Cis interactions in the Irf8 locus regulate stage-dependent enhancer activation

Individual elements within a superenhancer can act in a cooperative or temporal manner, but the underlying mechanisms remain obscure. We recently identified a

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

Heme-Mediated SPI-C Induction Promotes Monocyte Differentiation into Iron-Recycling Macrophages

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

Affinity-restricted memory B cells dominate recall responses to heterologous flaviviruses

Memory B cells (MBCs) can respond to heterologous antigens either by molding new specificities through secondary germinal centers (GCs) or selecting pre-existing clones without further affinity maturation. To distinguish these mechanisms in flavivirus infections and immunizations, we studied recall responses to envelope protein domain III (DIII). Conditional deletion of activation induced cytidine deaminase (AID) between heterologous challenges of West Nile, Japanese encephalitis, Zika, and Dengue viruses did not affect recall responses. DIII-specific MBCs were contained mostly within the plasma cell-biased CD80(+) subset and few GCs arose following heterologous boosters, demonstrating that recall responses are confined by pre-existing clonal diversity. Measurement of monoclonal antibody binding affinity to DIII proteins, timed AID deletion, single cell RNA-sequencing, and lineage tracing experiments point to selection of relatively low affinity MBCs as a mechanism to promote diversity. Engineering immunogens to avoid this MBC diversity may facilitate flavivirus type-specific vaccines with minimized potential for infection enhancement

Zbtb46 expression distinguishes classical dendritic cells and their committed progenitors from other immune lineages

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

Targeting of B and T lymphocyte associated (BTLA) prevents graft-versus-host disease without global immunosuppression

One-time treatment with an antibody against BTLA provides long-term protection against graft-versus-host disease without affecting effector T cell responses to tumors or pathogens

GWAS for systemic sclerosis identifies multiple risk loci and highlights fibrotic and vasculopathy pathways

Systemic sclerosis (SSc) is an autoimmune disease that shows one of the highest mortality rates among rheumatic diseases. We perform a large genome-wide association study (GWAS), and meta-analysis with previous GWASs, in 26,679 individuals and identify 27 independent genome-wide associated signals, including 13 new risk loci. The novel associations nearly double the number of genome-wide hits reported for SSc thus far. We define 95% credible sets of less than 5 likely causal variants in 12 loci. Additionally, we identify specific SSc subtype-associated signals. Functional analysis of high-priority variants shows the potential function of SSc signals, with the identification of 43 robust target genes through HiChIP. Our results point towards molecular pathways potentially involved in vasculopathy and fibrosis, two main hallmarks in SSc, and highlight the spectrum of critical cell types for the disease. This work supports a better understanding of the genetic basis of SSc and provides directions for future functional experiments

Development and Function of Classical Dendritic Cell Lineages

Classical dendritic cells (cDCs) are essential initiators of innate and adaptive immune responses to pathogens. However, study of these critical cells has been complicated by their similarity to other hematopoietic lineages by cell surface markers. We identified Zbtb46 as a transcription factor selectively expressed by cDCs and their committed progenitors, but not by plasmacytoid DCs (pDCs), monocytes, macrophages or other related 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 Zbtb46gfp mice recapitulated the cDC-specific expression of the native locus, being restricted to cDC precursors and lymphoid-organ and tissue-resident cDCs. Although cDCs developed in Zbtb46gfp/gfp (Zbtb46-deficient) mice, they maintained expression of granulocyte colony-stimulating factor and leukemia inhibitory factor receptors, which are normally down-regulated in order to restrict cDC responsiveness to non-cDC growth factors. Importantly, Zbtb46gfp mice in combination with mice expressing the diphtheria toxin receptor (DTR) under the control of the Zbtb46 locus allowed for specific visualization and depletion of cDCs in vivo. A further complication in the analysis of cDC function in vivo is the developmental specialization of distinct cDC subsets. Previous work in our lab generated mice deficient in the transcription factor BATF3, which resulted in the specific loss of CD8+ cDCs. Batf3-/- mice were unable to survive infection with the parasite Toxoplasma gondii or reject immunogenic tumors. In contrast, the role of the complementary CD11b+ cDC subset remains unclear. We identified Notch2 as a critical regulator of CD11b+ cDC differentiation; mice deficient in Notch2 in myeloid cells specifically lacked this population in vivo. In combination with Zbtb46DTR mice, we determined that Zbtb46+ Notch2-dependent cDCs were specifically required for survival after infection with the enterohemorrhagic bacterium Citrobacter rodentium. Following infection, CD11b+ cDCs were required for the induction of anti-microbial peptides from epithelial cells. Collectively, our studies have clarified the identity of the cDC lineage and provided the first demonstration of a non-redundant function of CD11b+ cDCs in response to pathogens in vivo. In humans, manipulation of this cDC subset may be useful for enhancing immunity to related enterohemorrhagic pathogens