The signaling symphony: T cell receptor tunes cytokine-mediated T cell differentiation

PubMed Central

Huang, Weishan; August, Avery

2015-01-01

T cell development, differentiation, and maintenance are orchestrated by 2 key signaling axes: the antigen-specific TCR and cytokine-mediated signals. The TCR signals the recognition of self- and foreign antigens to control T cell homeostasis for immune tolerance and immunity, which is regulated by a variety of cytokines to determine T cell subset homeostasis and differentiation. TCR signaling can synergize with or antagonize cytokine-mediated signaling to fine tune T cell fate; however, the latter is less investigated. Murine models with attenuated TCR signaling strength have revealed that TCR signaling can function as regulatory feedback machinery for T cell homeostasis and differentiation in differential cytokine milieus, such as IL-2-mediated Treg development; IL-7-mediated, naïve CD8+ T cell homeostasis; and IL-4-induced innate memory CD8+ T cell development. In this review, we discuss the symphonic cross-talk between TCR and cytokine-mediated responses that differentially control T cell behavior, with a focus on the negative tuning by TCR activation on the cytokine effects. PMID:25525115

Inhibition of Gαs/cAMP Signaling Decreases TCR-Stimulated IL-2 transcription in CD4(+) T Helper Cells.

PubMed

Hynes, Thomas R; Yost, Evan A; Yost, Stacy M; Hartle, Cassandra M; Ott, Braden J; Berlot, Catherine H

2015-07-06

The role of cAMP in regulating T cell activation and function has been controversial. cAMP is generally known as an immunosuppressant, but it is also required for generating optimal immune responses. As the effect of cAMP is likely to depend on its cellular context, the current study investigated whether the mechanism of activation of Gαs and adenylyl cyclase influences their effect on T cell receptor (TCR)-stimulated interleukin-2 (IL-2) mRNA levels. The effect of blocking Gs-coupled receptor (GsPCR)-mediated Gs activation on TCR-stimulated IL-2 mRNA levels in CD4(+) T cells was compared with that of knocking down Gαs expression or inhibiting adenylyl cyclase activity. The effect of knocking down Gαs expression on TCR-stimulated cAMP accumulation was compared with that of blocking GsPCR signaling. ZM-241385, an antagonist to the Gs-coupled A2A adenosine receptor (A2AR), enhanced TCR-stimulated IL-2 mRNA levels in primary human CD4(+) T helper cells and in Jurkat T cells. A dominant negative Gαs construct, GαsDN3, also enhanced TCR-stimulated IL-2 mRNA levels. Similar to GsPCR antagonists, GαsDN3 blocked GsPCR-dependent activation of both Gαs and Gβγ. In contrast, Gαs siRNA and 2',5'-dideoxyadenosine (ddA), an adenylyl cyclase inhibitor, decreased TCR-stimulated IL-2 mRNA levels. Gαs siRNA, but not GαsDN3, decreased TCR-stimulated cAMP synthesis. Potentiation of IL-2 mRNA levels by ZM-241385 required at least two days of TCR stimulation, and addition of ddA after three days of TCR stimulation enhanced IL-2 mRNA levels. GsPCRs play an inhibitory role in the regulation of TCR-stimulated IL-2 mRNA levels whereas Gαs and cAMP can play a stimulatory one. Additionally, TCR-dependent activation of Gαs does not appear to involve GsPCRs. These results suggest that the context of Gαs/cAMP activation and the stage of T cell activation and differentiation determine the effect on TCR-stimulated IL-2 mRNA levels.

Atomic structure of an alphabeta T cell receptor (TCR) heterodimer in complex with an anti-TCR fab fragment derived from a mitogenic antibody.

PubMed Central

Wang, J; Lim, K; Smolyar, A; Teng, M; Liu, J; Tse, A G; Liu, J; Hussey, R E; Chishti, Y; Thomson, C T; Sweet, R M; Nathenson, S G; Chang, H C; Sacchettini, J C; Reinherz, E L

1998-01-01

Each T cell receptor (TCR) recognizes a peptide antigen bound to a major histocompatibility complex (MHC) molecule via a clonotypic alphabeta heterodimeric structure (Ti) non-covalently associated with the monomorphic CD3 signaling components. A crystal structure of an alphabeta TCR-anti-TCR Fab complex shows an Fab fragment derived from the H57 monoclonal antibody (mAb), interacting with the elongated FG loop of the Cbeta domain, situated beneath the Vbeta domain. This loop, along with the partially exposed ABED beta sheet of Cbeta, and glycans attached to both Cbeta and Calpha domains, forms a cavity of sufficient size to accommodate a single non-glycosylated Ig domain such as the CD3epsilon ectodomain. That this asymmetrically localized site is embedded within the rigid constant domain module has implications for the mechanism of signal transduction in both TCR and pre-TCR complexes. Furthermore, quaternary structures of TCRs vary significantly even when they bind the same MHC molecule, as manifested by a unique twisting of the V module relative to the C module. PMID:9427737

T-bet Down-Modulation in Tolerized Th1 Effector CD4 Cells Confers a TCR-Distal Signaling Defect That Selectively Impairs IFN-γ Expression1

PubMed Central

Long, Meixiao; Slaiby, Aaron M.; Hagymasi, Adam T.; Mihalyo, Marianne A.; Lichtler, Alexander C.; Reiner, Steven L.; Adler, Adam J.

2010-01-01

When Th1 effector CD4 cells encounter tolerizing Ag in vivo, their capacity to express the effector cytokines IFN-γ and TNF-α is lost more rapidly than noneffector functions such as IL-2 production and proliferation. To localize the relevant intracellular signaling defects, cytokine expression was compared following restimulation with Ag vs agents that bypass TCR-proximal signaling. IFN-γ and TNF-α expression were both partially rescued when TCR-proximal signaling was bypassed, indicating that both TCR-proximal and -distal signaling defects impair the expression of these two effector cytokines. In contrast, bypassing TCR-proximal signaling fully rescued IL-2 expression. T-bet, a transcription and chromatin remodeling factor that is required to direct the differentiation of naive CD4 cells into IFN-γ -expressing Th1 effectors, was partially down-modulated in tolerized Th1 effectors. Enforcing T-bet expression during tolerization selectively rescued the ability to express IFN-γ, but not TNF-α. Conversely, expression of a dominant-negative T-bet in Th1 effectors selectively impaired the ability to express IFN-γ, but not TNF-α. Analysis of histone acetylation at the IFN-γ promoter further suggested that down-modulation of T-bet expression during Th1 effector CD4 cell tolerization does not impair IFN-γ expression potential through alterations in chromatin structure. PMID:16393991

Pontin is required for pre-TCR signaling at the β-selection checkpoint in T cell development.

PubMed

Boo, Kyungjin; Baek, Sung Hee; Lee, Ho

2014-04-25

Pontin is a chromatin remodeling factor that possesses both ATPase and DNA helicase activities. Based on high expression in lymphoid tissues, we examined whether Pontin has a T cell-specific function. We generated Pontin(f/f);Lck-Cre mice, in which Pontin can be conditionally deleted in T cells and then explored T cell-specific function of Pontin in vivo. Here, we show that specific abrogation of Pontin expression in T cells almost completely blocked development of αβ T cells at the β-selection checkpoint by inducing cell apoptosis indicating that Pontin is essential for early T cell development. Pontin-deficient thymocytes show a comparable expression level of T cell receptor (TCR)β chain, but have enhanced activation of p53 and Notch signaling compared to wild-type thymocytes. Intriguingly, the developmental block of αβ T cells can be partially rescued by loss of p53. Together, our data demonstrate a novel role of Pontin as a crucial regulator in pre-TCR signaling during T cell development. Copyright © 2014 Elsevier Inc. All rights reserved.

Diacylglycerol kinase α inactivation is an integral component of the costimulatory pathway that amplifies TCR signals.

PubMed

Arranz-Nicolás, Javier; Ogando, Jesús; Soutar, Denise; Arcos-Pérez, Raquel; Meraviglia-Crivelli, Daniel; Mañes, Santos; Mérida, Isabel; Ávila-Flores, Antonia

2018-06-01

The arsenal of cancer therapies has evolved to target T lymphocytes and restore their capacity to destroy tumor cells. T cells rely on diacylglycerol (DAG) to carry out their functions. DAG availability and signaling are regulated by the enzymes diacylglycerol kinase (DGK) α and ζ, whose excess function drives T cells into hyporesponsive states. Targeting DGKα is a promising strategy for coping with cancer; its blockade could reinstate T-cell attack on tumors while limiting tumor growth, due to positive DGKα functions in several oncogenic pathways. Here, we made a side-by-side comparison of the effects of commercial pharmacological DGK inhibitors on T-cell responses with those promoted by DGKα and DGKζ genetic deletion or silencing. We show the specificity for DGKα of DGK inhibitors I and II and the structurally similar compound ritanserin. Inhibitor treatment promoted Ras/ERK (extracellular signal-regulated kinase) signaling and AP-1 (Activator protein-1) transcription, facilitated DGKα membrane localization, reduced the requirement for costimulation, and cooperated with enhanced activation following DGKζ silencing/deletion. DGKiII and ritanserin had similar effects on TCR proximal signaling, but ritanserin counteracted long-term T-cell activation, an effect that was potentiated in DGKα -/- cells. In contrast with enhanced activation triggered by pharmacological inhibition, DGKα silencing/genetic deletion led to impaired Lck (lymphocyte-specific protein tyrosine kinase) activation and limited costimulation responses. Our results demonstrate that pharmacological inhibition of DGKα downstream of the TCR provides a gain-of-function effect that amplifies the DAG-dependent signaling cascade, an ability that could be exploited therapeutically to reinvigorate T cells to attack tumors.

Domain-swapped T cell receptors improve the safety of TCR gene therapy

PubMed Central

Bethune, Michael T; Gee, Marvin H; Bunse, Mario; Lee, Mark S; Gschweng, Eric H; Pagadala, Meghana S; Zhou, Jing; Cheng, Donghui; Heath, James R; Kohn, Donald B; Kuhns, Michael S; Uckert, Wolfgang; Baltimore, David

2016-01-01

T cells engineered to express a tumor-specific αβ T cell receptor (TCR) mediate anti-tumor immunity. However, mispairing of the therapeutic αβ chains with endogenous αβ chains reduces therapeutic TCR surface expression and generates self-reactive TCRs. We report a general strategy to prevent TCR mispairing: swapping constant domains between the α and β chains of a therapeutic TCR. When paired, domain-swapped (ds)TCRs assemble with CD3, express on the cell surface, and mediate antigen-specific T cell responses. By contrast, dsTCR chains mispaired with endogenous chains cannot properly assemble with CD3 or signal, preventing autoimmunity. We validate this approach in cell-based assays and in a mouse model of TCR gene transfer-induced graft-versus-host disease. We also validate a related approach whereby replacement of αβ TCR domains with corresponding γδ TCR domains yields a functional TCR that does not mispair. This work enables the design of safer TCR gene therapies for cancer immunotherapy. DOI: http://dx.doi.org/10.7554/eLife.19095.001 PMID:27823582

Pak2 is required for actin cytoskeleton remodeling, TCR signaling, and normal thymocyte development and maturation

PubMed Central

Phee, Hyewon; Au-Yeung, Byron B; Pryshchep, Olga; O'Hagan, Kyle Leonard; Fairbairn, Stephanie Grace; Radu, Maria; Kosoff, Rachelle; Mollenauer, Marianne; Cheng, Debra; Chernoff, Jonathan; Weiss, Arthur

2014-01-01

The molecular mechanisms that govern thymocyte development and maturation are incompletely understood. The P21-activated kinase 2 (Pak2) is an effector for the Rho family GTPases Rac and Cdc42 that regulate actin cytoskeletal remodeling, but its role in the immune system remains poorly understood. In this study, we show that T-cell specific deletion of Pak2 gene in mice resulted in severe T cell lymphopenia accompanied by marked defects in development, maturation, and egress of thymocytes. Pak2 was required for pre-TCR β-selection and positive selection. Surprisingly, Pak2 deficiency in CD4 single positive thymocytes prevented functional maturation and reduced expression of S1P1 and KLF2. Mechanistically, Pak2 is required for actin cytoskeletal remodeling triggered by TCR. Failure to induce proper actin cytoskeletal remodeling impaired PLCγ1 and Erk1/2 signaling in the absence of Pak2, uncovering the critical function of Pak2 as an essential regulator that governs the actin cytoskeleton-dependent signaling to ensure normal thymocyte development and maturation. DOI: http://dx.doi.org/10.7554/eLife.02270.001 PMID:24843022

Regulation of TCR Signaling to NF-kB

DTIC Science & Technology

2013-03-20

applications and benefits to basic research are discussed in detail in Chapter 4.      16   CHAPTER 2: Selective autophagy of the...To confirm the specificity of the chemical inhibitors of autophagy, we employed a genetic approach. Specifically, we in vitro differentiated Th2...MG132 treatment. These data provide strong genetic evidence that TCR-mediated degradation of endogenous Bcl10 is controlled by autophagy, with a

Association of CD147 and Calcium Exporter PMCA4 Uncouples IL-2 Expression from Early TCR Signaling.

