified some quantitative trait loci estimates and revealed 375 polymorphic MEIs in an active epigenomic state. Among these is an AluYh3 polymorphism whose chromatin state changed after infection and was associated with the expression of TRIM25, a gene that restricts influenza RNA synthesis. Our results demonstrate that graph genomes can reveal regulatory regions that would have been overlooked by other approaches.

3.
Cell Genom ; 3(5): 100292, 2023 May 10.
Article in English | MEDLINE | ID: mdl-37228757

ABSTRACT

Influenza A virus (IAV) infections are frequent every year and result in a range of disease severity. Here, we wanted to explore the potential contribution of transposable elements (TEs) to the variable human immune response. Transcriptome profiling in monocyte-derived macrophages from 39 individuals following IAV infection revealed significant inter-individual variation in viral load post-infection. Using transposase-accessible chromatin using sequencing (ATAC-seq), we identified a set of TE families with either enhanced or reduced accessibility upon infection. Of the enhanced families, 15 showed high variability between individuals and had distinct epigenetic profiles. Motif analysis showed an association with known immune regulators (e.g., BATFs, FOSs/JUNs, IRFs, STATs, NFkBs, NFYs, and RELs) in stably enriched families and with other factors in variable families, including KRAB-ZNFs. We showed that TEs and host factors regulating TEs were predictive of viral load post-infection. Our findings shed light on the role TEs and KRAB-ZNFs may play in inter-individual variation in immunity.

4.
Nat Metab ; 2(1): 97-109, 2020 01.
Article in English | MEDLINE | ID: mdl-32066997

ABSTRACT

The complex relationship between metabolic disease risk and body fat distribution in humans involves cellular characteristics which are specific to body fat compartments. Here we show depot-specific differences in the stromal vascual fraction of visceral and subcutaneous adipose tissue by performing single-cell RNA sequencing of tissue specimen from obese individuals. We characterize multiple immune cells, endothelial cells, fibroblasts, adipose and hematopoietic stem cell progenitors. Subpopulations of adipose-resident immune cells are metabolically active and associated with metabolic disease status and those include a population of potential dysfunctional CD8+ T cells expressing metallothioneins. We identify multiple types of adipocyte progenitors that are common across depots, including a subtype enriched in individuals with type 2 diabetes. Depot-specific analysis reveals a class of adipocyte progenitors unique to visceral adipose tissue, which shares common features with beige preadipocytes. Our human single-cell transcriptome atlas across fat depots provides a resource to dissect functional genomics of metabolic disease.


Subject(s)
Adipose Tissue/metabolism , Metabolic Diseases/metabolism , Single-Cell Analysis/methods , Adipocytes/metabolism , Adipose Tissue/cytology , Adult , Body Fat Distribution , Female , Gene Expression Profiling , High-Throughput Nucleotide Sequencing , Humans , Male , Metabolic Diseases/pathology , Middle Aged , Obesity/metabolism
5.
Mol Cell Biol ; 26(4): 1510-7, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16449660

ABSTRACT

Nucleokinesis has recently been suggested as a critical regulator of neuronal migration. Here we show that Disabled 1 (Dab1), which is required for neuronal positioning in mammals, regulates the nuclear position of postmitotic neurons in a phosphorylation-site dependent manner. Dab1 expression in the Drosophila visual system partially rescues nuclear position defects caused by a mutation in the Dynactin subunit Glued. Furthermore, we observed that a loss-of-function allele of amyloid precursor protein (APP)-like, a kinesin cargo receptor, enhanced the severity of a Dab1 overexpression phenotype characterized by misplaced nuclei in the adult retina. In mammalian neurons, overexpression of APP reduced the ability of Reelin to induce Dab1 tyrosine phosphorylation, suggesting an antagonistic relationship between APP family members and Dab1 function. This is the first evidence that signaling which regulates Dab1 tyrosine phosphorylation determines nuclear positioning through Dab1-mediated influences on microtubule motor proteins in a subset of neurons.


