- Clinical Pathology
Honors & Awards
Medical Scientist Training Program (MSTP), NIH (2006-2014)
Predoctoral Fellowship, American Heart Association (2012)
David M. Kipnis Dissertation Award, Washington University School of Medicine (2013)
Irvington Postdoctoral Fellowship, Cancer Research Institute (2016)
Bridge Scholar, Parker Institute for Cancer Immunotherapy (2017)
Career Award for Medical Scientists, Burroughs Wellcome Fund (2018)
Clinical Scientist Career Development Award (K08), National Cancer Institute (2018)
Innovative Technology Award, Bill and Melinda Gates Foundation (2018)
Michelson Prize for Human Immunology and Vaccine Research, Michelson Research Foundation (2018)
PhD Training:Washington University School Of Medicine RegistrarMO
Visiting Scholar, King's College London, Immunology (2005)
B.A., University of Illinois, Urbana-Champaign, Philosophy (2006)
B.S., University of Illinois, Urbana-Champaign, Molecular Biology (2006)
M.D., Washington University in St. Louis, Medicine (2014)
Ph.D., Washington University in St. Louis, Immunology (2014)
Residency, Stanford Hospital and Clinics, Clinical Pathology (2017)
Current Research and Scholarly Interests
Epigenomics, Cancer immunology, Single-cell genomics
Transcript-indexed ATAC-seq for precision immune profiling.
T cells create vast amounts of diversity in the genes that encode their T cell receptors (TCRs), which enables individual clones to recognize specific peptide-major histocompatibility complex (MHC) ligands. Here we combined sequencing of the TCR-encoding genes with assay for transposase-accessible chromatin with sequencing (ATAC-seq) analysis at the single-cell level to provide information on the TCR specificity and epigenomic state of individual T cells. By using this approach, termed transcript-indexed ATAC-seq (T-ATAC-seq), we identified epigenomic signatures in immortalized leukemic T cells, primary human T cells from healthy volunteers and primary leukemic T cells from patient samples. In peripheral blood CD4+ T cells from healthy individuals, we identified cis and trans regulators of naive and memory T cell states and found substantial heterogeneity in surface-marker-defined T cell populations. In patients with a leukemic form of cutaneous T cell lymphoma, T-ATAC-seq enabled identification of leukemic and nonleukemic regulatory pathways in T cells from the same individual by allowing separation of the signals that arose from the malignant clone from the background T cell noise. Thus, T-ATAC-seq is a new tool that enables analysis of epigenomic landscapes in clonal T cells and should be valuable for studies of T cell malignancy, immunity and immunotherapy.
View details for DOI 10.1038/s41591-018-0008-8
View details for PubMedID 29686426
Enhancer connectome in primary human cells identifies target genes of disease-associated DNA elements.
The challenge of linking intergenic mutations to target genes has limited molecular understanding of human diseases. Here we show that H3K27ac HiChIP generates high-resolution contact maps of active enhancers and target genes in rare primary human T cell subtypes and coronary artery smooth muscle cells. Differentiation of naive T cells into T helper 17 cells or regulatory T cells creates subtype-specific enhancer-promoter interactions, specifically at regions of shared DNA accessibility. These data provide a principled means of assigning molecular functions to autoimmune and cardiovascular disease risk variants, linking hundreds of noncoding variants to putative gene targets. Target genes identified with HiChIP are further supported by CRISPR interference and activation at linked enhancers, by the presence of expression quantitative trait loci, and by allele-specific enhancer loops in patient-derived primary cells. The majority of disease-associated enhancers contact genes beyond the nearest gene in the linear genome, leading to a fourfold increase in the number of potential target genes for autoimmune and cardiovascular diseases.
View details for DOI 10.1038/ng.3963
View details for PubMedID 28945252
Gene regulation in the immune system by long noncoding RNAs.
2017; 18 (9): 962–72
Long noncoding RNAs (lncRNAs) are emerging as critical regulators of gene expression in the immune system. Studies have shown that lncRNAs are expressed in a highly lineage-specific manner and control the differentiation and function of innate and adaptive cell types. In this Review, we focus on mechanisms used by lncRNAs to regulate genes encoding products involved in the immune response, including direct interactions with chromatin, RNA and proteins. In addition, we address new areas of lncRNA biology, such as the functions of enhancer RNAs, circular RNAs and chemical modifications to RNA in cellular processes. We emphasize critical gaps in knowledge and future prospects for the roles of lncRNAs in the immune system and autoimmune disease.
View details for DOI 10.1038/ni.3771
View details for PubMedID 28829444
- Long Noncoding RNA in Hematopoiesis and Immunity IMMUNITY 2015; 42 (5): 792-804
Runx1 and Cbf beta regulate the development of Flt3(+) dendritic cell progenitors and restrict myeloproliferative disorder
2014; 123 (19): 2968-2977
Runx1 and Cbfβ are critical for the establishment of definitive hematopoiesis and are implicated in leukemic transformation. Despite the absolute requirements for these factors in the development of hematopoietic stem cells and lymphocytes, their roles in the development of bone marrow progenitor subsets have not been defined. Here, we demonstrate that Cbfβ is essential for the development of Flt3(+) macrophage-dendritic cell (DC) progenitors in the bone marrow and all DC subsets in the periphery. Besides the loss of DC progenitors, pan-hematopoietic Cbfb-deficient mice also lack CD105(+) erythroid progenitors, leading to severe anemia at 3 to 4 months of age. Instead, Cbfb deficiency results in aberrant progenitor differentiation toward granulocyte-macrophage progenitors (GMPs), resulting in a myeloproliferative phenotype with accumulation of GMPs in the periphery and cellular infiltration of the liver. Expression of the transcription factor Irf8 is severely reduced in Cbfb-deficient progenitors, and overexpression of Irf8 restors DC differentiation. These results demonstrate that Runx proteins and Cbfβ restrict granulocyte lineage commitment to facilitate multilineage hematopoietic differentiation and thus identify their novel tumor suppressor function in myeloid leukemia.