PubMed

Supper, Verena; Schiller, Herbert B; Paster, Wolfgang; Forster, Florian; Boulègue, Cyril; Mitulovic, Goran; Leksa, Vladimir; Ohradanova-Repic, Anna; Machacek, Christian; Schatzlmaier, Philipp; Zlabinger, Gerhard J; Stockinger, Hannes

2016-02-01

The Ig superfamily member CD147 is upregulated following T cell activation and was shown to serve as a negative regulator of T cell proliferation. Thus, Abs targeting CD147 are being tested as new treatment strategies for cancer and autoimmune diseases. How CD147 mediates immunosuppression and whether association with other coreceptor complexes is needed have remained unknown. In the current study, we show that silencing of CD147 in human T cells increases IL-2 production without affecting the TCR proximal signaling components. We mapped the immunosuppressive moieties of CD147 to its transmembrane domain and Ig-like domain II. Using affinity purification combined with mass spectrometry, we determined the domain specificity of CD147 interaction partners and identified the calcium exporter plasma membrane calcium ATPase isoform 4 (PMCA4) as the interaction partner of the immunosuppressive moieties of CD147. CD147 does not control the proper membrane localization of PMCA4, but PMCA4 is essential for the CD147-dependent inhibition of IL-2 expression via a calcium-independent mechanism. In summary, our data show that CD147 interacts via its immunomodulatory domains with PMCA4 to bypass TCR proximal signaling and inhibit IL-2 expression. Copyright © 2016 by The American Association of Immunologists, Inc.

Src homology 2-domain containing leukocyte-specific phosphoprotein of 76 kDa is mandatory for TCR-mediated inside-out signaling, but dispensable for CXCR4-mediated LFA-1 activation, adhesion, and migration of T cells.

PubMed

Horn, Jessica; Wang, Xiaoqian; Reichardt, Peter; Stradal, Theresia E; Warnecke, Nicole; Simeoni, Luca; Gunzer, Matthias; Yablonski, Deborah; Schraven, Burkhart; Kliche, Stefanie

2009-11-01

Engagement of the TCR or of chemokine receptors such as CXCR4 induces adhesion and migration of T cells via so-called inside-out signaling pathways. The molecular processes underlying inside-out signaling events are as yet not completely understood. In this study, we show that TCR- and CXCR4-mediated activation of integrins critically depends on the membrane recruitment of the adhesion- and degranulation-promoting adapter protein (ADAP)/Src kinase-associated phosphoprotein of 55 kDa (SKAP55)/Rap1-interacting adapter protein (RIAM)/Rap1 module. We further demonstrate that the Src homology 2 domain containing leukocyte-specific phosphoprotein of 76 kDa (SLP76) is crucial for TCR-mediated inside-out signaling and T cell/APC interaction. Besides facilitating membrane recruitment of ADAP, SKAP55, and RIAM, SLP76 regulates TCR-mediated inside-out signaling by controlling the activation of Rap1 as well as Rac-mediated actin polymerization. Surprisingly, however, SLP76 is not mandatory for CXCR4-mediated inside-out signaling. Indeed, both CXCR4-induced T cell adhesion and migration are not affected by loss of SLP76. Moreover, after CXCR4 stimulation, the ADAP/SKAP55/RIAM/Rap1 module is recruited to the plasma membrane independently of SLP76. Collectively, our data indicate a differential requirement for SLP76 in TCR- vs CXCR4-mediated inside-out signaling pathways regulating T cell adhesion and migration.

IFN-γ regulates CD8+ memory T cell differentiation and survival in response to weak, but not strong, TCR signals.

PubMed

Stoycheva, Diana; Deiser, Katrin; Stärck, Lilian; Nishanth, Gopala; Schlüter, Dirk; Uckert, Wolfgang; Schüler, Thomas

2015-01-15

In response to primary Ag contact, naive mouse CD8(+) T cells undergo clonal expansion and differentiate into effector T cells. After pathogen clearance, most effector T cells die, and only a small number of memory T cell precursors (TMPs) survive to form a pool of long-lived memory T cells (TMs). Although high- and low-affinity CD8(+) T cell clones are recruited into the primary response, the TM pool consists mainly of high-affinity clones. It remains unclear whether the more efficient expansion of high-affinity clones and/or cell-intrinsic processes exclude low-affinity T cells from the TM pool. In this article, we show that the lack of IFN-γR signaling in CD8(+) T cells promotes TM formation in response to weak, but not strong, TCR agonists. The IFN-γ-sensitive accumulation of TMs correlates with reduced mammalian target of rapamycin activation and the accumulation of long-lived CD62L(hi)Bcl-2(hi)Eomes(hi) TMPs. Reconstitution of mammalian target of rapamycin or IFN-γR signaling is sufficient to block this process. Hence, our data suggest that IFN-γR signaling actively blocks the formation of TMPs responding to weak TCR agonists, thereby promoting the accumulation of high-affinity T cells finally dominating the TM pool. Copyright © 2015 by The American Association of Immunologists, Inc.

TCR revision generates functional CD4+ T cells.

PubMed

Hale, J Scott; Wubeshet, Maramawit; Fink, Pamela J

2010-12-01

CD4(+)Vβ5(+) peripheral T cells in C57BL/6 mice respond to encounter with a peripherally expressed endogenous superantigen by undergoing either deletion or TCR revision. In this latter process, cells lose surface Vβ5 expression and undergo RAG-dependent rearrangement of endogenous TCRβ genes, driving surface expression of novel TCRs. Although postrevision CD4(+)Vβ5(-)TCRβ(+) T cells accumulate with age in Vβ5 transgenic mice and bear a diverse TCR Vβ repertoire, it is unknown whether they respond to homeostatic and antigenic stimuli and thus may benefit the host. We demonstrate in this study that postrevision cells are functional. These cells have a high rate of steady-state homeostatic proliferation in situ, and they undergo extensive MHC class II-dependent lymphopenia-induced proliferation. Importantly, postrevision cells do not proliferate in response to the tolerizing superantigen, implicating TCR revision as a mechanism of tolerance induction and demonstrating that TCR-dependent activation of postrevision cells is not driven by the transgene-encoded receptor. Postrevision cells proliferate extensively to commensal bacterial Ags and can generate I-A(b)-restricted responses to Ag by producing IFN-γ following Listeria monocytogenes challenge. These data show that rescued postrevision T cells are responsive to homeostatic signals and recognize self- and foreign peptides in the context of self-MHC and are thus useful to the host.

TCR revision generates functional CD4+ T cells1

PubMed Central

Hale, J. Scott; Wubeshet, Maramawit; Fink, Pamela J.

2010-01-01

CD4+Vβ5+ peripheral T cells in B6 mice respond to encounter with a peripherally-expressed endogenous superantigen by undergoing either deletion or TCR revision. In this latter process, cells lose surface Vβ5 expression and undergo RAG-dependent rearrangement of endogenous TCRβ genes, driving surface expression of novel TCRs. While post-revision CD4+Vβ5−TCRβ+ T cells accumulate with age in Vβ5 transgenic mice and bear a diverse TCR Vβ repertoire, it is unknown whether they respond to homeostatic and antigenic stimuli, and thus may benefit the host. We now demonstrate that post-revision cells are functional. These cells have a high rate of steady-state homeostatic proliferation in situ and they undergo extensive MHC class II-dependent lymphopenia-induced proliferation. Importantly, post-revision cells do not proliferate in response to the tolerizing superantigen, implicating TCR revision as a mechanism of tolerance induction and demonstrating that TCR-dependent activation of post-revision cells is not driven by the transgene-encoded receptor. Post-revision cells proliferate extensively to commensal bacterial Ags and can generate I-Ab-restricted responses to Ag by producing IFNγ following Listeria monocytogenes challenge. These data show that rescued post-revision T cells are responsive to homeostatic signals and recognize self and foreign peptides in the context of self MHC, and are thus useful to the host. PMID:20971922

Increasing functional avidity of TCR-redirected T cells by removing defined N-glycosylation sites in the TCR constant domain

PubMed Central

Hauptrock, Beate; Malina, Victoria; Antunes, Edite; Voss, Ralf-Holger; Wolfl, Matthias; Strong, Roland; Theobald, Matthias; Greenberg, Philip D.

2009-01-01

Adoptive transfer of T lymphocytes transduced with a T cell receptor (TCR) to impart tumor reactivity has been reported as a potential strategy to redirect immune responses to target cancer cells (Schumacher, T.N. 2002. Nat. Rev. Immunol. 2:512–519). However, the affinity of most TCRs specific for shared tumor antigens that can be isolated is usually low. Thus, strategies to increase the affinity of TCRs or the functional avidity of TCR-transduced T cells might be therapeutically beneficial. Because glycosylation affects the flexibility, movement, and interactions of surface molecules, we tested if selectively removing conserved N-glycoslyation sites in the constant regions of TCR α or β chains could increase the functional avidity of T cells transduced with such modified TCRs. We observed enhanced functional avidity and improved recognition of tumor cells by T cells harboring TCR chains with reduced N-glycosylation (ΔTCR) as compared with T cells with wild-type (WT) TCR chains. T cells transduced with WT or ΔTCR chains bound tetramer equivalently at 4°C, but tetramer binding was enhanced at 37°C, predominantly as a result of reduced tetramer dissociation. This suggested a temperature-dependent mechanism such as TCR movement in the cell surface or structural changes of the TCR allowing improved multimerization. This strategy was effective with mouse and human TCRs specific for different antigens and, thus, should be readily translated to TCRs with any specificity. PMID:19171765

The N terminus of SKAP55 enables T cell adhesion to TCR and integrin ligands via distinct mechanisms

PubMed Central

Ophir, Michael J.; Liu, Beiyun C.

2013-01-01

The T cell receptor (TCR) triggers the assembly of “SLP-76 microclusters,” which mediate signals required for T cell activation. In addition to regulating integrin activation, we show that Src kinase–associated phosphoprotein of 55 kD (SKAP55) is required for microcluster persistence and movement, junctional stabilization, and integrin-independent adhesion via the TCR. These functions require the dimerization of SKAP55 and its interaction with the adaptor adhesion and degranulation-promoting adaptor protein (ADAP). A “tandem dimer” containing two ADAP-binding SKAP55 Src homology 3 (SH3) domains stabilized SLP-76 microclusters and promoted T cell adhesion via the TCR, but could not support adhesion to integrin ligands. Finally, the SKAP55 dimerization motif (DM) enabled the coimmunoprecipitation of the Rap1-dependent integrin regulator Rap1-GTP–interacting adaptor molecule (RIAM), the recruitment of talin into TCR-induced adhesive junctions, and “inside-out” signaling to β1 integrins. Our data indicate that SKAP55 dimers stabilize SLP-76 microclusters, couple SLP-76 to the force-generating systems responsible for microcluster movement, and enable adhesion via the TCR by mechanisms independent of RIAM, talin, and β1 integrins. PMID:24368808

TCR repertoires of intratumoral T-cell subsets.

PubMed

Linnemann, Carsten; Mezzadra, Riccardo; Schumacher, Ton N M

2014-01-01

The infiltration of human tumors by T cells is a common phenomenon, and over the past decades, it has become increasingly clear that the nature of such intratumoral T-cell populations can predict disease course. Furthermore, intratumoral T cells have been utilized therapeutically in clinical studies of adoptive T-cell therapy. In this review, we describe how novel methods that are either based on T-cell receptor (TCR) sequencing or on cancer exome analysis allow the analysis of the tumor reactivity and antigen-specificity of the intratumoral TCR repertoire with unprecedented detail. Furthermore, we discuss studies that have started to utilize these techniques to probe the link between cancer exomes and the intratumoral TCR pool. Based on the observation that both the cancer epitope repertoire and intratumoral TCR repertoire appear highly individual, we outline strategies, such as 'autologous TCR gene therapy', that exploit the tumor-resident TCR repertoire for the development of personalized immunotherapy. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

TCR and IL-7 Signaling Are Altered in the Absence of Functional GTPase of the Immune Associated Nucleotide Binding Protein 5 (GIMAP5)

PubMed Central

Chen, Xi-Lin; Serrano, Daniel; Ghobadi, Farnaz; Mayhue, Marian; Hoebe, Kasper; Ilangumaran, Subburaj; Ramanathan, Sheela

2016-01-01

GTPase of the immune associated nucleotide binding protein (GIMAP) family of proteins are expressed essentially in cells of the hematopoietic system. Mutation in the founding member of this gene family, Gimap5, results in the lymphopenic phenotype in Bio-Breeding diabetes prone rats. In mice, deletion of functional Gimap5 gene affects the survival and renewal of hematopoietic stem cells in addition to the defects observed in T cells. Here we show that T cells from OTII TCR-transgenic Gimap5sph/sph mice do not proliferate in response to its cognate antigen. Furthermore, T cells from Gimap5 mutant rats and mice show decreased phosphorylation of STAT5 following stimulation with IL-7. Our results suggest that functional Gimap5 is required for optimal signaling through TCR and IL-7R in T cells. PMID:27023180

The tyrosine phosphatase PTPN22 discriminates weak self peptides from strong agonist TCR signals.

PubMed

Salmond, Robert J; Brownlie, Rebecca J; Morrison, Vicky L; Zamoyska, Rose

2014-09-01

T cells must be tolerant of self antigens to avoid autoimmunity but responsive to foreign antigens to provide protection against infection. We found that in both naive T cells and effector T cells, the tyrosine phosphatase PTPN22 limited signaling via the T cell antigen receptor (TCR) by weak agonists and self antigens while not impeding responses to strong agonist antigens. T cells lacking PTPN22 showed enhanced formation of conjugates with antigen-presenting cells pulsed with weak peptides, which led to activation of the T cells and their production of inflammatory cytokines. This effect was exacerbated under conditions of lymphopenia, with the formation of potent memory T cells in the absence of PTPN22. Our data address how loss-of-function PTPN22 alleles can lead to the population expansion of effector and/or memory T cells and a predisposition to human autoimmunity.