Subject(s)
Drosophila/growth & development , Drosophila/genetics , Eye/growth & development , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Animals , Animals, Genetically Modified , Cell Adhesion Molecules, Neuronal/metabolism , Cell Movement , Drosophila/cytology , Drosophila/metabolism , Drosophila Proteins/deficiency , Drosophila Proteins/genetics , Extracellular Matrix Proteins/metabolism , Eye/cytology , Eye/metabolism , Female , Genes, Insect , Membrane Proteins/deficiency , Membrane Proteins/genetics , Mice , Microscopy, Electron, Scanning , Models, Biological , Mutation , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/deficiency , Neurons/cytology , Neurons/metabolism , Phenotype , Phosphorylation , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Reelin Protein , Serine Endopeptidases/metabolism , Tyrosine/chemistry
6.
Nat Genet ; 51(2): 258-266, 2019 02.
Article in English | MEDLINE | ID: mdl-30598549

ABSTRACT

Osteoporosis is a common aging-related disease diagnosed primarily using bone mineral density (BMD). We assessed genetic determinants of BMD as estimated by heel quantitative ultrasound in 426,824 individuals, identifying 518 genome-wide significant loci (301 novel), explaining 20% of its variance. We identified 13 bone fracture loci, all associated with estimated BMD (eBMD), in ~1.2 million individuals. We then identified target genes enriched for genes known to influence bone density and strength (maximum odds ratio (OR) = 58, P = 1 × 10-75) from cell-specific features, including chromatin conformation and accessible chromatin sites. We next performed rapid-throughput skeletal phenotyping of 126 knockout mice with disruptions in predicted target genes and found an increased abnormal skeletal phenotype frequency compared to 526 unselected lines (P < 0.0001). In-depth analysis of one gene, DAAM2, showed a disproportionate decrease in bone strength relative to mineralization. This genetic atlas provides evidence linking associated SNPs to causal genes, offers new insight into osteoporosis pathophysiology, and highlights opportunities for drug development.


Subject(s)
Bone Density/genetics , Genetic Predisposition to Disease/genetics , Osteoporosis/genetics , Adult , Aged , Animals , Female , Fractures, Bone/genetics , Genome-Wide Association Study/methods , Humans , Male , Mice , Mice, Knockout , Middle Aged , Phenotype , Polymorphism, Single Nucleotide/genetics
8.
Mol Oncol ; 12(6): 775-787, 2018 06.
Article in English | MEDLINE | ID: mdl-29316219

ABSTRACT

Glioblastoma (GBM) is the most common and deadly malignant brain cancer of glial cell origin, with a median patient survival of less than 20 months. Transcription factors FOXG1 and TLE1 promote GBM propagation by supporting maintenance of brain tumour-initiating cells (BTICs) with stem-like properties. Here, we characterize FOXG1 and TLE1 target genes in GBM patient-derived BTICs using ChIP-Seq and RNA-Seq approaches. These studies identify 150 direct FOXG1 targets, several of which are also TLE1 targets, involved in cell proliferation, differentiation, survival, chemotaxis and angiogenesis. Negative regulators of NOTCH signalling, including CHAC1, are among the transcriptional repression targets of FOXG1:TLE1 complexes, suggesting a crosstalk between FOXG1:TLE1 and NOTCH-mediated pathways in GBM. These results provide previously unavailable insight into the transcriptional programs underlying the tumour-promoting functions of FOXG1:TLE1 in GBM.


Subject(s)
Forkhead Transcription Factors/genetics , Gene Regulatory Networks , Glioblastoma/genetics , Glioblastoma/pathology , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Nerve Tissue Proteins/genetics , Repressor Proteins/genetics , Binding Sites , Brain Neoplasms/genetics , Brain Neoplasms/pathology , Cell Line, Tumor , Co-Repressor Proteins , Forkhead Transcription Factors/metabolism , Gene Expression Regulation, Neoplastic , Genome, Human , Hepatocyte Nuclear Factor 3-alpha/metabolism , Humans , Nerve Tissue Proteins/metabolism , Protein Binding , RNA, Messenger/genetics , RNA, Messenger/metabolism , Repressor Proteins/metabolism , Reproducibility of Results , gamma-Glutamylcyclotransferase/metabolism
9.
Mol Cell Biol ; 23(20): 7210-21, 2003 Oct.
Article in English | MEDLINE | ID: mdl-14517291