View details for DOI 10.1182/blood-2013-11-539643
View details for Web of Science ID 000335897900015
View details for PubMedID 24677539
View details for PubMedCentralID PMC4014839
L-Myc expression by dendritic cells is required for optimal T-cell priming
2014; 507 (7491): 243-?
The transcription factors c-Myc and N-Myc--encoded by Myc and Mycn, respectively--regulate cellular growth and are required for embryonic development. A third paralogue, Mycl1, is dispensable for normal embryonic development but its biological function has remained unclear. To examine the in vivo function of Mycl1 in mice, we generated an inactivating Mycl1(gfp) allele that also reports Mycl1 expression. We find that Mycl1 is selectively expressed in dendritic cells (DCs) of the immune system and controlled by IRF8, and that during DC development, Mycl1 expression is initiated in the common DC progenitor concurrent with reduction in c-Myc expression. Mature DCs lack expression of c-Myc and N-Myc but maintain L-Myc expression even in the presence of inflammatory signals such as granulocyte-macrophage colony-stimulating factor. All DC subsets develop in Mycl1-deficient mice, but some subsets such as migratory CD103(+) conventional DCs in the lung and liver are greatly reduced at steady state. Importantly, loss of L-Myc by DCs causes a significant decrease in in vivo T-cell priming during infection by Listeria monocytogenes and vesicular stomatitis virus. The replacement of c-Myc by L-Myc in immature DCs may provide for Myc transcriptional activity in the setting of inflammation that is required for optimal T-cell priming.
View details for DOI 10.1038/nature12967
View details for Web of Science ID 000332651800041
View details for PubMedID 24509714
View details for PubMedCentralID PMC3954917
- Notch2-dependent classical dendritic cells orchestrate intestinal immunity to attaching-and-effacing bacterial pathogens NATURE IMMUNOLOGY 2013; 14 (9): 937-?
Re(de)fining the dendritic cell lineage
2012; 13 (12): 1145-1154
Dendritic cells (DCs) are essential mediators of innate and adaptive immune responses. Study of these critical cells has been complicated by their similarity to other hematopoietic lineages, particularly monocytes and macrophages. Progress has been made in three critical areas of DC biology: the characterization of lineage-restricted progenitors in the bone marrow, the identification of cytokines and transcription factors required during differentiation, and the development of genetic tools for the visualization and depletion of DCs in vivo. Collectively, these advances have clarified the nature of the DC lineage and have provided novel insights into their function during health and disease.
View details for DOI 10.1038/ni.2467
View details for Web of Science ID 000311217900007
View details for PubMedID 23160217
View details for PubMedCentralID PMC3644874
Zbtb46 expression distinguishes classical dendritic cells and their committed progenitors from other immune lineages
JOURNAL OF EXPERIMENTAL MEDICINE
2012; 209 (6): 1135-1152
Distinguishing dendritic cells (DCs) from other cells of the mononuclear phagocyte system is complicated by the shared expression of cell surface markers such as CD11c. In this study, we identified Zbtb46 (BTBD4) as a transcription factor selectively expressed by classical DCs (cDCs) and their committed progenitors but not by plasmacytoid DCs (pDCs), monocytes, macrophages, or other lymphoid or myeloid lineages. Using homologous recombination, we replaced the first coding exon of Zbtb46 with GFP to inactivate the locus while allowing detection of Zbtb46 expression. GFP expression in Zbtb46(gfp/+) mice recapitulated the cDC-specific expression of the native locus, being restricted to cDC precursors (pre-cDCs) and lymphoid organ- and tissue-resident cDCs. GFP(+) pre-cDCs had restricted developmental potential, generating cDCs but not pDCs, monocytes, or macrophages. Outside the immune system, Zbtb46 was expressed in committed erythroid progenitors and endothelial cell populations. Zbtb46 overexpression in bone marrow progenitor cells inhibited granulocyte potential and promoted cDC development, and although cDCs developed in Zbtb46(gfp/gfp) (Zbtb46 deficient) mice, they maintained expression of granulocyte colony-stimulating factor and leukemia inhibitory factor receptors, which are normally down-regulated in cDCs. Thus, Zbtb46 may help enforce cDC identity by restricting responsiveness to non-DC growth factors and may serve as a useful marker to identify rare cDC progenitors and distinguish between cDCs and other mononuclear phagocyte lineages.
View details for DOI 10.1084/jem.20120030
View details for Web of Science ID 000304907800009
View details for PubMedID 22615127
View details for PubMedCentralID PMC3371733
Enhancer connectome nominates target genes of inherited risk variants from inflammatory skin disorders.
