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CXCR5 mediates homing of both B and follicular helper T (TFH) cells into follicles of secondary lymphoid organs. We found that CXCR5+CD8+T cells are present in human tonsils and follicular lymphoma, inhibit TFH-mediated B cell differentiation, and exhibit strong cytotoxic activity. Consistent with these findings, adoptive transfer of CXCR5+CD8+T cells into an animal model of lymphoma resulted in significantly greater antitumor activity than CXCR5−CD8+T cells. Furthermore, RNA-Seq-based transcriptional profiling revealed 77 differentially expressed genes unique to CXCR5+CD8+T cells. Among these, a signature comprised of 33 upregulated genes correlated with improved survival in follicular lymphoma patients. We also showed that CXCR5+CD8+T cells could be induced and expanded ex vivo using IL-23 plus TGF-β, suggesting a possible strategy to generate these cells for clinical application. In summary, our study identified CXCR5+CD8+T cells as a distinct T cell subset with ability to suppress TFH-mediated B cell differentiation, exert strong antitumor activity, and confer favorable prognosis in follicular lymphoma patients.

Dendritic cells (DCs) are crucial for mediating immune responses but, when deregulated, also contribute to immunological disorders, such as autoimmunity. The molecular mechanism underlying the function of DCs is incompletely understood. In this study, we have identified TANK-binding kinase 1 (TBK1), a master innate immune kinase, as an important regulator of DC function. DC-specific deletion of Tbk1 causes T cell activation and autoimmune symptoms and also enhances antitumor immunity in animal models of cancer immunotherapy. The TBK1-deficient DCs have up-regulated expression of co-stimulatory molecules and increased T cell–priming activity. We further demonstrate that TBK1 negatively regulates the induction of a subset of genes by type I interferon receptor (IFNAR). Deletion of IFNAR1 could largely prevent aberrant T cell activation and autoimmunity in DC-conditional Tbk1 knockout mice. These findings identify a DC-specific function of TBK1 in the maintenance of immune homeostasis and tolerance.

Immune checkpoint blockade (ICB) has revolutionized cancer treatment, yet quality of life and continuation of therapy can be constrained by immune-related adverse events (irAEs). Limited understanding of irAE mechanisms hampers development of approaches to mitigate their damage. To address this, we examined whether mice gained sensitivity to anti-CTLA-4 (αCTLA-4)–mediated toxicity upon disruption of gut homeostatic immunity. We found αCTLA-4 drove increased inflammation and colonic tissue damage in mice with genetic predisposition to intestinal inflammation, acute gastrointestinal infection, transplantation with a dysbiotic fecal microbiome, or dextran sodium sulfate administration. We identified an immune signature of αCTLA-4–mediated irAEs, including colonic neutrophil accumulation and systemic interleukin-6 (IL-6) release. IL-6 blockade combined with antibiotic treatment reduced intestinal damage and improved αCTLA-4 therapeutic efficacy in inflammation-prone mice. Intestinal immune signatures were validated in biopsies from patients with ICB colitis. Our work provides new preclinical models of αCTLA-4 intestinal irAEs, mechanistic insights into irAE development, and potential approaches to enhance ICB efficacy while mitigating irAEs.

G‐CSF stimulates the expression of the chemokine MIP‐2 in mature neutrophils by a direct transcriptional mechanism dependent upon STAT3. Neutrophil mobilization from the bone marrow is a critical aspect of the innate immune response, enabling a rapid deployment of phagocytes to infected or inflamed tissue. The cytokine G‐CSF, which is induced rapidly during infection, elicits a swift and potent mobilizing response, yet its mechanisms of action remain poorly understood. Here, we studied the role of G‐CSF and its principal signal transducer STAT3 in regulating expression of the neutrophil chemoattractant MIP‐2. Our studies revealed Gr‐1himature neutrophils as major sources of Cxcl2(MIP‐2) mRNA in bone marrow and G‐CSF‐responsive MIP‐2 protein production. Induction of Cxcl2was regulated directly by G‐CSF‐activated STAT3 via interaction at a STAT consensus element in the Cxcl2promoter. G‐CSF coordinately stimulated the association of STAT3, induction of the transcriptionally active H3K4me3 modification, and recruitment of RNA Pol II at the Cxcl2proximal promoter, as well as the promoter region of Il8rb, encoding the MIP‐2 receptor. These results suggest that the G‐CSF–STAT3 pathway directly regulates transcriptional events that induce neutrophil mobilization.