ABSTRACT

The tyrosine phosphorylation sites of the Disabled 1 (Dab1) docking protein are essential for the transmission of the Reelin signal, which regulates neuronal placement. Here we identify Nck beta as a phosphorylation-dependent, Dab1-interacting protein. The SH2 domain of Nck beta but not Nck alpha binds Dab1 phosphorylated on the Reelin-regulated site, Y220, or on Y232. Nck beta is coexpressed with Dab1 in the developing brain and in cultured neurons, where Reelin stimulation leads to the redistribution of Nck beta from the cell soma into neuronal processes. We found that tyrosine-phosphorylated Dab1 in synergy with Nck beta disrupts the actin cytoskeleton in transfected cells. In Drosophila melanogaster, exogenous expression of mouse Dab1 causes tyrosine phosphorylation site-dependent morphological changes in the compound eye. This phenotype is enhanced by overexpression of the Drosophila Nck protein Dock, suggesting a conserved interaction between the Disabled and Nck family members. We suggest a model in which Dab1 phosphorylation leads to the recruitment of Nck beta to the membrane, where it acts to remodel the actin cytoskeleton.


Subject(s)
Carrier Proteins/metabolism , Cell Adhesion Molecules, Neuronal/metabolism , Drosophila Proteins , Extracellular Matrix Proteins/metabolism , Nerve Tissue Proteins/physiology , Neurons/metabolism , Tyrosine/metabolism , Actins/metabolism , Animals , Cell Line , Cytoskeleton/metabolism , Drosophila melanogaster/metabolism , Humans , Immunohistochemistry , Mice , Microscopy, Electron, Scanning , Phosphorylation , Plasmids/metabolism , Precipitin Tests , Protein Binding , Protein Structure, Tertiary , Rats , Reelin Protein , Serine Endopeptidases , Time Factors , Two-Hybrid System Techniques , src Homology Domains
10.
Mol Cell Biol ; 24(3): 1378-86, 2004 Feb.
Article in English | MEDLINE | ID: mdl-14729980

ABSTRACT

The Reelin signaling cascade plays a crucial role in the correct positioning of neurons during embryonic brain development. Reelin binding to apolipoprotein E receptor 2 (ApoER2) and very-low-density-lipoprotein receptor (VLDLR) leads to phosphorylation of disabled 1 (Dab1), an adaptor protein which associates with the intracellular domains of both receptors. Coreceptors for Reelin have been postulated to be necessary for Dab1 phosphorylation. We show that bivalent agents specifically binding to ApoER2 or VLDLR are sufficient to mimic the Reelin signal. These agents induce Dab1 phosphorylation, activate members of the Src family of nonreceptor tyrosine kinases, modulate protein kinase B/Akt phosphorylation, and increase long-term potentiation in hippocampal slices. Induced dimerization of Dab1 in HEK293 cells leads to its phosphorylation even in the absence of Reelin receptors. The mechanism for and the sites of these phosphorylations are identical to those effected by Reelin in primary neurons. These results suggest that binding of Reelin, which exists as a homodimer in vivo, to ApoER2 and VLDLR induces clustering of ApoER2 and VLDLR. As a consequence, Dab1 becomes dimerized or oligomerized on the cytosolic side of the plasma membrane, constituting the active substrate for the kinase; this process seems to be sufficient to transmit the signal and does not appear to require any coreceptor.