The Journal of investigative dermatology
The vast majority of polymorphisms for human dermatologic diseases fall in non-coding DNA regions, leading to difficulty interpreting their functional significance. Recent work utilizing chromosome conformation capture (3C) technology in combination with chromatin immunoprecipitation (ChIP) has provided a systematic means of linking non-coding variants within active enhancer loci to putative gene targets. Here, we apply H3K27ac HiChIP high-resolution contact maps, generated from primary human T-cell subsets (CD4+ Naive, TH17, and Treg), to 21 dermatologic conditions associated with single nucleotide polymorphisms (SNPs) from 106 genome-wide association studies (GWAS). This "enhancer connectome" identified 1,492 HiChIP gene-targets from 542 non-coding SNPs (p<5.0x10-8). SNP-containing enhancers from inflammatory skin conditions were significantly enriched within the human leukocyte antigen (HLA)-locus, and also targeted several key factors from the JAK-STAT signaling pathway, while non-immune conditions did not. A focused profiling of systemic lupus erythematosus (SLE) HiChIP-genes identified enhancer interactions with factors important for effector CD4+ T-cell differentiation and function, including interferon regulatory factor 8 (IRF8) and members of the Ikaros family of zinc-finger proteins. Our results demonstrate the ability of the enhancer connectome to nominate functionally-relevant candidates from GWAS-identified variants, representing a powerful tool to guide future studies into the genomic regulatory mechanisms underlying dermatologic diseases.
View details for DOI 10.1016/j.jid.2018.09.011
View details for PubMedID 30315781
Notch2-dependent DC2s mediate splenic germinal center responses.
Proceedings of the National Academy of Sciences of the United States of America
CD4+ T follicular helper (TFH) cells support germinal center (GC) reactions promoting humoral immunity. Dendritic cell (DC) diversification into genetically distinct subsets allows for specialization in promoting responses against several types of pathogens. Whether any classical DC (cDC) subset is required for humoral immunity is unknown, however. We tested several genetic models that selectively ablate distinct DC subsets in mice for their impact on splenic GC reactions. We identified a requirement for Notch2-dependent cDC2s, but not Batf3-dependent cDC1s or Klf4-dependent cDC2s, in promoting TFH and GC B cell formation in response to sheep red blood cells and inactivated Listeria monocytogenes This effect was mediated independent of Il2ra and several Notch2-dependent genes expressed in cDC2s, including Stat4 and Havcr2 Notch2 signaling during cDC2 development also substantially reduced the efficiency of cDC2s for presentation of MHC class II-restricted antigens, limiting the strength of CD4 T cell activation. Together, these results demonstrate a nonredundant role for the Notch2-dependent cDC2 subset in supporting humoral immune responses.
View details for DOI 10.1073/pnas.1809925115
View details for PubMedID 30279176
Integrative analysis of single-cell genomics data by coupled nonnegative matrix factorizations.
Proceedings of the National Academy of Sciences of the United States of America
When different types of functional genomics data are generated on single cells from different samples of cells from the same heterogeneous population, the clustering of cells in the different samples should be coupled. We formulate this "coupled clustering" problem as an optimization problem and propose the method of coupled nonnegative matrix factorizations (coupled NMF) for its solution. The method is illustrated by the integrative analysis of single-cell RNA-sequencing (RNA-seq) and single-cell ATAC-sequencing (ATAC-seq) data.
View details for DOI 10.1073/pnas.1805681115
View details for PubMedID 29987051
Expression of the transcription factor ZBTB46 distinguishes human histiocytic disorders of classical dendritic cell origin.
Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc
Distinguishing classical dendritic cells from other myeloid cell types is complicated by the shared expression of cell surface markers. ZBTB46 is a zinc finger and BTB domain-containing transcription factor, which is expressed by dendritic cells and committed dendritic cell precursors, but not by plasmacytoid dendritic cells, monocytes, macrophages, or other immune cell populations. In this study, we demonstrate that expression of ZBTB46 identifies human dendritic cell neoplasms. We examined ZBTB46 expression in a range of benign and malignant histiocytic disorders and found that ZBTB46 is able to clearly define the dendritic cell identity of many previously unclassified histiocytic disease subtypes. In particular, all examined cases of Langerhans cell histiocytosis and histiocytic sarcoma expressed ZBTB46, while all cases of blastic plasmacytoid dendritic cell neoplasm, chronic myelomonocytic leukemia, juvenile xanthogranuloma, Rosai-Dorfman disease, and Erdheim-Chester disease failed to demonstrate expression of ZBTB46. Moreover, ZBTB46 expression clarified the identity of diagnostically challenging neoplasms, such as cases of indeterminate cell histiocytosis, classifying a fraction of these entities as dendritic cell malignancies. These findings clarify the lineage origins of human histiocytic disorders and distinguish dendritic cell disorders from all other myeloid neoplasms.
View details for DOI 10.1038/s41379-018-0052-4
View details for PubMedID 29743654
The chromatin accessibility landscape of primary human cancers.
Science (New York, N.Y.)
2018; 362 (6413)
We present the genome-wide chromatin accessibility profiles of 410 tumor samples spanning 23 cancer types from The Cancer Genome Atlas (TCGA). We identify 562,709 transposase-accessible DNA elements that substantially extend the compendium of known cis-regulatory elements. Integration of ATAC-seq (the assay for transposase-accessible chromatin using sequencing) with TCGA multi-omic data identifies a large number of putative distal enhancers that distinguish molecular subtypes of cancers, uncovers specific driving transcription factors via protein-DNA footprints, and nominates long-range gene-regulatory interactions in cancer. These data reveal genetic risk loci of cancer predisposition as active DNA regulatory elements in cancer, identify gene-regulatory interactions underlying cancer immune evasion, and pinpoint noncoding mutations that drive enhancer activation and may affect patient survival. These results suggest a systematic approach to understanding the noncoding genome in cancer to advance diagnosis and therapy.