A HYPOTHESIS ACCOUNTING FOR THE PARADOXICAL EXPRESSION OF THE D GENE SEGMENT IN THE BCR AND THE TCR

PubMed Central

Cohn, Melvin

2009-01-01

The D gene segment expressed in both the TCR and BCR has a challenging behavior that begs interpretation. It is incorporated in three reading frames in the rearranged transcription unit but is expressed in antigen-selected cells in a preferred frame. Why was it so important to waste 2/3 of newborn cells? The hypothesis is presented that the D region is framework playing a role in both the TCR and the BCR by determining whether a signal is transmitted to the cell upon interaction with a cognate ligand. This assumption operates in determining haplotype exclusion for the BCR and in regulating the signaling orientation for the TCR. Relevant data as well as a definitive experiment challenging the validity of this hypothesis, are discussed. PMID:18546143

T-cell synapse formation depends on antigen recognition but not CD3 interaction: studies with TCR:ζ, a candidate transgene for TCR gene therapy.

PubMed

Roszik, János; Sebestyén, Zsolt; Govers, Coen; Guri, Yakir; Szöor, Arpád; Pályi-Krekk, Zsuzsanna; Vereb, György; Nagy, Peter; Szöllosi, János; Debets, Reno

2011-05-01

T-cell receptors (TCRs) can be genetically modified to improve gene-engineered T-cell responses, a strategy considered critical for the success of clinical TCR gene therapy to treat cancers. TCR:ζ, which is a heterodimer of TCRα and β chains each coupled to complete human CD3ζ, overcomes issues of mis-pairing with endogenous TCR chains, shows high surface expression and mediates antigen-specific T-cell functions in vitro. In the current study, we further characterized TCR:ζ in gene-engineered T cells and assessed whether this receptor is able to interact with surface molecules and drive correct synapse formation in Jurkat T cells. The results showed that TCR:ζ mediates the formation of synaptic areas with antigen-positive target cells, interacts closely with CD8α and MHC class I (MHCI), and co-localizes with CD28, CD45 and lipid rafts, similar to WT TCR. TCR:ζ did not closely associate with endogenous CD3ε, despite its co-presence in immune synapses, and TCR:ζ showed enhanced synaptic accumulation in T cells negative for surface-expressed TCR molecules. Notably, synaptic TCR:ζ demonstrated lowered densities when compared with TCR in dual TCR T cells, a phenomenon that was related to both extracellular and intracellular CD3ζ domains present in the TCR:ζ molecule and responsible for enlarged synapse areas. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Protein kinase D2 is a digital amplifier of T cell receptor–stimulated diacylglycerol signaling in naïve CD8+ T cells

PubMed Central

Navarro, María N.; Feijoo-Carnero, Carmen; Arandilla, Alba Gonzalez; Trost, Matthias; Cantrell, Doreen A.

2016-01-01

Protein kinase D2 (PKD2) is a serine and threonine kinase that is activated in T cells by diacylglycerol and protein kinase C in response to stimulation of the T cell receptor (TCR) by antigen. We quantified the activation of PKD2 at the single-cell level and found that this kinase acts as a sensitive digital amplifier of TCR engagement, enabling CD8+ T cells to match the production of inflammatory cytokines to the quality and quantity of TCR ligands. There was a digital response pattern of PKD2 activation in response to TCR engagement, such that increasing the concentration and potency of TCR ligands increased the number of cells that exhibited activated PKD2. However, for each cell that responded to TCR stimulation, the entire cellular pool of PKD2 (~400,000 molecules) was activated. Moreover, PKD2 acted as an amplification checkpoint for antigen-stimulated digital cytokine responses and translated the differential strength of TCR signaling to determine the number of naïve CD8+ T cells that became effector cells. Together, these results provide insights into PKD family kinases and how they act digitally to amplify signaling networks controlled by the TCR. PMID:25336615

Modulation of TCRβ surface expression during TCR revision.

PubMed

Simmons, Kalynn B; Wubeshet, Maramawit; Ames, Kristina T; McMahan, Catherine J; Hale, J Scott; Fink, Pamela J

2012-01-01

TCR revision is a tolerance mechanism by which self-reactive TCRs expressed by mature CD4(+) peripheral T cells are replaced by receptors encoded by genes generated by post-thymic DNA rearrangement. The downmodulation of surface TCR expression initiates TCR revision, and serves as a likely trigger for the induction of the recombinase machinery. We show here in a Vβ5 transgenic mouse model system that downregulation of the self-reactive transgene-encoded TCR is not maintained by transgene loss or diminished transcription or translation. The downregulation of surface TCR expression likely occurs in two stages, only one of which requires tolerogen expression. Copyright © 2011 Elsevier Inc. All rights reserved.

TCR backscattering characterization for microwave remote sensing

NASA Astrophysics Data System (ADS)

Riccio, Giovanni; Gennarelli, Claudio

2014-05-01

A Trihedral Corner Reflector (TCR) is formed by three mutually orthogonal metal plates of various shapes and is a very important scattering structure since it exhibits a high monostatic Radar Cross Section (RCS) over a wide angular range. Moreover it is a handy passive device with low manufacturing costs and robust geometric construction, the maintenance of its efficiency is not difficult and expensive, and it can be used in all weather conditions (i.e., fog, rain, smoke, and dusty environment). These characteristics make it suitable as reference target and radar enhancement device for satellite- and ground-based microwave remote sensing techniques. For instance, TCRs have been recently employed to improve the signal-to-noise ratio of the backscattered signal in the case of urban ground deformation monitoring [1] and dynamic survey of civil infrastructures without natural corners as the Musmeci bridge in Basilicata, Italy [2]. The region of interest for the calculation of TCR's monostatic RCS is here confined to the first quadrant containing the boresight direction. The backscattering term is presented in closed form by evaluating the far-field scattering integral involving the contributions related to the direct illumination and the internal bouncing mechanisms. The Geometrical Optics (GO) laws allow one to determine the field incident on each TCR plate and the patch (integration domain) illuminated by it, thus enabling the use of a Physical Optics (PO) approximation for the corresponding surface current densities to consider for integration on each patch. Accordingly, five contributions are associated to each TCR plate: one contribution is due to the direct illumination of the whole internal surface; two contributions originate by the impinging rays that are simply reflected by the other two internal surfaces; and two contributions are related to the impinging rays that undergo two internal reflections. It is useful to note that the six contributions due to the

Optimization of T-cell Reactivity by Exploiting TCR Chain Centricity for the Purpose of Safe and Effective Antitumor TCR Gene Therapy.

PubMed

Ochi, Toshiki; Nakatsugawa, Munehide; Chamoto, Kenji; Tanaka, Shinya; Yamashita, Yuki; Guo, Tingxi; Fujiwara, Hiroshi; Yasukawa, Masaki; Butler, Marcus O; Hirano, Naoto

2015-09-01

Adoptive transfer of T cells redirected by a high-affinity antitumor T-cell receptor (TCR) is a promising treatment modality for cancer patients. Safety and efficacy depend on the selection of a TCR that induces minimal toxicity and elicits sufficient antitumor reactivity. Many, if not all, TCRs possess cross-reactivity to unrelated MHC molecules in addition to reactivity to target self-MHC/peptide complexes. Some TCRs display chain centricity, in which recognition of MHC/peptide complexes is dominated by one of the TCR hemi-chains. In this study, we comprehensively studied how TCR chain centricity affects reactivity to target self-MHC/peptide complexes and alloreactivity using the TCR, clone TAK1, which is specific for human leukocyte antigen-A*24:02/Wilms tumor 1(235-243) (A24/WT1(235)) and cross-reactive with B*57:01 (B57). The TAK1β, but not the TAK1α, hemi-chain possessed chain centricity. When paired with multiple clonotypic TCRα counter-chains encoding TRAV12-2, 20, 36, or 38-2, the de novo TAK1β-containing TCRs showed enhanced, weakened, or absent reactivity to A24/WT1(235) and/or to B57. T cells reconstituted with these TCRα genes along with TAK1β possessed a very broad range (>3 log orders) of functional and structural avidities. These results suggest that TCR chain centricity can be exploited to enhance desired antitumor TCR reactivity and eliminate unwanted TCR cross-reactivity. TCR reactivity to target MHC/peptide complexes and cross-reactivity to unrelated MHC molecules are not inextricably linked and are separable at the TCR sequence level. However, it is still mandatory to carefully monitor for possible harmful toxicities caused by adoptive transfer of T cells redirected by thymically unselected TCRs. ©2015 American Association for Cancer Research.

Src-like adaptor protein regulates TCR expression on thymocytes by linking the ubiquitin ligase c-Cbl to the TCR complex.

PubMed

Myers, Margaret D; Sosinowski, Tomasz; Dragone, Leonard L; White, Carmen; Band, Hamid; Gu, Hua; Weiss, Arthur

2006-01-01

The adaptor molecule SLAP and E3 ubiquitin ligase c-Cbl each regulate expression of T cell receptor (TCR)-CD3 on thymocytes. Here we provide genetic and biochemical evidence that both molecules function in the same pathway. TCR-CD3 expression was similar in the absence of SLAP and/or c-Cbl. SLAP and c-Cbl were found to interact, and their expression together downregulated CD3epsilon. This required multiple domains in SLAP and the ring finger of c-Cbl. Furthermore, expression of SLAP and c-Cbl together induced TCRzeta ubiquitination and degradation, preventing the accumulation of fully assembled recycling TCR complexes. These studies indicate that SLAP links the E3 ligase activity of c-Cbl to the TCR, allowing for stage-specific regulation of TCR expression.

TLR4 signaling in effector CD4+ T cells regulates TCR activation and experimental colitis in mice

PubMed Central

González-Navajas, José M.; Fine, Sean; Law, Jason; Datta, Sandip K.; Nguyen, Kim P.; Yu, Mandy; Corr, Maripat; Katakura, Kyoko; Eckman, Lars; Lee, Jongdae; Raz, Eyal

2010-01-01

TLRs sense various microbial products. Their function has been best characterized in DCs and macrophages, where they act as important mediators of innate immunity. TLR4 is also expressed on CD4+ T cells, but its physiological function on these cells remains unknown. Here, we have shown that TLR4 triggering on CD4+ T cells affects their phenotype and their ability to provoke intestinal inflammation. In a model of spontaneous colitis, Il10–/–Tlr4–/– mice displayed accelerated development of disease, with signs of overt colitis as early as 8 weeks of age, when compared with Il10–/– and Il10–/–Tlr9–/– mice, which did not develop colitis by 8 months. Similar results were obtained in a second model of colitis in which transfer of naive Il10–/–Tlr4–/– CD4+ T cells into Rag1–/– recipients sufficient for both IL-10 and TLR4 induced more aggressive colitis than the transfer of naive Il10–/– CD4+ T cells. Mechanistically, LPS stimulation of TLR4-bearing CD4+ T cells inhibited ERK1/2 activation upon subsequent TCR stimulation via the induction of MAPK phosphatase 3 (MKP-3). Our data therefore reveal a tonic inhibitory role for TLR4 signaling on subsequent TCR-dependent CD4+ T cell responses. PMID:20051628

PLAC1-specific TCR-engineered T cells mediate antigen-specific antitumor effects in breast cancer

PubMed Central

Li, Qiongshu; Liu, Muyun; Wu, Man; Zhou, Xin; Wang, Shaobin; Hu, Yuan; Wang, Youfu; He, Yixin; Zeng, Xiaoping; Chen, Junhui; Liu, Qubo; Xiao, Dong; Hu, Xiang; Liu, Weibin

2018-01-01

Placenta-specific 1 (PLAC1), a novel cancer-testis antigen (CTA), is expressed in a number of different human malignancies. It is frequently produced in breast cancer, serving a function in tumorigenesis. Adoptive immunotherapy using T cell receptor (TCR)-engineered T cells against CTA mediates objective tumor regression; however, to the best of our knowledge, targeting PLAC1 using engineered T cells has not yet been attempted. In the present study, the cDNAs encoding TCRα- and β-chains specific for human leukocyte antigen (HLA)-A*0201-restricted PLAC1 were cloned from a cytotoxic T-lymphocyte, generated by in vitro by the stimulation of CD8+ T cells using autologous HLA-A2+ dendritic cells loaded with a PLAC1-specific peptide (p28-36, VLCSIDWFM). The TCRα/β-chains were linked by a 2A peptide linker (TCRα-Thosea asigna virus-TCRβ), and the constructs were cloned into the lentiviral vector, followed by transduction into human cytotoxic (CD8+) T cells. The efficiency of transduction was up to 25.16%, as detected by PLAC1 multimers. TCR-transduced CD8+ T cells, co-cultured with human non-metastatic breast cancer MCF-7 cells (PLAC1+, HLA-A2+) and triple-negative breast cancer MDAMB-231 cells (PLAC1+, HLA-A2+), produced interferon γ and tumor necrosis factor α, suggesting TCR activation. Furthermore, the PLAC1 TCR-transduced CD8+ T cells efficiently and specifically identified and annihilated the HLA-A2+/PLAC1+ breast cancer cell lines in a lactate dehydrogenase activity assay. Western blot analysis demonstrated that TCR transduction stimulated the production of mitogen-activated protein kinase signaling molecules, extracellular signal-regulated kinases 1/2 and nuclear factor-κB, through phosphoinositide 3-kinase γ-mediated phosphorylation of protein kinase B in CD8+ T cells. Xenograft mouse assays revealed that PLAC1 TCR-transduced CD8+T cells significantly delayed the tumor progression in mice-bearing breast cancer compared with normal saline or negative