Cytokines and transcription factors play key roles in dendritic cell (DC) development, yet information about regulatory interactions between these signals remains limited. Here we show that the cytokines GM-CSF and Flt3L induce the transcriptional mediators Id2 and E2-2 and control DC lineage diversification by STAT–dependent pathways. We found that STAT5 is required for tissue CD103+ DC generation and plasmacytoid DC (pDC) suppression in steady state or response to GM-CSF. STAT5 stimulates GM-CSF–dependent expression of Id2, which controls CD103+ DC production and pDC inhibition. By contrast, pDCs, but not CD103+ DCs, are dependent on STAT3. Consistently, STAT3 stimulates Flt3L-responsive expression of the pDC regulator Tcf4 (E2-2). These data suggest that STATs contribute to DC development by controlling transcription factors involved in lineage differentiation.

Cytokines and transcription factors play key roles in dendritic cell (DC) development, yet information about regulatory interactions between these signals remains limited. Here we show that the cytokines GM-CSF and Flt3L induce the transcriptional mediators Id2 and E2-2 and control DC lineage diversification by STAT–dependent pathways. We found that STAT5 is required for tissue CD103+DC generation and plasmacytoid DC (pDC) suppression in steady state or response to GM-CSF. STAT5 stimulates GM-CSF–dependent expression of Id2, which controls CD103+DC production and pDC inhibition. By contrast, pDCs, but not CD103+DCs, are dependent on STAT3. Consistently, STAT3 stimulates Flt3L-responsive expression of the pDC regulator Tcf4(E2-2). These data suggest that STATs contribute to DC development by controlling transcription factors involved in lineage differentiation.

Review of the role for cytokines and cytokine‐responsive STAT transcription factors in dendritic cell development. DCs have a vital role in the immune system by recognizing exogenous or self‐antigens and eliciting appropriate stimulatory or tolerogenic adaptive immune responses. DCs also contribute to human autoimmune disease and, when depleted, to immunodeficiency. Moreover, DCs are being explored for potential use in clinical therapies including cancer treatment. Thus, understanding the molecular mechanisms that regulate DCs is crucial to improving treatments for human immune disease and cancer. DCs constitute a heterogeneous population including plasmacytoid (pDC) and classic (cDC) subsets; however, the majority of DCs residing in lymphoid organs and peripheral tissues in steady state share common progenitor populations, originating with hematopoietic stem cells. Like other hematopoietic lineages, DCs require extracellular factors including cytokines, as well as intrinsic transcription factors, to control lineage specification, commitment, and maturation. Here, we review recent findings on the roles for cytokines and cytokine‐activated STAT transcription factors in DC subset development. We also discuss how cytokines and STATs intersect with lineage‐regulatory transcription factors and how insight into the molecular basis of human disease has revealed transcriptional regulators of DCs. Whereas this is an emerging area with much work remaining, we anticipate that knowledge gained by delineating cytokine and transcription factor mechanisms will enable a better understanding of DC subset diversity, and the potential to manipulate these important immune cells for human benefit.

Glioma is one of the most common primary brain tumors. Despite surgical resection, radiotherapy, and chemotherapy, the prognosis of patients with malignant glioma remains poor. Programmed cell death 5 (PDCD5) is a newly described pro-apoptotic protein. Our previous study showed that PDCD5 downregulation in gliomas was associated with higher pathological grade. Here, we investigated the effect of PDCD5 on chemosensitivity of glioma cells and its mechanism. We demonstrated that overexpression or knockdown of PDCD5 had no significant effect on the proliferation of glioma cell lines (U87, U251, and T98G) in the absence of chemotherapeutic agents. However, PDCD5 overexpression effectively sensitized U87 cells to chemotherapeutic drugs (cisplatin, carboplatin, and vincristine) in a concentration-dependent manner, while its knockdown resulted in decreased chemosensitivity in U251, T98G, and U87 cells. Importantly, expression of PDCD5 also markedly inhibited tumor cell proliferation and colony formation in the presence of low doses of cisplatin. Furthermore, we found that PDCD5 expression promoted cisplatin-induced apoptosis, increased markedly the activation of caspase-3 and caspase-9, and decreased significantly the ratio of Bcl-2/Bax proteins, but had no effect on the activation of caspase-8. Taken together, our findings indicate that PDCD5 promotes chemosensitivity by activating mitochondria-related apoptotic pathway, and that the combination of PDCD5 and chemotherapeutic drugs such as cisplatin, is expected to be an effective therapeutic strategy for the malignant glioma.

