ABSTRACT
The regulation of neutrophil lifespan is critical for a circumscribed immune response. Neutrophils are sensitive to Fas/CD95 death receptor signaling in vitro, but it is unknown if Fas regulates neutrophil lifespan in vivo. We hypothesized that FasL-expressing CD8(+) T cells, which kill antigen-stimulated T cells during chronic viral infection, can also induce neutrophil death in tissues during infection. With the use of LysM-Cre Fas(fl/fl) mice, which lack Fas expression in macrophages and neutrophils, we show that Fas regulates neutrophil lifespan during lymphocytic choriomeningitis virus (LCMV) infection in the lung, peripheral blood, and spleen. Fas also contributed to the regulation of neutrophil numbers in the colon of Citrobacter rodentium-infected mice. To examine the effects of infection on Fas activation in neutrophils, we primed neutrophils with TLR ligands or IL-18, resulting in ablation of Fas death receptor signaling. These data provide the first in vivo genetic evidence that neutrophil lifespan is controlled by death receptor signaling and provide a mechanism to account for neutrophil resistance to Fas stimulation during infection.
Subject(s)
Cellular Senescence/immunology , Citrobacter rodentium/immunology , Enterobacteriaceae Infections/immunology , Gene Expression Regulation/immunology , Lymphocytic Choriomeningitis/immunology , Lymphocytic choriomeningitis virus/immunology , Neutrophils/immunology , fas Receptor/immunology , Animals , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/pathology , Cellular Senescence/genetics , Enterobacteriaceae Infections/genetics , Enterobacteriaceae Infections/pathology , Fas Ligand Protein/genetics , Fas Ligand Protein/immunology , Gene Expression Regulation/genetics , Interleukin-18/genetics , Interleukin-18/immunology , Lymphocytic Choriomeningitis/genetics , Lymphocytic Choriomeningitis/pathology , Macrophages/immunology , Macrophages/pathology , Mice , Mice, Knockout , Neutrophils/pathology , Signal Transduction/genetics , Signal Transduction/immunology , fas Receptor/geneticsABSTRACT
Homeotic (HOX) genes are dysregulated in multiple malignancies, including several AML subtypes. We demonstrate that H3K79 dimethylation (H3K79me2) is converted to monomethylation (H3K79me1) at HOX loci as hematopoietic cells mature, thus coinciding with a decrease in HOX gene expression. We show that H3K79 methyltransferase activity as well as H3K79me1-to-H3K79me2 conversion is regulated by the DOT1L cofactor AF10. AF10 inactivation reverses leukemia-associated epigenetic profiles, precludes abnormal HOXA gene expression, and impairs the transforming ability of MLL-AF9, MLL-AF6, and NUP98-NSD1 fusions-mechanistically distinct HOX-activating oncogenes. Furthermore, NUP98-NSD1-transformed cells are sensitive to small-molecule inhibition of DOT1L. Our findings demonstrate that pharmacological inhibition of the DOT1L/AF10 complex may provide therapeutic benefits in an array of malignancies with abnormal HOXA gene expression.
Subject(s)
Histones/metabolism , Homeodomain Proteins/metabolism , Leukemia, Myeloid, Acute/metabolism , Leukemia, Myeloid, Acute/pathology , Methyltransferases/metabolism , Transcription Factors/metabolism , Adenosine/analogs & derivatives , Adenosine/pharmacology , Animals , Bone Marrow Cells/metabolism , Epigenesis, Genetic , Gene Expression Regulation, Neoplastic/drug effects , HL-60 Cells , Humans , Methylation , Methyltransferases/antagonists & inhibitors , Mice , Mice, Transgenic , Molecular Sequence Data , Neoplasms, Experimental , Nuclear Pore Complex Proteins/metabolism , Nuclear Proteins/metabolism , Oncogene Proteins, Fusion/metabolism , Phenylurea Compounds/pharmacologySubject(s)
Apoptosis/drug effects , Cytokines/metabolism , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Mesenchymal Stem Cells/cytology , Proto-Oncogene Proteins c-kit/antagonists & inhibitors , Proto-Oncogene Proteins c-kit/genetics , Drug Resistance, Neoplasm , Humans , Leukemia, Myeloid, Acute/pathology , Tumor Cells, CulturedABSTRACT
The t(6;11)(q27;q23) is a recurrent chromosomal rearrangement that encodes the MLLAF6 fusion oncoprotein and is observed in patients with diverse hematologic malignancies. The presence of the t(6;11)(q27;q23) has been linked to poor overall survival in patients with AML. In this study, we demonstrate that MLL-AF6 requires continued activity of the histone-methyltransferase DOT1L to maintain expression of the MLL-AF6-driven oncogenic gene-expression program. Using gene-expression analysis and genome-wide chromatin immunoprecipitation studies followed by next generation sequencing, we found that MLL-fusion target genes display markedly high levels of histone 3 at lysine 79 (H3K79) dimethylation in murine MLL-AF6 leukemias as well as in ML2, a human myelomonocytic leukemia cell line bearing the t(6;11)(q27;q23) translocation. Targeted disruption of Dot1l using a conditional knockout mouse model inhibited leukemogenesis mediated by the MLL-AF6 fusion oncogene. Moreover, both murine MLL-AF6-transformed cells as well as the human MLL-AF6-positive ML2 leukemia cell line displayed specific sensitivity to EPZ0004777, a recently described, selective, small-molecule inhibitor of Dot1l. Dot1l inhibition resulted in significantly decreased proliferation, decreased expression of MLL-AF6 target genes, and cell cycle arrest of MLL-AF6-transformed cells. These results indicate that patients bearing the t(6;11)(q27;q23) translocation may benefit from therapeutic agents targeting aberrant H3K79 methylation.