View details for DOI 10.1126/science.aav1898
View details for PubMedID 30361341
- Pembrolizumab for advanced basal cell carcinoma: an investigator-initiated, proof-of-concept study. Journal of the American Academy of Dermatology 2018
Transforming Growth Factor Beta 3 (TGFB3) - a Novel Systemic Sclerosis Susceptibility Locus Involved in Fibrosis and Th17 Cell Development Identified By Genome-Wide Association Study in African Americans from the Genome Research in African American Scleroderma Patients Consortium
View details for Web of Science ID 000411824106434
Cells in Inflamed Islets of Autoimmune Diabetes Mice.
Journal of immunology
In NOD mice, CD11c(+) cells increase greatly with islet inflammation and contribute to autoimmune destruction of pancreatic β cells. In this study, we investigated their origin and mechanism of recruitment. CD11c(+) cells in inflamed islets resembled classical dendritic cells based on their transcriptional profile. However, the majority of these cells were not from the Zbtb46-dependent dendritic-cell lineage. Instead, monocyte precursors could give rise to CD11c(+) cells in inflamed islets. Chemokines Ccl5 and Ccl8 were persistently elevated in inflamed islets and the influx of CD11c(+) cells was partially dependent on their receptor Ccr5. Treatment with islet Ag-specific regulatory T cells led to a marked decrease of Ccl5 and Ccl8, and a reduction of monocyte recruitment. These results implicate a monocytic origin of CD11c(+) cells in inflamed islets and suggest that therapeutic regulatory T cells directly or indirectly regulate their influx by altering the chemotactic milieu in the islets.
View details for DOI 10.4049/jimmunol.1601062
View details for PubMedID 28550204
Revisiting the specificity of the MHC class II transactivator CIITA in classical murine dendritic cells in vivo.
European journal of immunology
Ciita was discovered for its role in regulating transcription of major histocompatibility complex class II (MHCII) genes. Subsequently, CIITA was predicted to control many other genes based on reporter and ChIP-seq analysis but few such predictions have been verified in vivo using Ciita(-/-) mice. Testing these predictions for classical dendritic cells (cDCs) has been particularly difficult, since Ciita(-/-) mice lack MHCII expression required to identify cDCs. However, recent identification of the cDC-specific transcription factor Zbtb46 allows identification of cDCs independently of MHCII expression. We crossed Zbtb46(gfp) mice onto the Ciita(-/-) background and found that all cDC lineages developed in vivo in the absence of Ciita. We then compared the complete transcriptional profile of wild-type and Ciita(-/-) cDCs to define the physiological footprint of CIITA for both immature and activated cDCs. We find that CIITA exerts a highly restricted control over only the MHCII, H2-DO and H2-DM genes, in DC1 and DC2 cDC subsets, but not over other proposed targets, including Ii. These findings emphasize the caveats needed in interpreting transcription factor binding sites identified by in vitro reporter analysis, or by ChIP-seq, which may not necessarily indicate their functional activity in vivo. This article is protected by copyright. All rights reserved.
View details for DOI 10.1002/eji.201747050
View details for PubMedID 28608405
An improved ATAC-seq protocol reduces background and enables interrogation of frozen tissues.
We present Omni-ATAC, an improved ATAC-seq protocol for chromatin accessibility profiling that works across multiple applications with substantial improvement of signal-to-background ratio and information content. The Omni-ATAC protocol generates chromatin accessibility profiles from archival frozen tissue samples and 50-μm sections, revealing the activities of disease-associated DNA elements in distinct human brain structures. The Omni-ATAC protocol enables the interrogation of personal regulomes in tissue context and translational studies.
View details for DOI 10.1038/nmeth.4396
View details for PubMedID 28846090
Discovery of stimulation-responsive immune enhancers with CRISPR activation.
The majority of genetic variants associated with common human diseases map to enhancers, non-coding elements that shape cell-type-specific transcriptional programs and responses to extracellular cues. Systematic mapping of functional enhancers and their biological contexts is required to understand the mechanisms by which variation in non-coding genetic sequences contributes to disease. Functional enhancers can be mapped by genomic sequence disruption, but this approach is limited to the subset of enhancers that are necessary in the particular cellular context being studied. We hypothesized that recruitment of a strong transcriptional activator to an enhancer would be sufficient to drive target gene expression, even if that enhancer was not currently active in the assayed cells. Here we describe a discovery platform that can identify stimulus-responsive enhancers for a target gene independent of stimulus exposure. We used tiled CRISPR activation (CRISPRa) to synthetically recruit a transcriptional activator to sites across large genomic regions (more than 100 kilobases) surrounding two key autoimmunity risk loci, CD69 and IL2RA. We identified several CRISPRa-responsive elements with chromatin features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours an autoimmunity risk variant. Using engineered mouse models, we found that sequence perturbation of the disease-associated Il2ra enhancer did not entirely block Il2ra expression, but rather delayed the timing of gene activation in response to specific extracellular signals. Enhancer deletion skewed polarization of naive T cells towards a pro-inflammatory T helper (TH17) cell state and away from a regulatory T cell state. This integrated approach identifies functional enhancers and reveals how non-coding variation associated with human immune dysfunction alters context-specific gene programs.
View details for DOI 10.1038/nature23875
View details for PubMedID 28854172
Chromatin Accessibility Landscape of Cutaneous T Cell Lymphoma and Dynamic Response to HDAC Inhibitors.