Plasmacytoid dendritic cells (pDCs) reside in bone marrrow and lymphoid organs in homeostatic conditions and typically secrete abundant quantities of type I interferons (IFNs) on Toll-like receptor triggering. Recently, a pDC population was identified within Peyer patches (PPs) of the gut that is distinguished by its lack of IFN production; however, the relationship of PP pDCs to pDCs in other organs has been unclear. We report that PP pDCs are derived from common DC progenitors and accumulate in response to Fms-like tyrosine kinase 3 ligand, yet appear divergent in transcription factor profile and surface marker phenotype, including reduced E2-2 and CCR9 expression. Type I IFN signaling via STAT1 has a cell-autonomous role in accrual of PP pDCs in vivo. Moreover, IFN-α enhances pDC generation from DC progenitors by a STAT1-dependent mechanism. pDCs that have been developed in the presence of IFN-α resemble PP pDCs, produce inflammatory cytokines, stimulate Th17 cell generation, and fail to secrete IFN-α on Toll-like receptor engagement. These results indicate that IFN-α influences the development and function of pDCs by inducing emergence of an inflammatory (Th17-inducing) antigen-presenting subset, and simultaneously regulating accumulation of pDCs in the intestinal microenvironment.

Plasmacytoid dendritic cells (pDCs) reside in bone marrrow and lymphoid organs in homeostatic conditions and typically secrete abundant quantities of type I interferons (IFNs) on Toll-like receptor triggering. Recently, a pDC population was identified within Peyer patches (PPs) of the gut that is distinguished by its lack of IFN production; however, the relationship of PP pDCs to pDCs in other organs has been unclear. We report that PP pDCs are derived from common DC progenitors and accumulate in response to Fms-like tyrosine kinase 3 ligand, yet appear divergent in transcription factor profile and surface marker phenotype, including reduced E2-2 and CCR9 expression. Type I IFN signaling via STAT1 has a cell-autonomous role in accrual of PP pDCs in vivo. Moreover, IFN-α enhances pDC generation from DC progenitors by a STAT1-dependent mechanism. pDCs that have been developed in the presence of IFN-α resemble PP pDCs, produce inflammatory cytokines, stimulate Th17 cell generation, and fail to secrete IFN-α on Toll-like receptor engagement. These results indicate that IFN-α influences the development and function of pDCs by inducing emergence of an inflammatory (Th17-inducing) antigen-presenting subset, and simultaneously regulating accumulation of pDCs in the intestinal microenvironment.

Granulocyte colony-stimulating factor (G-CSF) mediates “emergency” granulopoiesis during infection, a process that is mimicked by clinical G-CSF use, yet we understand little about the intracellular signaling cascades that control demand-driven neutrophil production. Using a murine model with conditional deletion of signal transducer and activator of transcription 3 (STAT3) in bone marrow, we investigated the cellular and molecular mechanisms of STAT3 function in the emergency granulopoiesis response to G-CSF administration or infection with Listeria monocytogenes, a pathogen that is restrained by G-CSF signaling in vivo. Our results show that STAT3 deficiency renders hematopoietic progenitor cells and myeloid precursors refractory to the growth-promoting functions of G-CSF or L monocytogenes infection. STAT3 is necessary for accelerating granulocyte cell-cycle progression and maturation in response to G-CSF. STAT3 directly controls G-CSF–dependent expression of CCAAT-enhancer-binding protein β (C/EBPβ), a crucial factor in the emergency granulopoiesis response. Moreover, STAT3 and C/EBPβ coregulate c-Myc through interactions with the c-myc promoter that control the duration of C/EBPα occupancy during demand-driven granulopoiesis. These results place STAT3 as an essential mediator of emergency granulopoiesis by its regulation of transcription factors that direct G-CSF–responsive myeloid progenitor expansion.

