Faculty Bibliography

Plasmacytoid dendritic cells (pDCs) mount powerful antiviral type I interferon (IFN-I) responses, yet only a fraction of pDCs produces high levels of IFN-I. Here we report that peripheral pDCs in naive mice comprise three subsets (termed A, B and C) that represent progressive differentiation stages. This heterogeneity was generated by tonic IFN-I signaling elicited in part by the cGAS/STING and TLR9 DNA-sensing pathways. A small 'IFN-I-naive' subset (pDC-A) could give rise to other subsets; it was expanded in STING deficiency or after the IFN-I receptor blockade, but was abolished by exogenous IFN-I. In response to RNA viruses, pDC-A showed increased Bcl2-dependent survival and superior IFN-I responses, but was susceptible to virus infection. Conversely, the majority of pDCs comprised the 'IFN-I-primed' subsets (pDC-B/C) that showed lower IFN-I responses and poor survival, but did not support virus replication. Thus, tonic IFN-I signaling decreases the cytokine-producing capacity and survival of pDCs but increases their virus resistance, facilitating optimal antiviral responses.

Breast tumors harbor dynamic microenvironments, with multiple immune cell types playing opposing roles during tumor progression and/or response to therapy. Tumor-associated macrophages promote mammary tumorigenesis, whereas the role of mammary tissue macrophages (MTMs) remains incompletely understood. High-dimensional immunostaining of murine mammary tumor progression revealed that MTMs were localized in the peritumoral stroma and associated with eosinophils, which were previously shown to facilitate antitumor T cell responses. The depletion of MTMs accelerated tumorigenesis in both spontaneous and orthotopically transplanted mammary tumor models. Upon induction of a productive antitumor response via the depletion of regulatory T cells, MTMs assumed an alternatively activated state and expressed eotaxins, thereby attracting eosinophils to peritumoral regions. MTMs expressed the receptor for the alarmin IL-33, which induced both MTM activation and eosinophil recruitment. These results suggest that MTMs can sense IL-33 and recruit eosinophils to facilitate antitumor immunity, a mechanism that may operate during tumor progression and be further enhanced during productive antitumor responses.

To determine the cellular and molecular basis of Castleman Disease (CD), we analyze the spatial proteome and transcriptome from a discovery (n = 9 cases) and validation (n = 13 cases) cohort of Unicentric CD, idiopathic Multicentric CD, HHV8-associated MCD, and reactive lymph nodes. CD shows increased stromal cells that form unique microenvironments. Interaction of activated follicular dendritic cell (FDC) cytoplasmic meshworks with mantle-zone B cells is associated with B-cell activation and differentiation. CXCL13+ FDCs, PDGFRA + T-zone reticular cells (TRC), and ACTA2-positive perivascular reticular cells (PRC) were the predominant source of increased VEGF expression and IL-6 signaling. MCD is characterized by increased TRC while UCD shows increased B-reticular cells (BRC). VEGF expression by FDCs is associated with peri-follicular neovascularization. FDC, TRC and PRC of CD activates JAK-STAT, TGFβ, and MAPK pathways via specific ligand-receptor interactions. Here, we show that stromal-cell activation and associated B cell activation and differentiation, neovascularization and stromal remodeling underlie CD.

Leukemia-driving mutations are thought to arise in hematopoietic stem cells (HSC), yet the natural history of their spread is poorly understood. We genetically induced mutations within endogenous murine HSC and traced them in unmanipulated animals. In contrast to mutations associated with clonal hematopoiesis (such as Tet2 deletion), the leukemogenic KrasG12D mutation dramatically accelerated HSC contribution to all hematopoietic lineages. The acceleration was mediated by KrasG12D-expressing multipotent progenitors (MPP) that lacked self-renewal but showed increased proliferation and aberrant transcriptome. The deletion of osteopontin, a secreted negative regulator of stem/progenitor cells, delayed the early expansion of mutant progenitors. KrasG12D-carrying cells showed increased CXCR4-driven motility in the bone marrow, and the blockade of CXCR4 reduced the expansion of MPP in vivo. Finally, therapeutic blockade of KRASG12D spared mutant HSC but reduced the expansion of mutant MPP and their mature progeny. Thus, transforming mutations facilitate their own spread from stem cells by reprogramming MPP, creating a preleukemic state via a two-component stem/progenitor circuit.