Here, we define the landscape and dynamics of active regulatory DNA in cutaneous T cell lymphoma (CTCL) by ATAC-seq. Analysis of 111 human CTCL and control samples revealed extensive chromatin signatures that distinguished leukemic, host, and normal CD4(+) T cells. We identify three dominant patterns of transcription factor (TF) activation that drive leukemia regulomes, as well as TF deactivations that alter host T cells in CTCL patients. Clinical response to histone deacetylase inhibitors (HDACi) is strongly associated with a concurrent gain in chromatin accessibility. HDACi causes distinct chromatin responses in leukemic and host CD4(+) T cells, reprogramming host T cells toward normalcy. These results provide a foundational framework to study personal regulomes in human cancer and epigenetic therapy.
View details for DOI 10.1016/j.ccell.2017.05.008
View details for PubMedID 28625481
ATAC-see reveals the accessible genome by transposase-mediated imaging and sequencing.
Spatial organization of the genome plays a central role in gene expression, DNA replication, and repair. But current epigenomic approaches largely map DNA regulatory elements outside of the native context of the nucleus. Here we report assay of transposase-accessible chromatin with visualization (ATAC-see), a transposase-mediated imaging technology that employs direct imaging of the accessible genome in situ, cell sorting, and deep sequencing to reveal the identity of the imaged elements. ATAC-see revealed the cell-type-specific spatial organization of the accessible genome and the coordinated process of neutrophil chromatin extrusion, termed NETosis. Integration of ATAC-see with flow cytometry enables automated quantitation and prospective cell isolation as a function of chromatin accessibility, and it reveals a cell-cycle dependence of chromatin accessibility that is especially dynamic in G1 phase. The integration of imaging and epigenomics provides a general and scalable approach for deciphering the spatiotemporal architecture of gene control.
View details for DOI 10.1038/nmeth.4031
View details for PubMedID 27749837
A Long Noncoding RNA lincRNA-EPS Acts as a Transcriptional Brake to Restrain Inflammation
2016; 165 (7): 1672-1685
Long intergenic noncoding RNAs (lincRNAs) are important regulators of gene expression. Although lincRNAs are expressed in immune cells, their functions in immunity are largely unexplored. Here, we identify an immunoregulatory lincRNA, lincRNA-EPS, that is precisely regulated in macrophages to control the expression of immune response genes (IRGs). Transcriptome analysis of macrophages from lincRNA-EPS-deficient mice, combined with gain-of-function and rescue experiments, revealed a specific role for this lincRNA in restraining IRG expression. Consistently, lincRNA-EPS-deficient mice manifest enhanced inflammation and lethality following endotoxin challenge in vivo. lincRNA-EPS localizes at regulatory regions of IRGs to control nucleosome positioning and repress transcription. Further, lincRNA-EPS mediates these effects by interacting with heterogeneous nuclear ribonucleoprotein L via a CANACA motif located in its 3' end. Together, these findings identify lincRNA-EPS as a repressor of inflammatory responses, highlighting the importance of lincRNAs in the immune system.
View details for DOI 10.1016/j.cell.2016.05.075
View details for Web of Science ID 000378062000018
View details for PubMedID 27315481
- Cellular morphology of BRAF V600E-positive Langerhans cell histiocytosis BLOOD 2015; 126 (15): 1857-1857
Heme-Mediated SPI-C Induction Promotes Monocyte Differentiation into Iron-Recycling Macrophages
2014; 156 (6): 1223-1234
Splenic red pulp macrophages (RPM) degrade senescent erythrocytes and recycle heme-associated iron. The transcription factor SPI-C is selectively expressed by RPM and is required for their development, but the physiologic stimulus inducing Spic is unknown. Here, we report that Spic also regulated the development of F4/80(+)VCAM1(+) bone marrow macrophages (BMM) and that Spic expression in BMM and RPM development was induced by heme, a metabolite of erythrocyte degradation. Pathologic hemolysis induced loss of RPM and BMM due to excess heme but induced Spic in monocytes to generate new RPM and BMM. Spic expression in monocytes was constitutively inhibited by the transcriptional repressor BACH1. Heme induced proteasome-dependent BACH1 degradation and rapid Spic derepression. Furthermore, cysteine-proline dipeptide motifs in BACH1 that mediate heme-dependent degradation were necessary for Spic induction by heme. These findings are the first example of metabolite-driven differentiation of a tissue-resident macrophage subset and provide new insights into iron homeostasis.
View details for DOI 10.1016/j.cell.2014.01.069
View details for Web of Science ID 000332945100012
View details for PubMedID 24630724
Embryonic and Adult-Derived Resident Cardiac Macrophages Are Maintained through Distinct Mechanisms at Steady State and during Inflammation
2014; 40 (1): 91-104
Cardiac macrophages are crucial for tissue repair after cardiac injury but are not well characterized. Here we identify four populations of cardiac macrophages. At steady state, resident macrophages were primarily maintained through local proliferation. However, after macrophage depletion or during cardiac inflammation, Ly6c(hi) monocytes contributed to all four macrophage populations, whereas resident macrophages also expanded numerically through proliferation. Genetic fate mapping revealed that yolk-sac and fetal monocyte progenitors gave rise to the majority of cardiac macrophages, and the heart was among a minority of organs in which substantial numbers of yolk-sac macrophages persisted in adulthood. CCR2 expression and dependence distinguished cardiac macrophages of adult monocyte versus embryonic origin. Transcriptional and functional data revealed that monocyte-derived macrophages coordinate cardiac inflammation, while playing redundant but lesser roles in antigen sampling and efferocytosis. These data highlight the presence of multiple cardiac macrophage subsets, with different functions, origins, and strategies to regulate compartment size.
