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ABSTRACT: In this review we discuss how the competition for cytokines between different cells of the immune system can shape the system wide immune response. We focus on interleukin-2 (IL-2) secretion by activated effector T cells (T(eff)) and on the competition for IL-2 consumption between T(eff) and regulatory T cells (T(reg)). We discuss the evidence for the mechanism in which the depletion of IL-2 by T(reg) cells would be sufficient to suppress an autoimmune response, yet not strong enough to prevent an immune response. We present quantitative estimations and summarize our modeling effort to show that the tug-of-war between T(reg) and T(eff) cells for IL-2 molecules can be won by T(reg) cells in the case of weak activation of T(eff) leading to the suppression of the immune response. Or, for strongly activated T(eff) cells, it can be won by T(eff) cells bringing about the activation of the whole adaptive immune system. Finally, we discuss some recent applications attempting to achieve clinical effects through the modulation of IL-2 consumption by T(reg) compartment.
Frontiers in immunology. 01/2012; 3:268.
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ABSTRACT: Polarization of the T cell microtubule-organizing center (MTOC) toward the antigen-presenting cell (APC) is driven by the accumulation of diacylglycerol (DAG) at the immunological synapse (IS). The mechanisms that couple DAG to the MTOC are not known. By single-cell photoactivation of the T cell antigen receptor (TCR), we found that three distinct isoforms of protein kinase C (PKC) were recruited by DAG to the IS in two steps. PKC-ɛ and PKC-η accumulated first in a broad region of membrane, whereas PKC-θ arrived later in a smaller zone. Functional experiments indicated that PKC-θ was required for MTOC reorientation and that PKC-ɛ and PKC-η operated redundantly to promote the recruitment of PKC-θ and subsequent polarization responses. Our results establish a previously uncharacterized role for PKC proteins in T cell polarity.
Nature Immunology 01/2011; 12(7):647-54. · 26.01 Impact Factor
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ABSTRACT: Understanding how the immune system decides between tolerance and activation by antigens requires addressing cytokine regulation as a highly dynamic process. We quantified the dynamics of interleukin-2 (IL-2) signaling in a population of T cells during an immune response by combining in silico modeling and single-cell measurements in vitro. We demonstrate that IL-2 receptor expression levels vary widely among T cells creating a large variability in the ability of the individual cells to consume, produce and participate in IL-2 signaling within the population. Our model reveals that at the population level, these heterogeneous cells are engaged in a tug-of-war for IL-2 between regulatory (T(reg)) and effector (T(eff)) T cells, whereby access to IL-2 can either increase the survival of T(eff) cells or the suppressive capacity of T(reg) cells. This tug-of-war is the mechanism enforcing, at the systems level, a core function of T(reg) cells, namely the specific suppression of survival signals for weakly activated T(eff) cells but not for strongly activated cells. Our integrated model yields quantitative, experimentally validated predictions for the manipulation of T(reg) suppression.
Molecular Systems Biology 11/2010; 6:437. · 8.63 Impact Factor
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ABSTRACT: The T cell receptor (TCR) is responsible for discriminating between self- and foreign-derived peptides, translating minute differences in amino-acid sequence into large differences in response. Because of the great variability in the TCR and its ligands, activation of T cells by foreign peptides is a quantitative process, dependent on a mix of upstream signals and downstream integration. Accordingly, quantitative data and computational models have shed light on many important aspects of this process: molecular noise in ligand recognition, spatial dynamics in T cell-APC (antigen presenting cell) interactions, graded versus all-or-none decision making by the TCR apparatus, mechanisms of peptide antagonism and synergism, and the tunability and robustness of activation thresholds. Though diverse in their formalism, these studies together paint a picture of how modeling has shaped and will continue to shape understanding of T cell immunobiology.