Granulocyte colony-stimulating factor (G-CSF) mediates “emergency” granulopoiesis during infection, a process that is mimicked by clinical G-CSF use, yet we understand little about the intracellular signaling cascades that control demand-driven neutrophil production. Using a murine model with conditional deletion of signal transducer and activator of transcription 3 (STAT3) in bone marrow, we investigated the cellular and molecular mechanisms of STAT3 function in the emergency granulopoiesis response to G-CSF administration or infection with Listeria monocytogenes, a pathogen that is restrained by G-CSF signaling in vivo. Our results show that STAT3 deficiency renders hematopoietic progenitor cells and myeloid precursors refractory to the growth-promoting functions of G-CSF or L monocytogenesinfection. STAT3 is necessary for accelerating granulocyte cell-cycle progression and maturation in response to G-CSF. STAT3 directly controls G-CSF–dependent expression of CCAAT-enhancer-binding protein β (C/EBPβ), a crucial factor in the emergency granulopoiesis response. Moreover, STAT3 and C/EBPβ coregulate c-Myc through interactions with the c-mycpromoter that control the duration of C/EBPα occupancy during demand-driven granulopoiesis. These results place STAT3 as an essential mediator of emergency granulopoiesis by its regulation of transcription factors that direct G-CSF–responsive myeloid progenitor expansion.

Neutrophil mobilization, the release of neutrophils from the bone marrow reserve into circulating blood, is important to increase peripheral neutrophil amounts during bacterial infections. Granulocyte colony-stimulating factor (G-CSF) and chemokines, such as macrophage-inflammatory protein-2 (MIP-2; CXCL2), can induce neutrophil mobilization, but the mechanism(s) they use remain unclear. Signal transducers and activator of transcription 3 (STAT3) is the principal intracellular signaling molecule activated upon G-CSF ligation of its receptor. Using a murine model with conditional STAT3 deletion in bone marrow, we demonstrated previously that STAT3 regulates acute G-CSF–responsive neutrophil mobilization and MIP-2–dependent neutrophil chemotaxis. In this study, we show STAT3 is also necessary for MIP-2–elicited neutrophil mobilization. STAT3 appears to function by controlling extracellular signal-regulated kinase (ERK) activation, which is important for MIP-2–mediated chemotaxis. In addition, we demonstrate that G-CSF stimulates the expression of the MIP-2 receptor via STAT3-dependent transcriptional activation of Il8rb. G-CSF treatment also induces STAT3-dependent changes in bone marrow chemokine expression levels which may further affect neutrophil retention and release. Taken together, our study demonstrates that STAT3 regulates multiple aspects of chemokine and chemokine receptor expression and function within the bone marrow, indicating a central role in the neutrophil mobilization response.

Neutrophil mobilization, the release of neutrophils from the bone marrow reserve into circulating blood, is important to increase peripheral neutrophil amounts during bacterial infections. Granulocyte colony-stimulating factor (G-CSF) and chemokines, such as macrophage-inflammatory protein-2 (MIP-2; CXCL2), can induce neutrophil mobilization, but the mechanism(s) they use remain unclear. Signal transducers and activator of transcription 3 (STAT3) is the principal intracellular signaling molecule activated upon G-CSF ligation of its receptor. Using a murine model with conditional STAT3 deletion in bone marrow, we demonstrated previously that STAT3 regulates acute G-CSF–responsive neutrophil mobilization and MIP-2–dependent neutrophil chemotaxis. In this study, we show STAT3 is also necessary for MIP-2–elicited neutrophil mobilization. STAT3 appears to function by controlling extracellular signal-regulated kinase (ERK) activation, which is important for MIP-2–mediated chemotaxis. In addition, we demonstrate that G-CSF stimulates the expression of the MIP-2 receptor via STAT3-dependent transcriptional activation of Il8rb. G-CSF treatment also induces STAT3-dependent changes in bone marrow chemokine expression levels which may further affect neutrophil retention and release. Taken together, our study demonstrates that STAT3 regulates multiple aspects of chemokine and chemokine receptor expression and function within the bone marrow, indicating a central role in the neutrophil mobilization response.

Blocking tumor-derived cytokine signaling in dendritic cells enables engineered dendritic cells to function as an antitumor vaccine.

Nguyen-Jackson: Amgen, Inc: Research Funding. Panopoulos:Amgen, Inc: Research Funding. Zhang:Amgen, Inc: Research Funding. Li:Amgen, Inc: Research Funding. Watowich:Amgen, Inc: Research Funding.

Zhang: Amgen: Research Funding. Nguyen-Jackson:Amgen: Research Funding. Watowich:Amgen, Inc: Research Funding.

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