The CD3-Zeta Chimeric Antigen Receptor Overcomes TCR Hypo-Responsiveness of Human Terminal Late-Stage T Cells

PubMed Central

Awerkiew, Sabine; Schmidt, Annette; Hombach, Andreas A.; Pfister, Herbert; Abken, Hinrich

2012-01-01

Adoptive therapy of malignant diseases with tumor-specific cytotoxic T cells showed remarkable efficacy in recent trials. Repetitive T cell receptor (TCR) engagement of target antigen, however, inevitably ends up in hypo-responsive cells with terminally differentiated KLRG-1+ CD57+ CD7− phenotype limiting their therapeutic efficacy. We here revealed that hypo-responsiveness of CMV-specific late-stage CD8+ T cells is due to reduced TCR synapse formation compared to younger cells. Membrane anchoring of TCR components contributes to T cell hypo-responsiveness since dislocation of galectin-3 from the synapse by swainsonine restored both TCR synapse formation and T cell response. Transgenic expression of a CD3-zeta signaling chimeric antigen receptor (CAR) recovered hypo-responsive T cells to full effector functions indicating that the defect is restricted to TCR membrane components while synapse formation of the transgenic CAR was not blocked. CAR engineered late-stage T cells released cytokines and mediated redirected cytotoxicity as efficiently as younger effector T cells. Our data provide a rationale for TCR independent, CAR mediated activation in the adoptive cell therapy to avoid hypo-responsiveness of late-stage T cells upon repetitive antigen encounter. PMID:22292024

SLAT promotes TCR-mediated, Rap1-dependent LFA-1 activation and adhesion through interaction of its PH domain with Rap1

PubMed Central

Côte, Marjorie; Fos, Camille; Canonigo-Balancio, Ann J.; Ley, Klaus; Bécart, Stéphane; Altman, Amnon

2015-01-01

ABSTRACT SLAT (also known as DEF6) promotes T cell activation and differentiation by regulating NFAT-Ca2+ signaling. However, its role in TCR-mediated inside-out signaling, which induces integrin activation and T cell adhesion, a central process in T cell immunity and inflammation, has not been explored. Here, we show that SLAT is crucial for TCR-induced adhesion to ICAM-1 and affinity maturation of LFA-1 in CD4+ T cells. Mechanistic studies revealed that SLAT interacts, through its PH domain, with a key component of inside-out signaling, namely the active form of the small GTPase Rap1 (which has two isoforms, Rap1A and Rap1B). This interaction has been further shown to facilitate the interdependent recruitment of Rap1 and SLAT to the T cell immunological synapse upon TCR engagement. Furthermore, a SLAT mutant lacking its PH domain drastically inhibited LFA-1 activation and CD4+ T cell adhesion. Finally, we established that a constitutively active form of Rap1, which is present at the plasma membrane, rescues the defective LFA-1 activation and ICAM-1 adhesion in SLAT-deficient (Def6−/−) T cells. These findings ascribe a new function to SLAT, and identify Rap1 as a target of SLAT function in TCR-mediated inside-out signaling. PMID:26483383

SLAT promotes TCR-mediated, Rap1-dependent LFA-1 activation and adhesion through interaction of its PH domain with Rap1.

PubMed

Côte, Marjorie; Fos, Camille; Canonigo-Balancio, Ann J; Ley, Klaus; Bécart, Stéphane; Altman, Amnon

2015-12-01

SLAT (also known as DEF6) promotes T cell activation and differentiation by regulating NFAT-Ca(2+) signaling. However, its role in TCR-mediated inside-out signaling, which induces integrin activation and T cell adhesion, a central process in T cell immunity and inflammation, has not been explored. Here, we show that SLAT is crucial for TCR-induced adhesion to ICAM-1 and affinity maturation of LFA-1 in CD4(+) T cells. Mechanistic studies revealed that SLAT interacts, through its PH domain, with a key component of inside-out signaling, namely the active form of the small GTPase Rap1 (which has two isoforms, Rap1A and Rap1B). This interaction has been further shown to facilitate the interdependent recruitment of Rap1 and SLAT to the T cell immunological synapse upon TCR engagement. Furthermore, a SLAT mutant lacking its PH domain drastically inhibited LFA-1 activation and CD4(+) T cell adhesion. Finally, we established that a constitutively active form of Rap1, which is present at the plasma membrane, rescues the defective LFA-1 activation and ICAM-1 adhesion in SLAT-deficient (Def6(-/-)) T cells. These findings ascribe a new function to SLAT, and identify Rap1 as a target of SLAT function in TCR-mediated inside-out signaling. © 2015. Published by The Company of Biologists Ltd.

CRISPR-mediated TCR replacement generates superior anticancer transgenic T cells.

PubMed

Legut, Mateusz; Dolton, Garry; Mian, Afsar Ali; Ottmann, Oliver G; Sewell, Andrew K

2018-01-18

Adoptive transfer of T cells genetically modified to express a cancer-specific T-cell receptor (TCR) has shown significant therapeutic potential for both hematological and solid tumors. However, a major issue of transducing T cells with a transgenic TCR is the preexisting expression of TCRs in the recipient cells. These endogenous TCRs compete with the transgenic TCR for surface expression and allow mixed dimer formation. Mixed dimers, formed by mispairing between the endogenous and transgenic TCRs, may harbor autoreactive specificities. To circumvent these problems, we designed a system where the endogenous TCR-β is knocked out from the recipient cells using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 (Cas9) technology, simultaneously with transduction with a cancer-reactive receptor of choice. This TCR replacement strategy resulted in markedly increased surface expression of transgenic αβ and γδ TCRs, which in turn translated to a stronger, and more polyfunctional, response of engineered T cells to their target cancer cell lines. Additionally, the TCR-plus-CRISPR-modified T cells were up to a thousandfold more sensitive to antigen than standard TCR-transduced T cells or conventional model proxy systems used for studying TCR activity. Finally, transduction with a pan-cancer-reactive γδ TCR used in conjunction with CRISPR/Cas9 knockout of the endogenous αβ TCR resulted in more efficient redirection of CD4 + and CD8 + T cells against a panel of established blood cancers and primary, patient-derived B-cell acute lymphoblastic leukemia blasts compared with standard TCR transfer. Our results suggest that TCR transfer combined with genome editing could lead to new, improved generations of cancer immunotherapies. © 2018 by The American Society of Hematology.

Organization of the resting TCR in nanoscale oligomers.

PubMed

Schamel, Wolfgang W A; Alarcón, Balbino

2013-01-01

Despite the low affinity of the T-cell antigen receptor (TCR) for its peptide/major histocompatibility complex (pMHC) ligand, T cells are very sensitive to their antigens. This paradox can be resolved if we consider that the TCR may be organized into pre-existing oligomers or nanoclusters. Such structures could improve antigen recognition by increasing the functional affinity (avidity) of the TCR-pMHC interaction and by allowing cooperativity between individual TCRs. Up to approximately 20 TCRs become tightly apposed in these nanoclusters, often in a linear manner, and such structures could reflect a relatively generalized phenomenon: the non-random concentration of membrane receptors in specific areas of the plasma membrane known as protein islands. The association of TCRs into nanoclusters can explain the enhanced kinetics of the pMHC-TCR interaction in two dimensional versus three dimensional systems, but also their existence calls for a revision of the TCR triggering models based on pMHC-induced TCR clustering. Interestingly, the B-cell receptor and the FcεRI have also been shown to form nanoclusters, suggesting that the formation of pre-existing receptor oligomers could be widely used in the immune system. © 2012 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

In TCR-Stimulated T-cells, N-ras Regulates Specific Genes and Signal Transduction Pathways

PubMed Central

Lynch, Stephen J.; Zavadil, Jiri; Pellicer, Angel

2013-01-01

It has been recently shown that N-ras plays a preferential role in immune cell development and function; specifically: N-ras, but not H-ras or K-ras, could be activated at and signal from the Golgi membrane of immune cells following a low level T-cell receptor stimulus. The goal of our studies was to test the hypothesis that N-ras and H-ras played distinct roles in immune cells at the level of the transcriptome. First, we showed via mRNA expression profiling that there were over four hundred genes that were uniquely differentially regulated either by N-ras or H-ras, which provided strong evidence in favor of the hypothesis that N-ras and H-ras have distinct functions in immune cells. We next characterized the genes that were differentially regulated by N-ras in T cells following a low-level T-cell receptor stimulus. Of the large pool of candidate genes that were differentially regulated by N-ras downstream of TCR ligation, four genes were verified in qRT-PCR-based validation experiments (Dntt, Slc9a6, Chst1, and Lars2). Finally, although there was little overlap between individual genes that were regulated by N-ras in unstimulated thymocytes and stimulated CD4+ T-cells, there was a nearly complete correspondence between the signaling pathways that were regulated by N-ras in these two immune cell types. PMID:23755101

Structure of the superantigen staphylococcal enterotoxin B in complex with TCR and peptide-MHC demonstrates absence of TCR-peptide contacts.

PubMed

Rödström, Karin E J; Elbing, Karin; Lindkvist-Petersson, Karin

2014-08-15

Superantigens are immune-stimulatory toxins produced by Staphylococcus aureus, which are able to interact with host immune receptors to induce a massive release of cytokines, causing toxic shock syndrome and possibly death. In this article, we present the x-ray structure of staphylococcal enterotoxin B (SEB) in complex with its receptors, the TCR and MHC class II, forming a ternary complex. The structure, in combination with functional analyses, clearly shows how SEB adopts a wedge-like position when binding to the β-chain of TCR, allowing for an interaction between the α-chain of TCR and MHC. Furthermore, the binding mode also circumvents contact between TCR and the peptide presented by MHC, which enables SEB to initiate a peptide-independent activation of T cells. Copyright © 2014 by The American Association of Immunologists, Inc.

Rag Deletion in Peripheral T Cells Blocks TCR Revision

PubMed Central

Hale, J. Scott; Ames, Kristina T.; Boursalian, Tamar E.; Fink, Pamela J.

2010-01-01

Mature CD4+Vβ5+ T cells that recognize a peripherally expressed endogenous superantigen are tolerized either by deletion or T cell receptor (TCR) revision. In Vβ5 transgenic mice, this latter tolerance pathway results in the appearance of CD4+Vβ5−TCRβ+ T cells, coinciding with Rag1, Rag2, and TdT expression and the accumulation of Vβ-DJβ recombination intermediates in peripheral CD4+ T cells. Because post-thymic RAG-dependent TCR rearrangement has remained controversial, we sought to definitively determine whether TCR revision is an extrathymic process that occurs in mature peripheral T cells. We now show that Rag deletion in post-positive selection T cells in Vβ5 transgenic mice blocks TCR revision in vivo, and that mature peripheral T cells sorted to remove cells bearing endogenous TCRβ chains can express newly generated TCRβ molecules in adoptive hosts. These findings unambiguously demonstrate post-thymic, RAG-dependent TCR rearrangement and define TCR revision as a tolerance pathway that targets mature peripheral CD4+ T cells. PMID:20435935

The mechanism of chromosome 7 inversion in human lymphocytes expressing chimeric gamma beta TCR.

PubMed

Retière, C; Halary, F; Peyrat, M A; Le Deist, F; Bonneville, M; Hallet, M M

1999-01-15

Functional chimeric TCR chains, encoded by V gamma J gamma C beta or V gamma J beta C beta hybrid gene TCR, are expressed at the surface of a small fraction of alpha beta T lymphocytes in healthy individuals. Their frequency is dramatically increased in patients with ataxia-telangiectasia, a syndrome associated with inherited genomic instability. As the TCR gamma and beta loci are in an inverted orientation on chromosome 7, the generation of such hybrid genes requires at least an inversion event. Until now, neither the sequences involved in this genetic mechanism nor the number of recombinations leading to the formation of functional transcriptional units have been characterized. In this manuscript, we demonstrate that at least two rearrangements, involving classical recombination signal sequence and the V(D)J recombinase complex, lead to the formation of productive hybrid genes. A primary inversion 7 event between D beta and J gamma genic segments generates C gamma V beta and C beta V gamma hybrid loci. Within the C gamma V beta locus, secondary rearrangements between V gamma and J gamma or V gamma and J beta elements generate functional genes. Besides, our results suggest that secondary rearrangements were blocked in the C beta V gamma locus of normal but not ataxia-telangiectasia T lymphocytes. We also provide formal evidence that the same D beta-3' recombination signal sequence can be used in successive rearrangements with J gamma and J beta genic segments, thus showing that a signal joint has been involved in a secondary recombination event.

TCR-independent CD28-mediated gene expression in peripheral blood lymphocytes from donors chronically infected with HIV-1.