Subject(s)
Cell Adhesion Molecules, Neuronal/metabolism , Extracellular Matrix Proteins/metabolism , Receptors, Cell Surface/metabolism , Signal Transduction/physiology , Animals , Dimerization , Nerve Tissue Proteins/metabolism , Rats , Reelin Protein , Serine Endopeptidases
11.
Nat Genet ; 46(1): 39-44, 2014 Jan.
Article in English | MEDLINE | ID: mdl-24316981

ABSTRACT

Embryonal tumors with multilayered rosettes (ETMRs) are rare, deadly pediatric brain tumors characterized by high-level amplification of the microRNA cluster C19MC. We performed integrated genetic and epigenetic analyses of 12 ETMR samples and identified, in all cases, C19MC fusions to TTYH1 driving expression of the microRNAs. ETMR tumors, cell lines and xenografts showed a specific DNA methylation pattern distinct from those of other tumors and normal tissues. We detected extreme overexpression of a previously uncharacterized isoform of DNMT3B originating at an alternative promoter that is active only in the first weeks of neural tube development. Transcriptional and immunohistochemical analyses suggest that C19MC-dependent DNMT3B deregulation is mediated by RBL2, a known repressor of DNMT3B. Transfection with individual C19MC microRNAs resulted in DNMT3B upregulation and RBL2 downregulation in cultured cells. Our data suggest a potential oncogenic re-engagement of an early developmental program in ETMR via epigenetic alteration mediated by an embryonic, brain-specific DNMT3B isoform.


Subject(s)
Brain Neoplasms/genetics , Chromosomes, Human, Pair 19 , DNA (Cytosine-5-)-Methyltransferases/genetics , Membrane Proteins/genetics , MicroRNAs/genetics , Neoplasms, Germ Cell and Embryonal/genetics , Child, Preschool , DNA (Cytosine-5-)-Methyltransferases/metabolism , DNA Methylation , Gene Expression Regulation, Neoplastic , Gene Fusion , Humans , Male , Protein Isoforms , Retinoblastoma-Like Protein p130/genetics , Xenograft Model Antitumor Assays , DNA Methyltransferase 3B
12.
J Cell Sci ; 121(11): 1869-75, 2008 Jun 01.
Article in English | MEDLINE | ID: mdl-18477607

ABSTRACT

The reelin signaling pathway regulates nervous system function after birth, in addition to its role in regulating neuronal positioning during embryogenesis. The receptor-dependent, reelin-induced tyrosine phosphorylation of the Dab1 docking protein is an established prerequisite for biological responses to this ligand. Here we show that the inactivation of a conditional Dab1 allele reduces process complexity in correctly positioned neurons in the CA1 region of the mouse hippocampus after birth. Reelin stimulation of cultured hippocampal neurons enhances dendritogenesis by approximately twofold and in a manner dependent on Src family kinases. This enhancement is blocked by reducing expression of Crk family proteins, adaptor molecules that interact with Dab1 in a tyrosine phosphorylation-dependent manner. Retrovirally expressed inhibitory RNAs used to reduce Crk and CrkL expression did not block BDNF-enhanced dendritogenesis or influence axonogenesis. Together, this demonstrates that the Crk family proteins are important downstream components of the reelin signaling pathway in the regulation of postnatal hippocampal dendritogenesis.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Cell Adhesion Molecules, Neuronal/physiology , Cell Differentiation/physiology , Dendrites/metabolism , Extracellular Matrix Proteins/physiology , Hippocampus/metabolism , Nerve Tissue Proteins/physiology , Nuclear Proteins/metabolism , Proto-Oncogene Proteins c-crk/metabolism , Serine Endopeptidases/physiology , Adaptor Proteins, Signal Transducing/genetics , Animals , Cell Shape/physiology , Cells, Cultured , Dendrites/ultrastructure , Down-Regulation/genetics , Hippocampus/embryology , Hippocampus/growth & development , Mice , Mice, Knockout , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/metabolism , Nuclear Proteins/genetics , Phosphorylation , Proto-Oncogene Proteins c-crk/genetics , Pyramidal Cells/cytology , Pyramidal Cells/metabolism , RNA, Small Interfering , Reelin Protein , Signal Transduction/physiology , Silver Staining , src-Family Kinases/genetics , src-Family Kinases/metabolism
13.
Mol Cell Neurosci ; 37(1): 178-86, 2008 Jan.
Article in English | MEDLINE | ID: mdl-18029196

