ABSTRACT
CD8+ T cells play a critical role in adaptive immunity, differentiating into CD8+ memory T cells that form the basis of protective cellular immunity. Vaccine efficacy is attributed to long-term protective immunity, and understanding the parameters that regulate development of CD8+ T cells is critical to the design of T cell-mediated vaccines. We show in this study using mouse models that two distinct parameters, TCR signal strength (regulated by the tyrosine kinase ITK) and Ag affinity, play important but separate roles in modulating the development of memory CD8+ T cells. Unexpectedly, our data reveal that reducing TCR signal strength along with reducing Ag affinity for the TCR leads to enhanced and accelerated development of CD8+ memory T cells. Additionally, TCR signal strength is able to regulate CD8+ T cell effector cytokine R production independent of TCR Ag affinity. Analysis of RNA-sequencing data reveals that genes for inflammatory cytokines/cytokine receptors are significantly altered upon changes in Ag affinity and TCR signal strength. Furthermore, our findings show that the inflammatory milieu is critical in regulating this TCR signal strength-mediated increase in memory development, as both CpG oligonucleotide treatment or cotransfer of wild-type and Itk-/- T cells eliminates the observed increase in memory cell formation. These findings suggest that TCR signal strength and Ag affinity independently contribute to CD8+ memory T cell development, which is modulated by inflammation, and suggest that manipulating TCR signal strength along with Ag affinity, may be used to tune the development of CD8+ memory T cells during vaccine development.
Subject(s)
Antigens/immunology , CD8-Positive T-Lymphocytes/immunology , Immunologic Memory/immunology , Receptors, Antigen, T-Cell/immunology , Signal Transduction/immunology , Animals , Cell Differentiation/immunology , Cytokines/immunology , Female , Inflammation/immunology , Male , Mice , Mice, Inbred C57BL , Protein-Tyrosine Kinases/immunologyABSTRACT
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
Specific reduction in the intake of proteins or amino acids (AAs) offers enormous health benefits, including increased life span, protection against age-associated disorders, and improved metabolic fitness and immunity. Cells respond to conditions of AA starvation by activating the amino acid starvation response (AAR). Here, we showed that mimicking AAR with halofuginone (HF) enhanced the magnitude and affinity of neutralizing, antigen-specific antibody responses in mice immunized with dengue virus envelope domain III protein (DENVrEDIII), a potent vaccine candidate against DENV. HF enhanced the formation of germinal centers (GCs) and increased the production of the cytokine IL-10 in the secondary lymphoid organs of vaccinated mice. Furthermore, HF promoted the transcription of genes associated with memory B cell formation and maintenance and maturation of GCs in the draining lymph nodes of vaccinated mice. The increased abundance of IL-10 in HF-preconditioned mice correlated with enhanced GC responses and may promote the establishment of long-lived plasma cells that secrete antigen-specific, high-affinity antibodies. Thus, these data suggest that mimetics of AA starvation could provide an alternative strategy to augment the efficacy of vaccines against dengue and other infectious diseases.
Subject(s)
Amino Acids/deficiency , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antibody Formation , Dengue Vaccines/pharmacology , Animals , Interleukin-10/immunology , Mice , Mice, Inbred BALB C , Piperidines/pharmacology , Quinazolinones/pharmacologyABSTRACT
Autoimmune diseases are characterized by impaired immune tolerance towards self-antigens, leading to enhanced immunity to self by dysfunctional B cells and/or T cells. The activation of these cells is controlled by non-receptor tyrosine kinases (NRTKs), which are critical mediators of antigen receptor and cytokine receptor signaling pathways. NRTKs transduce, amplify and sustain activating signals that contribute to autoimmunity, and are counter-regulated by protein tyrosine phosphatases (PTPs). The function of and interaction between NRTKs and PTPs during the development of autoimmunity could be key points of therapeutic interference against autoimmune diseases. In this review, we summarize the current state of knowledge of the functions of NRTKs and PTPs involved in B cell receptor (BCR), T cell receptor (TCR), and cytokine receptor signaling pathways that contribute to autoimmunity, and discuss their targeting for therapeutic approaches against autoimmune diseases.