PubMed Central

Wong, J G; Smithgall, M D; Haffar, O K

1997-01-01

Complete activation of peripheral blood T cells requires both T-cell receptor (TCR) stimulation and CD28 costimulation. Signalling pathways associated specifically with CD28 are not well understood, however, because ligation of CD28 in the absence of TCR stimulation does not give rise to cellular responses in normal cells. In peripheral blood lymphocytes (PBL) from donors chronically infected with human immunodeficiency virus-1 (HIV-1), CD28 can induce viral replication through an alternative pathway that does not require TCR ligation. We have exploited this observation to study CD28-mediated signal transduction using reverse transcriptase-mediated polymerase chain reaction (RT-PCR) to amplify viral RNA. Independent ligation of CD28 on donor PBL induced expression of the HIV-1 tat gene but not the interleukin-2 (IL-2) gene. Viral induction did not occur following pretreatment of cells with actinomycin D, suggesting it was mediated through transcriptional activation of the viral long terminal repeat (LTR). tat was induced in the presence of the protein kinase C inhibitor H-7, but was inhibited by cyclosporin A. Our results demonstrate that CD28 is linked directly to specific signalling pathways leading to de novo induction of genes in PBL. Images Figure 1 Figure 2 Figure 3 PMID:9135558

Phosphoproteomics analyses show subnetwork systems in T-cell receptor signaling.

PubMed

Hatano, Atsushi; Matsumoto, Masaki; Nakayama, Keiichi I

2016-10-01

A key issue in the study of signal transduction is how multiple signaling pathways are systematically integrated into the cell. We have now performed multiple phosphoproteomics analyses focused on the dynamics of the T-cell receptor (TCR) signaling network and its subsystem mediated by the Ca 2+ signaling pathway. Integration of these phosphoproteomics data sets and extraction of components of the TCR signaling network dependent on Ca 2+ signaling showed unexpected phosphorylation kinetics for candidate substrates of the Ca 2+ -dependent phosphatase calcineurin (CN) during TCR stimulation. Detailed characterization of the TCR-induced phosphorylation of a novel CN substrate, Itpkb, showed that phosphorylation of this protein is regulated by both CN and the mitogen-activated protein kinase Erk in a competitive manner. Phosphorylation of additional CN substrates was also found to be regulated by Erk and CN in a similar manner. The combination of multiple phosphoproteomics approaches thus showed two major subsystems mediated by Erk and CN in the TCR signaling network, with these subsystems regulating the phosphorylation of a group of proteins in a competitive manner. © 2016 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

High-throughput identification of antigen-specific TCRs by TCR gene capture.

PubMed

Linnemann, Carsten; Heemskerk, Bianca; Kvistborg, Pia; Kluin, Roelof J C; Bolotin, Dmitriy A; Chen, Xiaojing; Bresser, Kaspar; Nieuwland, Marja; Schotte, Remko; Michels, Samira; Gomez-Eerland, Raquel; Jahn, Lorenz; Hombrink, Pleun; Legrand, Nicolas; Shu, Chengyi Jenny; Mamedov, Ilgar Z; Velds, Arno; Blank, Christian U; Haanen, John B A G; Turchaninova, Maria A; Kerkhoven, Ron M; Spits, Hergen; Hadrup, Sine Reker; Heemskerk, Mirjam H M; Blankenstein, Thomas; Chudakov, Dmitriy M; Bendle, Gavin M; Schumacher, Ton N M

2013-11-01

The transfer of T cell receptor (TCR) genes into patient T cells is a promising approach for the treatment of both viral infections and cancer. Although efficient methods exist to identify antibodies for the treatment of these diseases, comparable strategies to identify TCRs have been lacking. We have developed a high-throughput DNA-based strategy to identify TCR sequences by the capture and sequencing of genomic DNA fragments encoding the TCR genes. We establish the value of this approach by assembling a large library of cancer germline tumor antigen-reactive TCRs. Furthermore, by exploiting the quantitative nature of TCR gene capture, we show the feasibility of identifying antigen-specific TCRs in oligoclonal T cell populations from either human material or TCR-humanized mice. Finally, we demonstrate the ability to identify tumor-reactive TCRs within intratumoral T cell subsets without knowledge of antigen specificities, which may be the first step toward the development of autologous TCR gene therapy to target patient-specific neoantigens in human cancer.

Normalized Synergy Predicts That CD8 Co-Receptor Contribution to T Cell Receptor (TCR) and pMHC Binding Decreases As TCR Affinity Increases in Human Viral-Specific T Cells

PubMed Central

Williams, Chad M.; Schonnesen, Alexandra A.; Zhang, Shu-Qi; Ma, Ke-Yue; He, Chenfeng; Yamamoto, Tori; Eckhardt, S. Gail; Klebanoff, Christopher A.; Jiang, Ning

2017-01-01

The discovery of naturally occurring T cell receptors (TCRs) that confer specific, high-affinity recognition of pathogen and cancer-associated antigens remains a major goal in cellular immunotherapies. The contribution of the CD8 co-receptor to the interaction between the TCR and peptide-bound major histocompatibility complex (pMHC) has previously been correlated with the activation and responsiveness of CD8+ T cells. However, these studies have been limited to model systems of genetically engineered hybridoma TCRs or transgenic mouse TCRs against either a single epitope or an array of altered peptide ligands. CD8 contribution in a native human antigen-specific T cell response remains elusive. Here, using Hepatitis C Virus-specific precursor CTLs spanning a large range of TCR affinities, we discovered that the functional responsiveness of any given TCR correlated with the contribution of CD8 to TCR/pMHC binding. Furthermore, we found that CD8 contribution to TCR/pMHC binding in the two-dimensional (2D) system was more accurately reflected by normalized synergy (CD8 cooperation normalized by total TCR/pMHC bonds) rather than synergy (total CD8 cooperation) alone. While synergy showed an increasing trend with TCR affinity, normalized synergy was demonstrated to decrease with the increase of TCR affinity. Critically, normalized synergy was shown to correlate with CTL functionality and peptide sensitivity, corroborating three-dimensional (3D) analysis of CD8 contribution with respect to TCR affinity. In addition, we identified TCRs that were independent of CD8 for TCR/pMHC binding. Our results resolve the current discrepancy between 2D and 3D analysis on CD8 contribution to TCR/pMHC binding, and demonstrate that naturally occurring high-affinity TCRs are more capable of CD8-independent interactions that yield greater functional responsiveness even with CD8 blocking. Taken together, our data suggest that addition of the normalized synergy parameter to our previously

Inverted repeats in the promoter as an autoregulatory sequence for TcrX in Mycobacterium tuberculosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhattacharya, Monolekha; Das, Amit Kumar, E-mail: amitk@hijli.iitkgp.ernet.in

Highlights: Black-Right-Pointing-Pointer The regulatory sequences recognized by TcrX have been identified. Black-Right-Pointing-Pointer The regulatory region comprises of inverted repeats segregated by 30 bp region. Black-Right-Pointing-Pointer The mode of binding of TcrX with regulatory sequence is unique. Black-Right-Pointing-Pointer In silico TcrX-DNA docked model binds one of the inverted repeats. Black-Right-Pointing-Pointer Both phosphorylated and unphosphorylated TcrX binds regulatory sequence in vitro. -- Abstract: TcrY, a histidine kinase, and TcrX, a response regulator, constitute a two-component system in Mycobacterium tuberculosis. tcrX, which is expressed during iron scarcity, is instrumental in the survival of iron-dependent M. tuberculosis. However, the regulator of tcrX/Y has notmore » been fully characterized. Crosslinking studies of TcrX reveal that it can form oligomers in vitro. Electrophoretic mobility shift assays (EMSAs) show that TcrX recognizes two regions in the promoter that are comprised of inverted repeats separated by {approx}30 bp. The dimeric in silico model of TcrX predicts binding to one of these inverted repeat regions. Site-directed mutagenesis and radioactive phosphorylation indicate that D54 of TcrX is phosphorylated by H256 of TcrY. However, phosphorylated and unphosphorylated TcrX bind the regulatory sequence with equal efficiency, which was shown with an EMSA using the D54A TcrX mutant.« less

Cutting Edge: Rag deletion in peripheral T cells blocks TCR revision.

PubMed

Hale, J Scott; Ames, Kristina T; Boursalian, Tamar E; Fink, Pamela J

2010-06-01

Mature CD4(+)Vbeta5(+) T cells that recognize a peripherally expressed endogenous superantigen are tolerized either by deletion or TCR revision. In Vbeta5 transgenic mice, this latter tolerance pathway results in the appearance of CD4(+)Vbeta5(-)TCRbeta(+) T cells, coinciding with Rag1, Rag2, and TdT expression and the accumulation of V(beta)-DJ(beta) recombination intermediates in peripheral CD4(+) T cells. Because postthymic RAG-dependent TCR rearrangement has remained controversial, we sought to definitively determine whether TCR revision is an extrathymic process that occurs in mature peripheral T cells. We show in this study that Rag deletion in post-positive selection T cells in Vbeta5 transgenic mice blocks TCR revision in vivo and that mature peripheral T cells sorted to remove cells bearing endogenous TCRbeta-chains can express newly generated TCRbeta molecules in adoptive hosts. These findings unambiguously demonstrate postthymic, RAG-dependent TCR rearrangement and define TCR revision as a tolerance pathway that targets mature peripheral CD4(+) T cells.

Suppression of lethal autoimmunity by regulatory T cells with a single TCR specificity

PubMed Central

Hemmers, Saskia; Schizas, Michail; Faire, Mehlika B.; Konopacki, Catherine; Schmidt-Supprian, Marc; Germain, Ronald N.

2017-01-01

The regulatory T cell (T reg cell) T cell receptor (TCR) repertoire is highly diverse and skewed toward recognition of self-antigens. TCR expression by T reg cells is continuously required for maintenance of immune tolerance and for a major part of their characteristic gene expression signature; however, it remains unknown to what degree diverse TCR-mediated interactions with cognate self-antigens are required for these processes. In this study, by experimentally switching the T reg cell TCR repertoire to a single T reg cell TCR, we demonstrate that T reg cell function and gene expression can be partially uncoupled from TCR diversity. An induced switch of the T reg cell TCR repertoire to a random repertoire also preserved, albeit to a limited degree, the ability to suppress lymphadenopathy and T helper cell type 2 activation. At the same time, these perturbations of the T reg cell TCR repertoire led to marked immune cell activation, tissue inflammation, and an ultimately severe autoimmunity, indicating the importance of diversity and specificity for optimal T reg cell function. PMID:28130403

PU.1 regulates TCR expression by modulating GATA-3 activity

PubMed Central

Chang, Hua-Chen; Han, Ling; Jabeen, Rukhsana; Carotta, Sebastian; Nutt, Stephen L.; Kaplan, Mark H.

2009-01-01

The Ets transcription factor PU.1 is a master regulator for the development of multiple lineages during hematopoiesis. The expression pattern of PU.1 is dynamically regulated during early T lineage development in the thymus. We previously revealed that PU.1 delineates heterogeneity of effector Th2 populations. In this study, we further define the function of PU.1 on the Th2 phenotype using mice that specifically lack PU.1 in T cells using an lck-Cre transgene with a conditional Sfpi1 allele (Sfpi1lck-/-). While deletion of PU.1 by the lck-Cre transgene does not affect T cell development, Sfpi1lck-/- T cells have a lower activation threshold than wild type T cells. When TCR engagement is limiting, Sfpi1lck-/- T cells cultured in Th2 polarizing conditions secrete higher levels of Th2 cytokines and have greater cytokine homogeneity than wild type cells. We show that PU.1 modulates the levels of TCR expression in CD4+ T cells by regulating the DNA-binding activity of GATA-3 and limiting GATA-3 regulation of TCR gene expression. GATA-3 dependent regulation of TCR expression is also observed in Th1 and Th2 cells. In CD4+ T cells, PU.1 expression segregates into subpopulations of cells that have lower levels of surface TCR, suggesting that PU.1 contributes to the heterogeneity of TCR expression. Thus, we have identified a mechanism whereby increased GATA-3 function in the absence of the antagonizing activity of PU.1 leads to increased TCR expression, a reduced activation threshold and increased homogeneity in Th2 populations. PMID:19801513

Molecular Mechanisms of Bcl10-Mediated NF-kB Signal Transduction

DTIC Science & Technology

2006-03-08

recruiting and activating the kinase, Akt , which is a critical mediator of pro-survival signals (3) (Figure 3). Figure 3. TCR-induced signaling...kinase and Akt rather than through upstream intermediates initiated by TCR ligation (34, 70). This suggests that TCR stimulation and CD28 co...P. Vito. 2004. Physical and functional interaction of CARMA1 and CARMA3 with Ikappa kinase gamma- NFkappaB essential modulator. J Biol Chem 279

End-binding protein 1 controls signal propagation from the T cell receptor

PubMed Central

Martín-Cófreces, Noa B; Baixauli, Francesc; López, María J; Gil, Diana; Monjas, Alicia; Alarcón, Balbino; Sánchez-Madrid, Francisco

2012-01-01

The role of microtubules (MTs) in the control and dynamics of the immune synapse (IS) remains unresolved. Here, we show that T cell activation requires the growth of MTs mediated by the plus-end specific protein end-binding 1 (EB1). A direct interaction of the T cell receptor (TCR) complex with EB1 provides the molecular basis for EB1 activity promoting TCR encounter with signalling vesicles at the IS. EB1 knockdown alters TCR dynamics at the IS and prevents propagation of the TCR activation signal to LAT, thus inhibiting activation of PLCγ1 and its localization to the IS. These results identify a role for EB1 interaction with the TCR in controlling TCR sorting and its connection with the LAT/PLCγ1 signalosome. PMID:22922463

Detection test of wireless network signal strength and GPS positioning signal in underground pipeline

NASA Astrophysics Data System (ADS)

Li, Li; Zhang, Yunwei; Chen, Ling

2018-03-01

In order to solve the problem of selecting positioning technology for inspection robot in underground pipeline environment, the wireless network signal strength and GPS positioning signal testing are carried out in the actual underground pipeline environment. Firstly, the strength variation of the 3G wireless network signal and Wi-Fi wireless signal provided by China Telecom and China Unicom ground base stations are tested, and the attenuation law of these wireless signals along the pipeline is analyzed quantitatively and described. Then, the receiving data of the GPS satellite signal in the pipeline are tested, and the attenuation of GPS satellite signal under underground pipeline is analyzed. The testing results may be reference for other related research which need to consider positioning in pipeline.

c-Cbl promotes T cell receptor-induced thymocyte apoptosis by activating the phosphatidylinositol 3-kinase/Akt pathway.