View details for DOI 10.1016/j.immuni.2013.11.019
View details for Web of Science ID 000331476900012
View details for PubMedID 24439267
Extrathymic Aire-Expressing Cells Are a Distinct Bone Marrow-Derived Population that Induce Functional Inactivation of CD4(+) T Cells
2013; 39 (3): 560-572
The autoimmune regulator (Aire) is essential for prevention of autoimmunity; its role is best understood in the thymus, where it promotes self-tolerance through tissue-specific antigen (TSA) expression. Recently, extrathymic Aire-expressing cells (eTACs) have been described in murine secondary lymphoid organs, but the identity of such cells and their role in immune tolerance remains unclear. Here we have shown that eTACs are a discrete major histocompatibility complex class II (MHC II)(hi), CD80(lo), CD86(lo), epithelial cell adhesion molecule (EpCAM)(hi), CD45(lo) bone marrow-derived peripheral antigen-presenting cell (APC) population. We also have demonstrated that eTACs can functionally inactivate CD4⁺ T cells through a mechanism that does not require regulatory T cells (Treg) and is resistant to innate inflammatory stimuli. Together, these findings further define eTACs as a distinct tolerogenic cell population in secondary lymphoid organs.
View details for DOI 10.1016/j.immuni.2013.08.005
View details for Web of Science ID 000330949600018
View details for PubMedID 23993652
Bcl11a Controls Flt3 Expression in Early Hematopoietic Progenitors and Is Required for pDC Development In Vivo
2013; 8 (5)
Bcl11a is a transcription factor known to regulate lymphoid and erythroid development. Recent bioinformatic analysis of global gene expression patterns has suggested a role for Bcl11a in the development of dendritic cell (DC) lineages. We tested this hypothesis by analyzing the development of DC and other lineages in Bcl11a (-/-) mice. We found that Bcl11a was required for expression of IL-7 receptor (IL-7R) and Flt3 in early hematopoietic progenitor cells. In addition, we found severely decreased numbers of plasmacytoid dendritic cells (pDCs) in Bcl11a (-/-) fetal livers and in the bone marrow of Bcl11a (-/-) fetal liver chimeras. Moreover, Bcl11a (-/-) cells showed severely impaired in vitro development of Flt3L-derived pDCs and classical DCs (cDCs). In contrast, we found normal in vitro development of DCs from Bcl11a (-/-) fetal liver cells treated with GM-CSF. These results suggest that the persistent cDC development observed in Bcl11a (-/-) fetal liver chimeras reflects derivation from a Bcl11a- and Flt3-independent pathway in vivo.
View details for DOI 10.1371/journal.pone.0064800
View details for Web of Science ID 000319799900108
View details for PubMedID 23741395
View details for PubMedCentralID PMC3669380
Ly6C(hi) Monocytes in the Inflamed Colon Give Rise to Proinflammatory Effector Cells and Migratory Antigen-Presenting Cells
2012; 37 (6): 1076-1090
Ly6C(hi) monocytes seed the healthy intestinal lamina propria to give rise to resident CX(3)CR1(+) macrophages that contribute to the maintenance of gut homeostasis. Here we report on two alternative monocyte fates in the inflamed colon. We showed that CCR2 expression is essential to the recruitment of Ly6C(hi) monocytes to the inflamed gut to become the dominant mononuclear cell type in the lamina propria during settings of acute colitis. In the inflammatory microenvironment, monocytes upregulated TLR2 and NOD2, rendering them responsive to bacterial products to become proinflammatory effector cells. Ablation of Ly6C(hi) monocytes ameliorated acute gut inflammation. With time, monocytes differentiated into migratory antigen-presenting cells capable of priming naive T cells, thus acquiring hallmarks reminiscent of dendritic cells. Collectively, our results highlight cellular dynamics in the inflamed colon and the plasticity of Ly6C(hi) monocytes, marking them as potential targets for inflammatory bowel disease (IBD) therapy.
View details for DOI 10.1016/j.immuni.2012.08.026
View details for Web of Science ID 000312575000015
View details for PubMedID 23219392
Compensatory dendritic cell development mediated by BATF-IRF interactions
2012; 490 (7421): 502-?
The AP1 transcription factor Batf3 is required for homeostatic development of CD8α(+) classical dendritic cells that prime CD8 T-cell responses against intracellular pathogens. Here we identify an alternative, Batf3-independent pathway in mice for CD8α(+) dendritic cell development operating during infection with intracellular pathogens and mediated by the cytokines interleukin (IL)-12 and interferon-γ. This alternative pathway results from molecular compensation for Batf3 provided by the related AP1 factors Batf, which also functions in T and B cells, and Batf2 induced by cytokines in response to infection. Reciprocally, physiological compensation between Batf and Batf3 also occurs in T cells for expression of IL-10 and CTLA4. Compensation among BATF factors is based on the shared capacity of their leucine zipper domains to interact with non-AP1 factors such as IRF4 and IRF8 to mediate cooperative gene activation. Conceivably, manipulating this alternative pathway of dendritic cell development could be of value in augmenting immune responses to vaccines.