Subject(s)
Autoimmune Diseases/immunology , Protein Tyrosine Phosphatases/metabolism , Protein-Tyrosine Kinases/metabolism , Animals , Autoimmune Diseases/enzymology , Autoimmune Diseases/therapy , Autoimmunity/immunology , Humans , Immune Tolerance/immunology , Receptors, Antigen, B-Cell/immunology , Receptors, Antigen, T-Cell/immunology , Receptors, Cytokine/immunology , Signal Transduction/immunologyABSTRACT
Type 1 regulatory CD4+ T (Tr1) cells express high levels of the immunosuppressive cytokine IL-10 but not the master transcription factor Foxp3, and can suppress inflammation and promote immune tolerance. In order to identify and obtain viable Tr1 cells for research and clinical applications, co-expression of CD49b and LAG3 has been proposed as a unique surface signature for both human and mouse Tr1 cells. However, recent studies have revealed that this pattern of co-expression is dependent on the stimulating conditions and the differentiation stage of the CD4+ T cells. Here, using an IL-10GFP/Foxp3RFP dual reporter transgenic murine model, we demonstrate that co-expression of CD49b and LAG3 is not restricted to the Foxp3- Tr1 cells, but is also observed in Foxp3+ T regulatory (Treg) cells and CD8+ T cells that produce IL-10. Our data indicate that IL-10-producing Tr1 cells, Treg cells and CD8+ T cells are all capable of co-expressing LAG3 and CD49b in vitro following differentiation under IL-10-inducing conditions, and in vivo following pathogenic insult or infection in the pulmonary mucosa. Our findings urge caution in the use of LAG3/CD49b co-expression as sole markers to identify Tr1 cells, since it may mark IL-10-producing T cell lineages more broadly, including the Foxp3- Tr1 cells, Foxp3+ Treg cells, and CD8+ T cells.
Subject(s)
Antigens, CD/metabolism , CD8-Positive T-Lymphocytes/metabolism , Integrin alpha2/metabolism , Interleukin-10/metabolism , T-Lymphocytes, Regulatory/metabolism , Animals , Biomarkers/metabolism , CD8-Positive T-Lymphocytes/immunology , Cell Differentiation , Cell Lineage , Cells, Cultured , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/metabolism , Humans , Immune Tolerance , Interleukin-10/genetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , T-Lymphocytes, Regulatory/immunology , Lymphocyte Activation Gene 3 ProteinABSTRACT
Type 1 regulatory T (Tr1) cells differentiate in response to signals engaging the T cell receptor (TCR), express high levels of the immunosuppressive cytokine IL-10, but not Foxp3, and can suppress inflammation and promote immune tolerance. Here we show that ITK, an important modulator of TCR signalling, is required for the TCR-induced development of Tr1 cells in various organs, and in the mucosal system during parasitic and viral infections. ITK kinase activity is required for mouse and human Tr1 cell differentiation. Tr1 cell development and suppressive function of Itk deficient cells can be restored by the expression of the transcription factor interferon regulatory factor 4 (IRF4). Downstream of ITK, Ras activity is responsible for Tr1 cell induction, as expression of constitutively active HRas rescues IRF4 expression and Tr1 cell differentiation in Itk-/- cells. We conclude that TCR/ITK signalling through the Ras/IRF4 pathway is required for functional development of Tr1 cells.
Subject(s)
Interferon Regulatory Factors/metabolism , Protein-Tyrosine Kinases/metabolism , T-Lymphocytes, Regulatory/physiology , ras Proteins/metabolism , Animals , Cell Differentiation , Humans , Interferon Regulatory Factors/genetics , Mice, Inbred C57BL , Mice, Transgenic , Orthomyxoviridae Infections/immunology , Positive Regulatory Domain I-Binding Factor 1/metabolism , Protein-Tyrosine Kinases/genetics , Signal Transduction , Strongylida Infections/immunology , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/metabolism , ras Proteins/geneticsABSTRACT
8E5/LAV cells harbor a single HIV provirus, and are used frequently to generate standards for HIV genome quantification. Using flow cytometry-based in situ mRNA hybridization validated by qPCR, we find that different batches of 8E5 cells contain varying numbers of cells lacking viral mRNA and/or viral genomes. These findings raise concerns for studies employing 8E5 cells for quantitation, and highlight the value of mRNA FISH and flow cytometry in the detection and enumeration of HIV-positive cells.