PubMed

Thien, Christine B F; Dagger, Samantha A; Steer, James H; Koentgen, Frank; Jansen, Elisa S; Scott, Clare L; Langdon, Wallace Y

2010-04-02

The ability of thymocytes to assess T cell receptor (TCR) signaling strength and initiate the appropriate downstream response is crucial for determining their fate. We have previously shown that a c-Cbl RING finger mutant knock-in mouse, in which the E3 ubiquitin ligase activity of c-Cbl is inactivated, is highly sensitive to TCR-induced death signals that cause thymic deletion. This high intensity signal involves the enhanced tyrosine phosphorylation of the mutant c-Cbl protein promoting a marked increase in the activation of Akt. Here we show that this high intensity signal in c-Cbl RING finger mutant thymocytes also promotes the enhanced induction of two mediators of TCR-directed thymocyte apoptosis, Nur77 and the pro-apoptotic Bcl-2 family member, Bim. In contrast, a knock-in mouse harboring a mutation at Tyr-737, the site in c-Cbl that activates phosphatidylinositol 3-kinase, shows reduced TCR-mediated responses including suppression of Akt activation, a reduced induction of Nur77 and Bim, and greater resistance to thymocyte death. These findings identify tyrosine-phosphorylated c-Cbl as a critical sensor of TCR signal strength that regulates the engagement of death-promoting signals.

SLAP deficiency increases TCR avidity leading to altered repertoire and negative selection of cognate antigen-specific CD8+ T cells

PubMed Central

Friend, Samantha F.; Peterson, Lisa K.; Kedl, Ross M.; Dragone, Leonard L.

2014-01-01

How T cell receptor (TCR) avidity influences CD8+ T cell development and repertoire selection is not yet fully understood. To fill this gap, we utilized Src-like adaptor protein (SLAP)-deficient mice as a tool to increase TCR avidity on double positive (DP) thymocytes. We generated SLAP−/− mice with the transgenic MHC class I-restricted TCR (OT-1) and SLAP−/− Vβ5 mice, expressing only the β-chain of the TCR OT-1 transgene, to examine the effects of increased TCR surface levels on CD8+ T cell development and repertoire selection. In comparing SLAP−/− OT-1 and Vβ5 mice with wild-type controls, we performed compositional analysis and assessed thymocyte signaling by measuring CD5 levels. In addition, we performed tetramer and compositional staining to measure affinity for the cognate antigen, ovalbumin (OVA) peptide, presented by MHC. Furthermore, we quantified differences in α-chain repertoire in SLAP−/− Vβ5 mice. We have found that SLAP−/− OT-1 mice have fewer CD8+ thymocytes but have increased CD5 expression. SLAP−/− OT-1 mice have fewer DP thymocytes expressing Vα2, signifying increased endogenous α-chain rearrangement, and more non-OVA-specific CD8+ splenocytes upon tetramer staining. Our data demonstrate that SLAP−/− Vβ5 mice also have fewer OVA-specific cells and increased Vα2 usage in the peripheral Vβ5 CD8+ T cells that were non-OVA-specific, demonstrating differences in α-chain repertoire. These studies provide direct evidence that increased TCR avidity in DP thymocytes enhances CD8+ T cell negative selection deleting thymocytes with specificity for cognate antigen, an antigen the mature T cells may never encounter. Collectively, these studies provide new insights into how TCR avidity during CD8+ T cell development influences repertoire selection. PMID:22956467

SLAP deficiency increases TCR avidity leading to altered repertoire and negative selection of cognate antigen-specific CD8+ T cells.

PubMed

Friend, Samantha F; Peterson, Lisa K; Kedl, Ross M; Dragone, Leonard L

2013-03-01

How T cell receptor (TCR) avidity influences CD8(+) T cell development and repertoire selection is not yet fully understood. To fill this gap, we utilized Src-like adaptor protein (SLAP)-deficient mice as a tool to increase TCR avidity on double positive (DP) thymocytes. We generated SLAP(-/-) mice with the transgenic MHC class I-restricted TCR (OT-1) and SLAP(-/-) Vβ5 mice, expressing only the β-chain of the TCR OT-1 transgene, to examine the effects of increased TCR surface levels on CD8(+) T cell development and repertoire selection. In comparing SLAP(-/-) OT-1 and Vβ5 mice with wild-type controls, we performed compositional analysis and assessed thymocyte signaling by measuring CD5 levels. In addition, we performed tetramer and compositional staining to measure affinity for the cognate antigen, ovalbumin (OVA) peptide, presented by MHC. Furthermore, we quantified differences in α-chain repertoire in SLAP(-/-) Vβ5 mice. We have found that SLAP(-/-) OT-1 mice have fewer CD8(+) thymocytes but have increased CD5 expression. SLAP(-/-) OT-1 mice have fewer DP thymocytes expressing Vα2, signifying increased endogenous α-chain rearrangement, and more non-OVA-specific CD8(+) splenocytes upon tetramer staining. Our data demonstrate that SLAP(-/-) Vβ5 mice also have fewer OVA-specific cells and increased Vα2 usage in the peripheral Vβ5 CD8(+) T cells that were non-OVA-specific, demonstrating differences in α-chain repertoire. These studies provide direct evidence that increased TCR avidity in DP thymocytes enhances CD8(+) T cell negative selection deleting thymocytes with specificity for cognate antigen, an antigen the mature T cells may never encounter. Collectively, these studies provide new insights into how TCR avidity during CD8(+) T cell development influences repertoire selection.

An optimized single chain TCR scaffold relying on the assembly with the native CD3-complex prevents residual mispairing with endogenous TCRs in human T-cells

PubMed Central

Knies, Diana; Klobuch, Sebastian; Xue, Shao-An; Birtel, Matthias; Echchannaoui, Hakim; Yildiz, Oezlem; Omokoko, Tana; Guillaume, Philippe; Romero, Pedro; Stauss, Hans; Sahin, Ugur; Herr, Wolfgang; Theobald, Matthias; Thomas, Simone; Voss, Ralf-Holger

2016-01-01

Immunotherapy of cancer envisions the adoptive transfer of T-cells genetically engineered with tumor-specific heterodimeric α/β T-cell receptors (TCRα/β). However, potential mispairing of introduced TCRα/β-chains with endogenous β/α-ones may evoke unpredictable autoimmune reactivities. A novel single chain (sc)TCR format relies on the fusion of the Vα-Linker-Vβ-fragment to the TCR Cβ-domain and coexpression of the TCR Cα-domain capable of recruiting the natural CD3-complex for full and hence, native T-cell signaling. Here, we tested whether such a gp100(280-288)- or p53(264-272) tumor antigen-specific scTCR is still prone to mispairing with TCRα. In a human Jurkat-76 T-cell line lacking endogenous TCRs, surface expression and function of a scTCR could be reconstituted by any cointroduced TCRα-chain indicating mispairing to take place on a molecular basis. In contrast, transduction into human TCRα/β-positive T-cells revealed that mispairing is largely reduced. Competition experiments in Jurkat-76 confirmed the preference of dcTCR to selfpair and to spare scTCR. This also allowed for the generation of dc/scTCR-modified cytomegalovirus/tumor antigen-bispecific T-cells to augment T-cell activation in CMV-infected tumor patients. Residual mispairing was prevented by strenghtening the Vα-Li-Vβ-fragment through the design of a novel disulfide bond between a Vα- and a linker-resident residue close to Vβ. Multimer-stainings, and cytotoxicity-, IFNγ-secretion-, and CFSE-proliferation-assays, the latter towards dendritic cells endogenously processing RNA-electroporated gp100 antigen proved the absence of hybrid scTCR/TCRα-formation without impairing avidity of scTCR/Cα in T-cells. Moreover, a fragile cytomegalovirus pp65(495-503)-specific scTCR modified this way acquired enhanced cytotoxicity. Thus, optimized scTCR/Cα inhibits residual TCR mispairing to accomplish safe adoptive immunotherapy for bulk endogenous TCRα/β-positive T-cells. PMID:27028870

Evolution of the antigen-specific CD8+ TCR repertoire across the life span: evidence for clonal homogenization of the old TCR repertoire.

PubMed

Rudd, Brian D; Venturi, Vanessa; Davenport, Miles P; Nikolich-Zugich, Janko

2011-02-15

Defects in T cell responses against pathogens and reduced diversity of TCRs have been described at both extremes of the life span. Yet, we still lack information on how Ag-specific T cell populations are maintained and/or altered from birth to old age. In this study, for the first time to our knowledge, we provide insight into Ag-specific TCR repertoire changes over the life span at the single-cell level. We have examined the TCR diversity of the primary CD8(+) T cell response to the immunodominant HSV-1 epitope HSV glycoprotein B 495-502 (HSV gB(498-505); SSIEFARL) (gB-8p) in neonatal, adult, and old C57BL/6 mice. The global distinctive features of the gB-8p-specific TCR repertoire were preserved in mice of different ages. However, both old and especially neonatal mice exhibited significant decreases in TCR diversity compared with that of adult mice. Still, although the neonatal Ag-specific repertoire comprised expectedly shorter germline-biased CDR3β lengths, the repertoire was surprisingly complex, and only a minority of responding cells lacked random nucleotide additions. Changes with aging included increased use of the already dominant TCRVβ10 family, a trend for lower content of the TCR containing the germline WG motif in the CDR3, and a remarkable sharing of one dominant clonotype between individual old mice, implying operation of selective mechanisms. Implications for the rational design of vaccines for neonates and the elderly are discussed.

Deconstructing Ras Signaling in the Thymus

PubMed Central

Kortum, Robert L.; Sommers, Connie L.; Pinski, John M.; Alexander, Clayton P.; Merrill, Robert K.; Li, Wenmei; Love, Paul E.

2012-01-01

Thymocytes must transit at least two distinct developmental checkpoints, governed by signals that emanate from either the pre-T cell receptor (pre-TCR) or the TCR to the small G protein Ras before emerging as functional T lymphocytes. Recent studies have shown a role for the Ras guanine exchange factor (RasGEF) Sos1 at the pre-TCR checkpoint. At the second checkpoint, the quality of signaling through the TCR is interrogated to ensure the production of an appropriate T cell repertoire. Although RasGRP1 is the only confirmed RasGEF required at the TCR checkpoint, current models suggest that the intensity and character of Ras activation, facilitated by both Sos and RasGRP1, will govern the boundary between survival (positive selection) and death (negative selection) at this stage. Using mouse models, we have assessed the independent and combined roles for the RasGEFs Sos1, Sos2, and RasGRP1 during thymocyte development. Although Sos1 was the dominant RasGEF at the pre-TCR checkpoint, combined Sos1/RasGRP1 deletion was required to effectively block development at this stage. Conversely, while RasGRP1 deletion efficiently blocked positive selection, combined RasGRP1/Sos1 deletion was required to block negative selection. This functional redundancy in RasGEFs during negative selection may act as a failsafe mechanism ensuring appropriate central tolerance. PMID:22586275

The adapter protein SLP-76 mediates "outside-in" integrin signaling and function in T cells.

PubMed

Baker, R G; Hsu, C J; Lee, D; Jordan, M S; Maltzman, J S; Hammer, D A; Baumgart, T; Koretzky, G A

2009-10-01

The adapter protein SH2 domain-containing leukocyte protein of 76 kDa (SLP-76) is an essential mediator of signaling from the T-cell antigen receptor (TCR). We report here that SLP-76 also mediates signaling downstream of integrins in T cells and that SLP-76-deficient T cells fail to support adhesion to integrin ligands. In response to both TCR and integrin stimulation, SLP-76 relocalizes to surface microclusters that colocalize with phosphorylated signaling proteins. Disruption of SLP-76 recruitment to the protein named LAT (linker for activation of T cells) inhibits SLP-76 clustering downstream of the TCR but not downstream of integrins. Conversely, an SLP-76 mutant unable to bind ADAP (adhesion and degranulation-promoting adapter protein) forms clusters following TCR but not integrin engagement and fails to support T-cell adhesion to integrin ligands. These findings demonstrate that SLP-76 relocalizes to integrin-initiated signaling complexes by a mechanism different from that employed during TCR signaling and that SLP-76 relocalization corresponds to SLP-76-dependent integrin function in T cells.

Ikaros promotes rearrangement of TCR alpha genes in an Ikaros null thymoma cell line

PubMed Central

Collins, Bernard; Clambey, Eric T.; Scott-Browne, James; White, Janice; Marrack, Philippa; Hagman, James; Kappler, John W.