View details for DOI 10.1038/nature11531
View details for Web of Science ID 000310196200034
View details for PubMedID 22992524
View details for PubMedCentralID PMC3482832
Cross-dressed CD8 alpha(+)/CD103(+) dendritic cells prime CD8(+) T cells following vaccination
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2012; 109 (31): 12716-12721
Activation of naïve cluster of differentiation (CD)8(+) cytotoxic T lymphocytes (CTLs) is a tightly regulated process, and specific dendritic cell (DC) subsets are typically required to activate naive CTLs. Potential pathways for antigen presentation leading to CD8(+) T-cell priming include direct presentation, cross-presentation, and cross-dressing. To distinguish between these pathways, we designed single-chain trimer (SCT) peptide-MHC class I complexes that can be recognized as intact molecules but cannot deliver antigen to MHC through conventional antigen processing. We demonstrate that cross-dressing is a robust pathway of antigen presentation following vaccination, capable of efficiently activating both naïve and memory CD8(+) T cells and requires CD8α(+)/CD103(+) DCs. Significantly, immune responses induced exclusively by cross-dressing were as strong as those induced exclusively through cross-presentation. Thus, cross-dressing is an important pathway of antigen presentation, with important implications for the study of CD8(+) T-cell responses to viral infection, tumors, and vaccines.
View details for DOI 10.1073/pnas.1203468109
View details for Web of Science ID 000307538200087
View details for PubMedID 22802630
Dual actions of Meis1 inhibit erythroid progenitor development and sustain general hematopoietic cell proliferation
2012; 120 (2): 335-346
Myeloid ecotropic viral integration site 1 (Meis1) forms a heterodimer with Pbx1 that augments Hox-dependent gene expression and is associated with leukemogenesis and HSC self-renewal. Here we identified 2 independent actions of Meis1 in hematopoietic development: one regulating cellular proliferation and the other involved in megakaryocyte lineage development. First, we found that endogenous Mesp1 indirectly induces Meis1 and Meis2 in endothelial cells derived from embryonic stem cells. Overexpression of Meis1 and Meis2 greatly enhanced the formation of hematopoietic colonies from embryonic stem cells, with the exception of erythroid colonies, by maintaining hematopoietic progenitor cells in a state of proliferation. Second, overexpression of Meis1 repressed the development of early erythroid progenitors, acting in vivo at the megakaryocyte-erythroid progenitor stage to skew development away from erythroid generation and toward megakaryocyte development. This previously unrecognized action of Meis1 may explain the embryonic lethality observed in Meis1(-/-) mice that arises from failure of lymphatic-venous separation and can result as a consequence of defective platelet generation. These results show that Meis1 exerts 2 independent functions, with its role in proliferation of hematopoietic progenitors acting earlier in development from its influence on the fate choice at the megakaryocyte-erythroid progenitor between megakaryocytic and erythroid development.
View details for DOI 10.1182/blood-2012-01-403139
View details for Web of Science ID 000307412400018
View details for PubMedCentralID PMC3628121
IRF-8 extinguishes neutrophil production and promotes dendritic cell lineage commitment in both myeloid and lymphoid mouse progenitors
2012; 119 (9): 2003-2012
While most blood lineages are assumed to mature through a single cellular and developmental route downstream of HSCs, dendritic cells (DCs) can be derived from both myeloid and lymphoid progenitors in vivo. To determine how distinct progenitors can generate similar downstream lineages, we examined the transcriptional changes that accompany loss of in vivo myeloid potential as common myeloid progenitors differentiate into common DC progenitors (CDPs), and as lymphoid-primed multipotent progenitors (LMPPs) differentiate into all lymphoid progenitors (ALPs). Microarray studies revealed that IFN regulatory factor 8 (IRF-8) expression increased during each of these transitions. Competitive reconstitutions using Irf8(-/-) BM demonstrated cell-intrinsic defects in the formation of CDPs and all splenic DC subsets. Irf8(-/-) common myeloid progenitors and, unexpectedly, Irf8(-/-) ALPs produced more neutrophils in vivo than their wild-type counterparts at the expense of DCs. Retroviral expression of IRF-8 in multiple progenitors led to reduced neutrophil production and increased numbers of DCs, even in the granulocyte-macrophage progenitor (GMP), which does not normally possess conventional DC potential. These data suggest that IRF-8 represses a neutrophil module of development and promotes convergent DC development from multiple lymphoid and myeloid progenitors autonomously of cellular context.
View details for DOI 10.1182/blood-2011-06-364976
View details for Web of Science ID 000300949500010
View details for PubMedID 22238324
View details for PubMedCentralID PMC3311244
Transcription factor networks in dendritic cell development
SEMINARS IN IMMUNOLOGY
2011; 23 (5): 388-397
Dendritic cells (DCs) are a heterogeneous population within the mononuclear phagocyte system (MPS) that derive from bone marrow precursors. Commitment and specification of hematopoietic progenitors to the DC lineage is critical for the proper induction of both immunity and tolerance. This review summarizes the important cytokines and transcription factors required for differentiation of the DC lineage as well as further diversification into specific DC subsets. We highlight recent advances in the characterization of immediate DC precursors arising from the common myeloid progenitor (CMP). Particular emphasis is placed on the corresponding temporal expression of relevant factors involved in regulating developmental options.