Plasmacytoid dendritic cells (pDCs) represent a unique cell type within the innate immune system. Their defining property is the recognition of pathogen-derived nucleic acids through endosomal Toll-like receptors and the ensuing production of type I interferon and other soluble mediators, which orchestrate innate and adaptive responses. We review several aspects of pDC biology that have recently come to the fore. We discuss emerging questions regarding the lineage affiliation and origin of pDCs and argue that these cells constitute an integral part of the dendritic cell lineage. We emphasize the specific function of pDCs as innate sentinels of virus infection, particularly their recognition of and distinct response to virus-infected cells. This essential evolutionary role of pDCs has been particularly important for the control of coronaviruses, as demonstrated by the recent COVID-19 pandemic. Finally, we highlight the key contribution of pDCs to systemic lupus erythematosus, in which therapeutic targeting of pDCs is currently underway.

The development of dendritic cells (DCs), including antigen-presenting conventional DCs (cDCs) and cytokine-producing plasmacytoid DCs (pDCs), is controlled by the growth factor Flt3 ligand (Flt3L) and its receptor Flt3. We genetically dissected Flt3L-driven DC differentiation using CRISPR-Cas9-based screening. Genome-wide screening identified multiple regulators of DC differentiation including subunits of TSC and GATOR1 complexes, which restricted progenitor growth but enabled DC differentiation by inhibiting mTOR signaling. An orthogonal screen identified the transcriptional repressor Trim33 (TIF-1γ) as a regulator of DC differentiation. Conditional targeting in vivo revealed an essential role of Trim33 in the development of all DCs, but not of monocytes or granulocytes. In particular, deletion of Trim33 caused rapid loss of DC progenitors, pDCs, and the cross-presenting cDC1 subset. Trim33-deficient Flt3⁺ progenitors up-regulated pro-inflammatory and macrophage-specific genes but failed to induce the DC differentiation program. Collectively, these data elucidate mechanisms that control Flt3L-driven differentiation of the entire DC lineage and identify Trim33 as its essential regulator.

The hierarchical model of hematopoiesis posits that self-renewing, multipotent hematopoietic stem cells (HSCs) give rise to all blood cell lineages. While this model accounts for hematopoiesis in transplant settings, its applicability to steady-state hematopoiesis remains to be clarified. Here, we used inducible clonal DNA barcoding of endogenous adult HSCs to trace their contribution to major hematopoietic cell lineages in unmanipulated animals. While the majority of barcodes were unique to a single lineage, we also observed frequent barcode sharing between multiple lineages, specifically between lymphocytes and myeloid cells. These results suggest that both single-lineage and multilineage contributions by HSCs collectively drive continuous hematopoiesis, and highlight a close relationship of myeloid and lymphoid development.

Autoantibodies to chromatin and dsDNA are a hallmark of systemic lupus erythematosus (SLE). In a mouse model of monogenic human SLE caused by DNASE1L3 deficiency, the anti-DNA response is dependent on endosomal nucleic acid-sensing TLRs TLR7 and TLR9. In this study, we report that this response also required TLR2, a surface receptor for microbial products that is primarily expressed on myeloid cells. Cell transfers into lymphopenic DNASE1L3-deficient mice showed that TLR2 was required for anti-DNA Ab production by lymphocytes. TLR2 was detectably expressed on B cells and facilitated the production of IL-6 by B cells activated in the presence of microbial products. Accordingly, treatment with broad-spectrum antibiotics or Ab-mediated blockade of IL-6 delayed the anti-DNA response in DNASE1L3-deficient mice. These studies reveal an unexpected B cell-intrinsic role of TLR2 in systemic autoreactivity to DNA, and they suggest that microbial products may synergize with self-DNA in the activation of autoreactive B cells in SLE.

The chromosome 9p21 locus comprises several tumor suppressor genes including MTAP, CDKN2A and CDKN2B, and its homo- or heterozygous deletion is associated with reduced survival in multiple cancer types. We report that mice with germline monoallelic deletion or induced biallelic deletion of the 9p21-syntenic locus (9p21s) developed a fatal myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN)-like disease associated with aberrant trabecular bone formation and/or fibrosis in the bone marrow (BM). Reciprocal BM transfers and conditional targeting of 9p21s suggested that the disease originates in the BM stroma. Single-cell analysis of 9p21s-deficient BM stroma revealed the expansion of chondrocyte and osteogenic precursors, reflected in increased osteogenic differentiation in vitro. It also showed reduced expression of factors maintaining hematopoietic stem/progenitor cells, including Cxcl12. Accordingly, 9p21s-deficient mice showed reduced levels of circulating Cxcl12 and concomitant upregulation of the pro-fibrotic chemokine Cxcl13 and osteogenesis- and fibrosis-related multifunctional glycoprotein Osteopontin (OPN)/Spp1. Our study highlights the potential of mutations in the BM microenvironment to drive MDS/MPN-like disease.