2014-01-01

Summary Ikaros is important in the development and maintenance of the lymphoid system, functioning in part by associating with chromatin-remodeling complexes. We have studied the functions of Ikaros in the transition from pre-T cell to the CD4+CD8+ thymocyte using an Ikaros null CD4−CD8− mouse thymoma cell line (JE131). We demonstrate that this cell line carries a single functional TCR β gene rearrangement and expresses a surface pre-TCR. JE131 cells also carry non-functional rearrangements on both alleles of their TCR α loci. Retroviral re-introduction of Ikaros dramatically increased the rate of transcription in the α locus and TCR Vα/Jα recombination resulting in the appearance of many new αβTCR+ cells. The process is RAG dependent, requires SWI/SNF chromatin-remodeling complexes and is coincident with the binding of Ikaros to the TCR α enhancer. Furthermore, knockdown of Mi2/NuRD complexes increased the frequency of TCR α rearrangement. Our data suggest that Ikaros controls Vα/Jα recombination in T cells by controlling access of the transcription and recombination machinery to the TCR α loci. The JE131 cell line should prove to be a very useful tool for studying the molecular details of this and other processes involved in the pre-T cell to αβTCR+ CD4+CD8+ thymocyte transition. PMID:23172374

Ikaros promotes rearrangement of TCR α genes in an Ikaros null thymoma cell line.

PubMed

Collins, Bernard; Clambey, Eric T; Scott-Browne, James; White, Janice; Marrack, Philippa; Hagman, James; Kappler, John W

2013-02-01

Ikaros is important in the development and maintenance of the lymphoid system, functioning in part by associating with chromatin-remodeling complexes. We have studied the functions of Ikaros in the transition from pre-T cell to the CD4(+) CD8(+) thymocyte using an Ikaros null CD4(-) CD8(-) mouse thymoma cell line (JE131). We demonstrate that this cell line carries a single functional TCR β gene rearrangement and expresses a surface pre-TCR. JE131 cells also carry nonfunctional rearrangements on both alleles of their TCR α loci. Retroviral reintroduction of Ikaros dramatically increased the rate of transcription in the α locus and TCR Vα/Jα recombination resulting in the appearance of many new αβTCR(+) cells. The process is RAG dependent, requires switch/sucrose nonfermentable chromatin-remodeling complexes and is coincident with the binding of Ikaros to the TCR α enhancer. Furthermore, knockdown of Mi2/nucleosome remodeling and deacetylase complexes increased the frequency of TCR α rearrangement. Our data suggest that Ikaros controls Vα/Jα recombination in T cells by controlling access of the transcription and recombination machinery to the TCR α loci. The JE131 cell line should prove to be a very useful tool for studying the molecular details of this and other processes involved in the pre-T cell to αβTCR(+) CD4(+) CD8(+) thymocyte transition. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Measuring Soil Moisture using the Signal Strength of Buried Bluetooth Devices.

NASA Astrophysics Data System (ADS)

Hut, R.; Campbell, C. S.

2015-12-01

A low power bluetooth Low Energy (BLE) device is burried 20cm into the soil and a smartphone is placed on top of the soil to test if bluetooth signal strength can be related to soil moisture. The smartphone continuesly records and stores bluetooth signal strength of the device. The soil is artifcially wetted and drained. Results show a relation between BLE signal strength and soil moisture that could be used to measure soil moisture using these off-the-shelf consumer electronics. This opens the possibily to develop sensors that can be buried into the soil, possibly below the plow-line. These sensors can measure local parameters such as electric conductivity, ph, pressure, etc. Readings would be uploaded to a device on the surface using BLE. The signal strength of this BLE would be an (additional) measurement of soil moisture.

Bcl-2-interacting mediator of cell death influences autoantigen-driven deletion and TCR revision

PubMed Central

Hale, J. Scott; Nelson, Lisa T.; Simmons, Kalynn B.; Fink, Pamela J.

2010-01-01

Peripheral CD4+Vβ5+ T cells are tolerized to an endogenous mouse mammary tumor virus superantigen either by deletion or TCR revision. Through TCR revision, RAG reexpression mediates extrathymic TCRβ rearrangement and results in a population of post-revision CD4+Vβ5− T cells expressing revised TCRβ chains. We have hypothesized that cell death pathways regulate the selection of cells undergoing TCR revision to ensure the safety and utility of the post-revision population. Here, we investigate the role of Bim-mediated cell death in autoantigen-driven deletion and TCR revision. Bim deficiency and Bcl-2 overexpression in Vβ5 transgenic (Tg) mice both impair peripheral deletion. Vβ5 Tg Bim deficient and Bcl-2 Tg mice exhibit an elevated frequency of CD4+ T cells expressing both the transgene-encoded Vβ5 chain and a revised TCRβ chain. We now show that these dual-TCR expressing cells are TCR revision intermediates, and that the population of RAG-expressing, revising CD4+ T cells is increased in Bim deficient Vβ5 Tg mice. These findings support a role for Bim and Bcl-2 in regulating the balance of survival versus apoptosis in peripheral T cells undergoing RAG-dependent TCR rearrangements during TCR revision, thereby ensuring the utility of the post-revision repertoire. PMID:21148799

Bcl-2-interacting mediator of cell death influences autoantigen-driven deletion and TCR revision.

PubMed

Hale, J Scott; Nelson, Lisa T; Simmons, Kalynn B; Fink, Pamela J

2011-01-15

Peripheral CD4(+)Vβ5(+) T cells are tolerized to an endogenous mouse mammary tumor virus superantigen either by deletion or TCR revision. Through TCR revision, RAG reexpression mediates extrathymic TCRβ rearrangement and results in a population of postrevision CD4(+)Vβ5(-) T cells expressing revised TCRβ chains. We have hypothesized that cell death pathways regulate the selection of cells undergoing TCR revision to ensure the safety and utility of the postrevision population. In this study, we investigate the role of Bcl-2-interacting mediator of cell death (Bim)-mediated cell death in autoantigen-driven deletion and TCR revision. Bim deficiency and Bcl-2 overexpression in Vβ5 transgenic (Tg) mice both impair peripheral deletion. Vβ5 Tg Bim-deficient and Bcl-2 Tg mice exhibit an elevated frequency of CD4(+) T cells expressing both the transgene-encoded Vβ5 chain and a revised TCRβ chain. We now show that these dual-TCR-expressing cells are TCR revision intermediates and that the population of RAG-expressing, revising CD4(+) T cells is increased in Bim-deficient Vβ5 Tg mice. These findings support a role for Bim and Bcl-2 in regulating the balance of survival versus apoptosis in peripheral T cells undergoing RAG-dependent TCR rearrangements during TCR revision, thereby ensuring the utility of the postrevision repertoire.

47 CFR 80.753 - Signal strength requirements at the service area contour.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 47 Telecommunication 5 2010-10-01 2010-10-01 false Signal strength requirements at the service... Station VHF Coverage § 80.753 Signal strength requirements at the service area contour. (a) The requirements for reception by a marine VHF shipboard receiver are satisfied if the field strength from the...

47 CFR 80.753 - Signal strength requirements at the service area contour.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 47 Telecommunication 5 2011-10-01 2011-10-01 false Signal strength requirements at the service... Station VHF Coverage § 80.753 Signal strength requirements at the service area contour. (a) The requirements for reception by a marine VHF shipboard receiver are satisfied if the field strength from the...

A Co-Receptor Independent Transgenic Human TCR Mediates Anti-Tumor and Anti-Self Immunity in Mice

PubMed Central

Mehrotra, Shikhar; Al-Khami, Amir A.; Klarquist, Jared; Husain, Shahid; Naga, Osama; Eby, Jonathan M.; Murali, Anuradha K.; Lyons, Gretchen E.; Li, Mingli; Spivey, Natali D.; Norell, Håkan; Martins da Palma, Telma; Onicescu, Georgiana; Diaz-Montero, C. Marcela; Garrett-Mayer, Elizabeth; Cole, David J.; Le Poole, I. Caroline; Nishimura, Michael I.

2013-01-01

Recent advancements in T cell immunotherapy suggest that T cells engineered with high affinity T cell receptors (TCR) can offer better tumor regression. However, whether a high affinity TCR alone is sufficient to control tumor growth, or the T cell subset bearing the TCR is also important remains unclear. Using the human tyrosinase epitope reactive, CD8 independent, high affinity TCR isolated from MHC class-I restricted CD4+ T cells obtained from tumor infiltrating lymphocytes of a metastatic melanoma patient, we developed a novel TCR transgenic mouse with a C57BL/6 background. This HLA-A2 restricted TCR was positively selected on both CD4+ and CD8+ single-positive (SP) cells. However, when the TCR transgenic mouse was developed with an HLA-A2 background, the transgenic TCR was primarily expressed by CD3+CD4-CD8- double-negative (DN) T cells. TIL 1383I TCR transgenic CD4+, CD8+ and CD4-CD8- T cells were functional and retained the ability to control tumor growth without the need for vaccination or cytokine support in vivo. Furthermore, the HLA-A2+/human tyrosinase TCR double transgenic mice developed spontaneous hair depigmentation and had visual defects that progressed with age. Our data show that the expression of the high affinity TIL 1383I TCR alone in CD3+ T cells is sufficient to control the growth of murine and human melanoma and the presence or absence of CD4 and CD8 co-receptors had little effect on its functional capacity. PMID:22798675

Vehicle Motions as Inferred from Radio-signal- Strength Records

NASA Technical Reports Server (NTRS)

Pilkington, W. C.

1958-01-01

Considerable data on various parameters of a satellite and its launching vehicle can be obtained from the received-signal-strength records at the various satellite receiver stations. The doppler shifts of the satellite can be used to determine the orbital characteristics quite accurately. (Ref. 1 discusses the rapid determination of these parameters.) Information concerning the velocity increment and direction of each stage can be obtained if doppler data are received by several stations during the high-speed staging. The variations in signal strength during launch can be used to find the spin rate of the satellite before, during, and after staging, and to find variations in attitude, including precession, during launch. With spin and precession frequencies available, any changes in the ratios of moments of inertia can be determined. In this paper the launchings of the Explorer satellites will be used as a basis for the generalized conclusions concerning utilization of radio-signal-strength records. The discussion includes a description of the Explorer launching system.

Monoclonal TCR-redirected tumor cell killing.

PubMed

Liddy, Nathaniel; Bossi, Giovanna; Adams, Katherine J; Lissina, Anna; Mahon, Tara M; Hassan, Namir J; Gavarret, Jessie; Bianchi, Frayne C; Pumphrey, Nicholas J; Ladell, Kristin; Gostick, Emma; Sewell, Andrew K; Lissin, Nikolai M; Harwood, Naomi E; Molloy, Peter E; Li, Yi; Cameron, Brian J; Sami, Malkit; Baston, Emma E; Todorov, Penio T; Paston, Samantha J; Dennis, Rebecca E; Harper, Jane V; Dunn, Steve M; Ashfield, Rebecca; Johnson, Andy; McGrath, Yvonne; Plesa, Gabriela; June, Carl H; Kalos, Michael; Price, David A; Vuidepot, Annelise; Williams, Daniel D; Sutton, Deborah H; Jakobsen, Bent K

2012-06-01

T cell immunity can potentially eradicate malignant cells and lead to clinical remission in a minority of patients with cancer. In the majority of these individuals, however, there is a failure of the specific T cell receptor (TCR)–mediated immune recognition and activation process. Here we describe the engineering and characterization of new reagents termed immune-mobilizing monoclonal TCRs against cancer (ImmTACs). Four such ImmTACs, each comprising a distinct tumor-associated epitope-specific monoclonal TCR with picomolar affinity fused to a humanized cluster of differentiation 3 (CD3)-specific single-chain antibody fragment (scFv), effectively redirected T cells to kill cancer cells expressing extremely low surface epitope densities. Furthermore, these reagents potently suppressed tumor growth in vivo. Thus, ImmTACs overcome immune tolerance to cancer and represent a new approach to tumor immunotherapy.

Male facial appearance signals physical strength to women.

PubMed

Fink, Bernhard; Neave, Nick; Seydel, Hanna

2007-01-01

Previous studies showed that male faces with extreme features that are likely to be associated with testosterone (T) are perceived as dominant and masculine. Women were reported to prefer masculinized male faces, as they may consider T markers to be an "honest" indication of good health. However, it is also likely that female preferences for certain male faces arise from the fact that dominant- and masculine-looking males are signaling characteristics which may be beneficial in intrasexual conflict, and thereby also indicate potential achievers of high status, an important factor in female mate selection. Although numerous studies were built on this assumption, nothing is known about the relationship between perceived facial dominance and physical strength in men. We measured hand-grip strength, as a measure of overall physical strength, in a sample of 32 male students, and recorded age, body weight, and height. Seventy-nine women rated facial images of these men for dominance, masculinity, and attractiveness. After controlling for age and body weight, hand-grip strength was found to correlate significantly positively with all three measures. The present data thus support the supposition that a male's physical strength is also signaled via facial characteristics of dominance and masculinity, which are considered attractive by women.

Mitochondria-derived hydrogen peroxide selectively enhances T cell receptor-initiated signal transduction.