View details for DOI 10.1016/j.smim.2011.08.009
View details for Web of Science ID 000297447300010
View details for PubMedID 21924924
View details for PubMedCentralID PMC4010935
Targeting of B and T lymphocyte associated (BTLA) prevents graft-versus-host disease without global immunosuppression
JOURNAL OF EXPERIMENTAL MEDICINE
2010; 207 (12): 2551-2559
Graft-versus-host disease (GVHD) causes significant morbidity and mortality in allogeneic hematopoietic stem cell transplantation (aHSCT), preventing its broader application to non-life-threatening diseases. We show that a single administration of a nondepleting monoclonal antibody specific for the coinhibitory immunoglobulin receptor, B and T lymphocyte associated (BTLA), permanently prevented GVHD when administered at the time of aHSCT. Once GVHD was established, anti-BTLA treatment was unable to reverse disease, suggesting that its mechanism occurs early after aHSCT. Anti-BTLA treatment prevented GVHD independently of its ligand, the costimulatory tumor necrosis factor receptor herpesvirus entry mediator (HVEM), and required BTLA expression by donor-derived T cells. Furthermore, anti-BTLA treatment led to the relative inhibition of CD4(+) forkhead box P3(-) (Foxp3(-)) effector T cell (T eff cell) expansion compared with precommitted naturally occurring donor-derived CD4(+) Foxp3(+) regulatory T cell (T reg cell) and allowed for graft-versus-tumor (GVT) effects as well as robust responses to pathogens. These results suggest that BTLA agonism rebalances T cell expansion in lymphopenic hosts after aHSCT, thereby preventing GVHD without global immunosuppression. Thus, targeting BTLA with a monoclonal antibody at the initiation of aHSCT therapy might reduce limitations imposed by histocompatibility and allow broader application to treatment of non-life-threatening diseases.
View details for DOI 10.1084/jem.20102017
View details for Web of Science ID 000285505000003
View details for PubMedID 21078889
Commercially Available Outbred Mice for Genome-Wide Association Studies
2010; 6 (9)
Genome-wide association studies using commercially available outbred mice can detect genes involved in phenotypes of biomedical interest. Useful populations need high-frequency alleles to ensure high power to detect quantitative trait loci (QTLs), low linkage disequilibrium between markers to obtain accurate mapping resolution, and an absence of population structure to prevent false positive associations. We surveyed 66 colonies for inbreeding, genetic diversity, and linkage disequilibrium, and we demonstrate that some have haplotype blocks of less than 100 Kb, enabling gene-level mapping resolution. The same alleles contribute to variation in different colonies, so that when mapping progress stalls in one, another can be used in its stead. Colonies are genetically diverse: 45% of the total genetic variation is attributable to differences between colonies. However, quantitative differences in allele frequencies, rather than the existence of private alleles, are responsible for these population differences. The colonies derive from a limited pool of ancestral haplotypes resembling those found in inbred strains: over 95% of sequence variants segregating in outbred populations are found in inbred strains. Consequently it is possible to impute the sequence of any mouse from a dense SNP map combined with inbred strain sequence data, which opens up the possibility of cataloguing and testing all variants for association, a situation that has so far eluded studies in completely outbred populations. We demonstrate the colonies' potential by identifying a deletion in the promoter of H2-Ea as the molecular change that strongly contributes to setting the ratio of CD4+ and CD8+ lymphocytes.
View details for DOI 10.1371/journal.pgen.1001085
View details for Web of Science ID 000282369200055
View details for PubMedID 20838427
Enhanced thymic selection of FoxP3(+) regulatory T cells in the NOD mouse model of autoimmune diabetes
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2007; 104 (46): 18181-18186
FoxP3(+)CD4(+) regulatory T cells (Tregs) play a key role in the maintenance of peripheral self-tolerance, and it has been suggested that diabetes-susceptible nonobese diabetic (NOD) mice are defective in the generation and numbers of Tregs. We found thymic selection of Tregs to be under genetic control. Fetal thymic organ cultures on the NOD background required 3- to 10-fold more antigen than corresponding cultures on the B6 background for optimal induction of Tregs, but once the threshold for induction was reached the NOD background yielded close to 10-fold more Tregs. This increased selection of Tregs was also found in nontransgenic NOD mice in fetal through adult stages. This trait did not map to the MHC, idd3, or the chromosome 3 (Chr3) regions that control clonal deletion, but mainly to two regions on Chr1 and Chr11. Thus, NOD mice do not have a global defect in the generation or maintenance of Tregs; if anything, they show the opposite.
View details for DOI 10.1073/pnas.0708899104
View details for Web of Science ID 000251077000052
View details for PubMedID 17991775
- Cytokines in type 2 diabetes INTERLEUKINS 2006; 74: 405-441
IL-1 beta-mediated innate immunity is amplified in the db/db mouse model of type 2 diabetes
JOURNAL OF IMMUNOLOGY
2005; 174 (8): 4991-4997
Chronic inflammation appears to play a critical role in type 2 diabetes and its complications. Here we tested the hypothesis that this inflammatory dysregulation affects the IL-1beta system and has functional consequences in the brain. Diabetic, db/db, and nondiabetic, db/+, mice were administered i.p. LPS, a potent cytokine inducer, at a dose of 100 microg/kg/mouse. db/db mouse innate immune-associated sickness behavior was 14.8, 33, 44.7, and 34% greater than that of db/+ mice at 2, 4, 8, and 12 h, respectively. When a fixed dose of LPS was used (5 microg/mouse), db/db mouse sickness was again enhanced 18.4, 22.2, and 14.5% at 4, 8, and 12 h as compared with db/+ mice. In diabetic mice, peritoneal macrophages produced more IL-1beta in response to LPS, and peritoneal levels of IL-1beta induced by LPS were increased. Importantly, IL-1R antagonist and type 2 IL-1 receptor (IL-1R2) failed to up-regulate in response to LPS in db/db mice. Finally, both peripheral and central administration of IL-1beta, itself, induced sickness in db/db mice that mimicked the effects of peripheral LPS and was significantly greater than that seen in db/+ mice. Taken together, these results indicate that IL-1beta-mediated innate immunity is augmented in db/db mice both at the periphery and in the brain, and the mechanism is due to diabetes-associated loss of IL-1beta counterregulation.
View details for Web of Science ID 000228234600069
View details for PubMedID 15814729