Cold Spring Harbor perspectives in biology 06/2010; 2(6):a005538. · 9.40 Impact Factor
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ABSTRACT: Fluorescence correlation spectroscopy (FCS) is a single-molecule fluorescence technique used to monitor molecular dynamics. It can be applied to living cells expressing fluorescently labeled proteins and lipids to determine the diffusion timescales and the total number of diffusing fluorescent molecules in the cell. A practical step-by-step approach to performing FCS with a commercial spectroscopy/microscopy system, the Zeiss Confocor 3, how to set up live-cell FCS experiments, acquire reliable data, and finally how to analyze the data acquired, are all described in this unit.
Current protocols in cell biology / editorial board, Juan S. Bonifacino ... [et al.] 12/2009; Chapter 4:Unit 4.24.
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Gabrielle A Rizzuto,
Taha Merghoub,
Daniel Hirschhorn-Cymerman,
Cailian Liu,
Alexander M Lesokhin,
Diana Sahawneh,
Hong Zhong,
Katherine S Panageas,
Miguel-Angel Perales, Grégoire Altan-Bonnet,
Jedd D Wolchok,
Alan N Houghton
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ABSTRACT: A primary goal of cancer immunotherapy is to improve the naturally occurring, but weak, immune response to tumors. Ineffective responses to cancer vaccines may be caused, in part, by low numbers of self-reactive lymphocytes surviving negative selection. Here, we estimated the frequency of CD8(+) T cells recognizing a self-antigen to be <0.0001% ( approximately 1 in 1 million CD8(+) T cells), which is so low as to preclude a strong immune response in some mice. Supplementing this repertoire with naive antigen-specific cells increased vaccine-elicited tumor immunity and autoimmunity, but a threshold was reached whereby the transfer of increased numbers of antigen-specific cells impaired functional benefit, most likely because of intraclonal competition in the irradiated host. We show that cells primed at precursor frequencies below this competitive threshold proliferate more, acquire polyfunctionality, and eradicate tumors more effectively. This work demonstrates the functional relevance of CD8(+) T cell precursor frequency to tumor immunity and autoimmunity. Transferring optimized numbers of naive tumor-specific T cells, followed by in vivo activation, is a new approach that can be applied to human cancer immunotherapy. Further, precursor frequency as an isolated variable can be exploited to augment efficacy of clinical vaccine strategies designed to activate any antigen-specific CD8(+) T cells.
Journal of Experimental Medicine 04/2009; 206(4):849-66. · 13.85 Impact Factor
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ABSTRACT: In T cells, the stochasticity of protein expression could contribute to the useful diversification of biological functions within a clonal population or interfere with accurate antigen discrimination. Combining computer modeling and single-cell measurements, we examined how endogenous variation in the expression levels of signaling proteins might affect antigen responsiveness during T cell activation. We found that the CD8 co-receptor fine-tunes activation thresholds, whereas the soluble hematopoietic phosphatase 1 (SHP-1) digitally regulates cell responsiveness. Stochastic variation in the expression of these proteins generates substantial diversity of activation within a clonal population of T cells, but co-regulation of CD8 and SHP-1 levels ultimately limits this very diversity. These findings reveal how eukaryotic cells can draw on regulated variation in gene expression to achieve phenotypic variability in a controlled manner.
Science 09/2008; 321(5892):1081-4. · 31.20 Impact Factor
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Molecular Immunology 03/2008; 45(3):619-31. · 2.90 Impact Factor
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ABSTRACT: CD8+ T cells have a crucial role in resistance to pathogens and can kill malignant cells; however, some critical functions of these lymphocytes depend on helper activity provided by a distinct population of CD4+ T cells. Cooperation between these lymphocyte subsets involves recognition of antigens co-presented by the same dendritic cell, but the frequencies of such antigen-bearing cells early in an infection and of the relevant naive T cells are both low. This suggests that an active mechanism facilitates the necessary cell-cell associations. Here we demonstrate that after immunization but before antigen recognition, naive CD8+ T cells in immunogen-draining lymph nodes upregulate the chemokine receptor CCR5, permitting these cells to be attracted to sites of antigen-specific dendritic cell-CD4+ T cell interaction where the cognate chemokines CCL3 and CCL4 (also known as MIP-1alpha and MIP-1beta) are produced. Interference with this actively guided recruitment markedly reduces the ability of CD4+ T cells to promote memory CD8+ T-cell generation, indicating that an orchestrated series of differentiation events drives nonrandom cell-cell interactions within lymph nodes, optimizing CD8+ T-cell immune responses involving the few antigen-specific precursors present in the naive repertoire.
