title: TCR signaling intensity controls CD8+ T cell responsiveness to TGF-β.
abstract: DGK-ζ is a negative regulator of TCR signaling that causes degradation of the second messenger DAG, terminating DAG-mediated activation of Ras and PKCθ. Cytotoxic T cells deficient in DGK-ζ demonstrate enhanced effector functions in vitro and antitumor activity in vivo, perhaps because of insensitivity to inhibitory cytokines. We sought to determine whether the enhanced responsiveness of DGK-ζ-deficient T cells renders them insensitive to the inhibitory cytokine TGF-β and to determine how the loss of DGK-ζ facilitates this insensitivity. We identified decreased transcriptional and functional responses to TGF-β in CD8(+) DGK-ζ(-/-) T cells but preserved TGF-β-mediated conversion of naïve DGK-ζ(-/-) CD4(+) T cells to a regulatory T cell phenotype. Decreased CD8(+) T cell responsiveness to TGF-β did not result from impaired canonical TGF-β signal transduction, because similar levels of TGF-β-R and intracellular Smad components were identified in WT and DGK-ζ(-/-) CD8(+) T cells, and TGF-β-mediated activation of Smad2 was unchanged. Instead, an enhanced TCR signal strength was responsible for TGF-β insensitivity, because (i) loss of DGK-ζ conferred resistance to TGF-β-mediated inhibition of Erk phosphorylation, (ii) TGF-β insensitivity could be recapitulated by exogenous addition of the DAG analog PMA, and (iii) TGF-β sensitivity could be observed in DGK-ζ-deficient T cells at limiting dilutions of TCR stimulation. These data indicate that enhanced TCR signal transduction in the absence of DGK-ζ makes T cells relatively insensitive to TGF-β, in a manner independent of Smads, a finding with practical implications in the development of immunotherapies that target TGF-β.
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Vidhyalakshmi
authors.initials: V
authors.lastname: Arumugam
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Theresa
authors.initials: T
authors.lastname: Bluemn
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Erin
authors.initials: E
authors.lastname: Wesley
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Amanda M
authors.initials: AM
authors.lastname: Schmidt
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Taku
authors.initials: T
authors.lastname: Kambayashi
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
authors.forename: Subramaniam
authors.initials: S
authors.lastname: Malarkannan
authors.affiliation: *Blood Research Institute, Department of Microbiology and Molecular Genetics, and Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin, USA; and Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA mriese@mcw.edu.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Riese
mesh_terms: D000818:Animals; D018414:CD8-Positive T-Lymphocytes / Q000166:cytology / Q000276:immunology*; D019852:Diacylglycerol Kinase / Q000235:genetics / Q000276:immunology*; D051379:Mice; D018345:Mice, Knockout; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology*; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology*; D051899:Smad2 Protein / Q000235:genetics / Q000276:immunology; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000276:immunology*
chemical_list: D011948:Receptors, Antigen, T-Cell; D051899:Smad2 Protein; C497783:Smad2 protein, mouse; D016212:Transforming Growth Factor beta; D019852:Diacylglycerol Kinase; C512480:diacylglycerol kinase zeta, mouse
keywords: Diacylglycerol; Smad2; diacylglycerol kinase ζ