ABSTRACT

The Dab1 docking protein is required for the proper organization of brain laminae and for a signal transduction pathway initiated by Reelin binding to the ApoER2 and VLDLR receptors on the cell surface of neurons. Dab1 physically interacts with APP; however, it is not known whether the APP gene influences Dab1 function. Here we demonstrate a genetic interaction between Dab1 and APP. Dab1-hypomorphic animals have neuronal ectopias in the neocortex and reduced cerebellar volume, possibly a consequence of Purkinje cell misplacement. These phenotypes are exacerbated in transgenic animals overexpressing a mutant form of APP, APP(swe), which is characterized by increased processing at the beta-secretase site. The Dab1-hypomorphic phenotype is improved in the cerebellum of animals that are deficient for APP. Together this suggests that APP expression constrains the consequences of Dab1 activity during brain development.


Subject(s)
Amyloid beta-Protein Precursor/genetics , Brain/anatomy & histology , Brain/growth & development , Phosphoproteins/genetics , Adaptor Proteins, Signal Transducing , Animals , Animals, Newborn , Gene Expression Regulation , Mice , Mice, Transgenic , Protein Binding/genetics , Reelin Protein
14.
J Cell Sci ; 117(Pt 19): 4527-36, 2004 Sep 01.
Article in English | MEDLINE | ID: mdl-15316068

ABSTRACT

Reelin-induced Dab1 tyrosine phosphorylation has been implicated in the regulation of neuronal positioning during brain development. The downstream consequences of Dab1 tyrosine phosphorylation are not fully understood, however. Here we identify CrkII, CrkL and Dock1 in complexes bound to tyrosine-phosphorylated Dab1, through mass spectrometry. The CrkII-Dab1 interaction requires tyrosine phosphorylation of Dab1 at residues 220 or 232 and is promoted by Reelin treatment of embryonic forebrain neurons. Unlike other CrkII binding proteins, such as paxillin and p130Cas, expression of Dab1 interfered with CrkII-dependent cell migration of Nara Bladder Tumor II (NBT-II) cells, in a tyrosine phosphorylation-site dependent manner. Overexpression of CrkIIGFP rescued the migration of these cells, suggesting that Dab1 makes Crk a limiting factor for migration. The Dock1-Dab1 association is indirect and requires CrkII. In organisms such as Drosophila melanogaster and Caenorhabditis elegans, signaling complexes, which contain Crk and Dock1 family members are conserved and act through Rac. We show that a rough-eye phenotype in Drosophila caused by exogenous expression of tyrosine-phosphorylated mouse Dab1RFP is partially rescued by a loss-of-function mutation in myoblast city, a Dock1-like gene in Drosophila. We propose a model that tyrosine-phosphorylated Dab1 engages the conserved Crk-Dock1-Rac signaling cassette, but when bound to Dab1 this signaling complex does not support migration.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Cell Adhesion Molecules, Neuronal/pharmacology , Cell Movement/physiology , Extracellular Matrix Proteins/pharmacology , Nerve Tissue Proteins/metabolism , Nuclear Proteins/metabolism , Proto-Oncogene Proteins/metabolism , Amino Acid Sequence , Animals , Cell Movement/drug effects , Cells, Cultured , Collagen/metabolism , Cytoskeletal Proteins/genetics , Cytoskeletal Proteins/metabolism , Cytoskeleton/metabolism , Drosophila/genetics , Drosophila/metabolism , Drosophila/ultrastructure , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Embryo, Mammalian/metabolism , Embryo, Nonmammalian , Mass Spectrometry , Mice , Microscopy, Electron, Scanning , Molecular Sequence Data , Neurons/metabolism , Paxillin , Phosphoproteins/metabolism , Phosphorylation , Protein Binding/drug effects , Proto-Oncogene Proteins c-crk , Reelin Protein , Serine Endopeptidases , rac GTP-Binding Proteins
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