Nature 05/2006; 440(7086):890-5. · 36.28 Impact Factor
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ABSTRACT: T-lymphocyte activation displays a remarkable combination of speed, sensitivity, and discrimination in response to peptide-major histocompatibility complex (pMHC) ligand engagement of clonally distributed antigen receptors (T cell receptors or TCRs). Even a few foreign pMHCs on the surface of an antigen-presenting cell trigger effective signaling within seconds, whereas 1 x 10(5)-1 x 10(6) self-pMHC ligands that may differ from the foreign stimulus by only a single amino acid fail to elicit this response. No existing model accounts for this nearly absolute distinction between closely related TCR ligands while also preserving the other canonical features of T-cell responses. Here we document the unexpected highly amplified and digital nature of extracellular signal-regulated kinase (ERK) activation in T cells. Based on this observation and evidence that competing positive- and negative-feedback loops contribute to TCR ligand discrimination, we constructed a new mathematical model of proximal TCR-dependent signaling. The model made clear that competition between a digital positive feedback based on ERK activity and an analog negative feedback involving SH2 domain-containing tyrosine phosphatase (SHP-1) was critical for defining a sharp ligand-discrimination threshold while preserving a rapid and sensitive response. Several nontrivial predictions of this model, including the notion that this threshold is highly sensitive to small changes in SHP-1 expression levels during cellular differentiation, were confirmed by experiment. These results combining computation and experiment reveal that ligand discrimination by T cells is controlled by the dynamics of competing feedback loops that regulate a high-gain digital amplifier, which is itself modulated during differentiation by alterations in the intracellular concentrations of key enzymes. The organization of the signaling network that we model here may be a prototypic solution to the problem of achieving ligand selectivity, low noise, and high sensitivity in biological responses.
PLoS Biology 12/2005; 3(11):e356. · 11.45 Impact Factor
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ABSTRACT: In order to discern aggregation in solution, we present a quantum mechanical analog of the photon statistics from fluorescent molecules diffusing through a focused beam. A generating functional is developed to fully describe the experimental physical system as well as the statistics. Histograms of the measured time delay between photon counts are fit by an analytical solution describing the static as well as diffusing regimes. To determine empirical fitting parameters, fluorescence correlation spectroscopy is used in parallel to the photon counting. For expedient analysis, we find that the distribution's deviation from a single Poisson shows a difference between two single fluor monomers or a double fluor aggregate of the same total intensities. Initial studies were performed on fixed-state aggregates limited to dimerization. However preliminary results on reactive species suggest that the method can be used to characterize any aggregating system.
Physical Review E 06/2004; 69(5 Pt 1):051916. · 2.26 Impact Factor
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ABSTRACT: We report the first measurement of the dynamics of bubble formation in double-stranded DNA. Fluctuations of fluorescence of a synthetic DNA construct, internally tagged with a fluorophore and a quencher, are monitored by fluorescence correlation spectroscopy. The relaxation dynamics follow a multistate relaxation kinetics, with a characteristic time scale of 50 microseconds. A simple model of bubble dynamics based on constant zipping-unzipping rates is proposed to account for our experimental data. The role of different secondary structures stabilizing the open bubble is tested.
Physical Review Letters 05/2003; 90(13):138101. · 7.37 Impact Factor