PubMed

Gill, Tejpal; Levine, Alan D

2013-09-06

T cell receptor (TCR)-initiated signal transduction is reported to increase production of intracellular reactive oxygen species, such as superoxide (O2˙(-)) and hydrogen peroxide (H2O2), as second messengers. Although H2O2 can modulate signal transduction by inactivating protein phosphatases, the mechanism and the subcellular localization of intracellular H2O2 as a second messenger of the TCR are not known. The antioxidant enzyme superoxide dismutase (SOD) catalyzes the dismutation of highly reactive O2˙(-) into H2O2 and thus acts as an intracellular generator of H2O2. As charged O2˙(-) is unable to diffuse through intracellular membranes, cells express distinct SOD isoforms in the cytosol (Cu,Zn-SOD) and mitochondria (Mn-SOD), where they locally scavenge O2˙(-) leading to production of H2O2. A 2-fold organelle-specific overexpression of either SOD in Jurkat T cell lines increases intracellular production of H2O2 but does not alter the levels of intracellular H2O2 scavenging enzymes such as catalase, membrane-bound peroxiredoxin1 (Prx1), and cytosolic Prx2. We report that overexpression of Mn-SOD enhances tyrosine phosphorylation of TCR-associated membrane proximal signal transduction molecules Lck, LAT, ZAP70, PLCγ1, and SLP76 within 1 min of TCR cross-linking. This increase in mitochondrial H2O2 specifically modulates MAPK signaling through the JNK/cJun pathway, whereas overexpressing Cu,Zn-SOD had no effect on any of these TCR-mediated signaling molecules. As mitochondria translocate to the immunological synapse during TCR activation, we hypothesize this translocation provides the effective concentration of H2O2 required to selectively modulate downstream signal transduction pathways.

A CD22-reactive TCR from the T-cell allorepertoire for the treatment of acute lymphoblastic leukemia by TCR gene transfer

PubMed Central

Jahn, Lorenz; Hagedoorn, Renate S.; van der Steen, Dirk M.; Hombrink, Pleun; Kester, Michel G.D.; Schoonakker, Marjolein P.; de Ridder, Daniëlle; van Veelen, Peter A.; Falkenburg, J.H. Frederik; Heemskerk, Mirjam H.M.

2016-01-01

CD22 is currently evaluated as a target-antigen for the treatment of B-cell malignancies using chimeric antigen receptor (CAR)-engineered T-cells or monoclonal antibodies (mAbs). CAR- and mAbs-based immunotherapies have been successfully applied targeting other antigens, however, occurrence of refractory disease to these interventions urges the identification of additional strategies. Here, we identified a TCR recognizing the CD22-derived peptide RPFPPHIQL (CD22RPF) presented in human leukocyte antigen (HLA)-B*07:02. To overcome tolerance to self-antigens such as CD22, we exploited the immunogenicity of allogeneic HLA. CD22RPF-specific T-cell clone 9D4 was isolated from a healthy HLA-B*07:02neg individual, efficiently produced cytokines upon stimulation with primary acute lymphoblastic leukemia and healthy B-cells, but did not react towards healthy hematopoietic and nonhematopoietic cell subsets, including dendritic cells (DCs) and macrophages expressing low levels of CD22. Gene transfer of TCR-9D4 installed potent CD22-specificity onto recipient CD8+ T-cells that recognized and lysed primary B-cell leukemia. TCR-transduced T-cells spared healthy CD22neg hematopoietic cell subsets but weakly lysed CD22low-expressing DCs and macrophages. CD22-specific TCR-engineered T-cells could form an additional immunotherapeutic strategy with a complementary role to CAR- and antibody-based interventions in the treatment of B-cell malignancies. However, CD22 expression on non-B-cells may limit the attractiveness of CD22 as target-antigen in cellular immunotherapy. PMID:27689397

A CD22-reactive TCR from the T-cell allorepertoire for the treatment of acute lymphoblastic leukemia by TCR gene transfer.

PubMed

Jahn, Lorenz; Hagedoorn, Renate S; van der Steen, Dirk M; Hombrink, Pleun; Kester, Michel G D; Schoonakker, Marjolein P; de Ridder, Daniëlle; van Veelen, Peter A; Falkenburg, J H Frederik; Heemskerk, Mirjam H M

2016-11-01

CD22 is currently evaluated as a target-antigen for the treatment of B-cell malignancies using chimeric antigen receptor (CAR)-engineered T-cells or monoclonal antibodies (mAbs). CAR- and mAbs-based immunotherapies have been successfully applied targeting other antigens, however, occurrence of refractory disease to these interventions urges the identification of additional strategies. Here, we identified a TCR recognizing the CD22-derived peptide RPFPPHIQL (CD22RPF) presented in human leukocyte antigen (HLA)-B*07:02. To overcome tolerance to self-antigens such as CD22, we exploited the immunogenicity of allogeneic HLA. CD22RPF-specific T-cell clone 9D4 was isolated from a healthy HLA-B*07:02neg individual, efficiently produced cytokines upon stimulation with primary acute lymphoblastic leukemia and healthy B-cells, but did not react towards healthy hematopoietic and nonhematopoietic cell subsets, including dendritic cells (DCs) and macrophages expressing low levels of CD22. Gene transfer of TCR-9D4 installed potent CD22-specificity onto recipient CD8+ T-cells that recognized and lysed primary B-cell leukemia. TCR-transduced T-cells spared healthy CD22neg hematopoietic cell subsets but weakly lysed CD22low-expressing DCs and macrophages. CD22-specific TCR-engineered T-cells could form an additional immunotherapeutic strategy with a complementary role to CAR- and antibody-based interventions in the treatment of B-cell malignancies. However, CD22 expression on non-B-cells may limit the attractiveness of CD22 as target-antigen in cellular immunotherapy.

CD8+ TCR repertoire formation is guided primarily by the peptide component of the antigenic complex.

PubMed

Koning, Dan; Costa, Ana I; Hoof, Ilka; Miles, John J; Nanlohy, Nening M; Ladell, Kristin; Matthews, Katherine K; Venturi, Vanessa; Schellens, Ingrid M M; Borghans, Jose A M; Kesmir, Can; Price, David A; van Baarle, Debbie

2013-02-01

CD8(+) T cells recognize infected or dysregulated cells via the clonotypically expressed αβ TCR, which engages Ag in the form of peptide bound to MHC class I (MHC I) on the target cell surface. Previous studies have indicated that a diverse Ag-specific TCR repertoire can be beneficial to the host, yet the determinants of clonotypic diversity are poorly defined. To better understand the factors that govern TCR repertoire formation, we conducted a comprehensive clonotypic analysis of CD8(+) T cell populations directed against epitopes derived from EBV and CMV. Neither pathogen source nor the restricting MHC I molecule were linked with TCR diversity; indeed, both HLA-A and HLA-B molecules were observed to interact with an overlapping repertoire of expressed TRBV genes. Peptide specificity, however, markedly impacted TCR diversity. In addition, distinct peptides sharing HLA restriction and viral origin mobilized TCR repertoires with distinct patterns of TRBV gene usage. Notably, no relationship was observed between immunodominance and TCR diversity. These findings provide new insights into the forces that shape the Ag-specific TCR repertoire in vivo and highlight a determinative role for the peptide component of the peptide-MHC I complex on the molecular frontline of CD8(+) T cell-mediated immune surveillance.

Crammed signaling motifs in the T-cell receptor.

PubMed

Borroto, Aldo; Abia, David; Alarcón, Balbino

2014-09-01

Although the T cell antigen receptor (TCR) is long known to contain multiple signaling subunits (CD3γ, CD3δ, CD3ɛ and CD3ζ), their role in signal transduction is still not well understood. The presence of at least one immunoreceptor tyrosine-based activation motif (ITAM) in each CD3 subunit has led to the idea that the multiplication of such elements essentially serves to amplify signals. However, the evolutionary conservation of non-ITAM sequences suggests that each CD3 subunit is likely to have specific non-redundant roles at some stage of development or in mature T cell function. The CD3ɛ subunit is paradigmatic because in a relatively short cytoplasmic sequence (∼55 amino acids) it contains several docking sites for proteins involved in intracellular trafficking and signaling, proteins whose relevance in T cell activation is slowly starting to be revealed. In this review we will summarize our current knowledge on the signaling effectors that bind directly to the TCR and we will propose a hierarchy in their response to TCR triggering. Copyright © 2014 Elsevier B.V. All rights reserved.

T follicular helper and T follicular regulatory cells have different TCR specificity

PubMed Central

Maceiras, Ana Raquel; Almeida, Silvia Cristina Paiva; Mariotti-Ferrandiz, Encarnita; Chaara, Wahiba; Jebbawi, Fadi; Six, Adrien; Hori, Shohei; Klatzmann, David; Faro, Jose; Graca, Luis

2017-01-01

Immunization leads to the formation of germinal centres (GCs) that contain both T follicular helper (Tfh) and T follicular regulatory (Tfr) cells. Whether T-cell receptor (TCR) specificity defines the differential functions of Tfh and Tfr cells is unclear. Here we show that antigen-specific T cells after immunization are preferentially recruited to the GC to become Tfh cells, but not Tfr cells. Tfh cells, but not Tfr cells, also proliferate efficiently on restimulation with the same immunizing antigen in vitro. Ex vivo TCR repertoire analysis shows that immunization induces oligoclonal expansion of Tfh cells. By contrast, the Tfr pool has a TCR repertoire that more closely resembles that of regulatory T (Treg) cells. Our data thus indicate that the GC Tfh and Tfr pools are generated from distinct TCR repertoires, with Tfh cells expressing antigen-responsive TCRs to promote antibody responses, and Tfr cells expressing potentially autoreactive TCRs to suppress autoimmunity. PMID:28429709

Cutting Edge: TCR Revision Affects Predominantly Foxp3− Cells and Skews Them toward the Th17 Lineage1

PubMed Central

Zehn, Dietmar; Bevan, Michael J.; Fink, Pamela J.

2009-01-01

CD4+ T cells respond to peripheral endogenous superantigen stimulation by undergoing deletion or TCR revision. The latter involves RAG re-expression, TCR gene rearrangement, and expression of a novel TCR. TCR-revised T cells are functional and express a diverse TCR repertoire. Because TCR revision harbors the potential to create self-reactivity, it is important to explore whether T cells known to be self-reactive (regulatory T cells) or those involved in autoimmunity (Th17 cells) arise from TCR revision. Interestingly, we observed that Foxp3+ cells are excluded from revising their TCR and that only a small fraction of postrevision cells expresses Foxp3. In contrast, Th17 cells are 20 times more frequent among revised than among C57BL/6 CD4+ T cells, indicating that postrevision cells are biased toward the Th17 lineage. The link between Th17 differentiation and TCR revision might be highly relevant to the role of Th17 cells in promoting autoimmunity. PMID:17947636

TCR-pMHC encounter differentially regulates transcriptomes of tissue-resident CD8 T cells.

PubMed

Yoshizawa, Akihiro; Bi, Kevin; Keskin, Derin B; Zhang, Guanglan; Reinhold, Bruce; Reinherz, Ellis L

2018-01-01

To investigate the role of TCR-pMHC interaction in regulating lung CD8 tissue-resident T cell (T R ) differentiation, polyclonal responses were compared against NP 366-374 /D b and PA 224-233 /D b , two immunodominant epitopes that arise during influenza A infection in mice. Memory niches distinct from iBALTs develop within the lamina propria, supporting CD103 + and CD103 - CD8 T R generation and intraepithelial translocation. Gene set enrichment analysis (GSEA) and weighted gene co-expression network analysis (WGCNA) identify dominant TCR, adherens junction, RIG-I-like and NOD-like pattern recognition receptor as well as TGF-β signaling pathways and memory signatures among PA 224-233 /D b T cells consistent with T resident memory (T RM ) status. In contrast, NP 366-374 /D b T cells exhibit enrichment of effector signatures, upregulating pro-inflammatory mediators even among T RM . While NP 366-374 /D b T cells manifest transcripts linked to canonical exhaustion pathways, PA 224-233 /D b T cells exploit P2rx7 purinoreceptor attenuation. The NP 366-374 /D b CD103 + subset expresses the antimicrobial lactotransferrin whereas PA 224-233 /D b CD103 + utilizes pore-forming mpeg-1, with <22% of genes correspondingly upregulated in CD103 + (or CD103 - ) subsets of both specificities. Thus, TCR-pMHC interactions among T R and antigen presenting cells in a tissue milieu strongly impact CD8 T cell biology. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Intrathymic selection of NK1.1+α/β T cell antigen receptor (TCR)+ cells in transgenic mice bearing TCR specific for chicken ovalbumin and restricted to I-Ad

PubMed Central

Iwabuchi, Chikako; Iwabuchi, Kazuya; Nakagawa, Ken-ichi; Takayanagi, Toshiaki; Nishihori, Hiroki; Tone, Saori; Ogasawara, Kazumasa; Good, Robert A.; Onoé, Kazunori

1998-01-01

Generation and negative selection of NK1.1+α/β T cell receptor (TCR)+ thymocytes were analyzed using TCR-transgenic (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice and Rag-1−/−/DO10 mice, which had been established by breeding and backcrossing between Rag-1−/− and DO10 mice. Almost all T cells from these mice were shown to bear Vα13/Vβ8.2 that is specific for chicken ovalbumin (cOVA) and restricted to I-Ad. A normal proportion of the NK1.1+ Vα13/Vβ8.2+ thymocytes was generated in these mice. However, the actual cell number of both NK1.1+ and NK1.1− thymocytes in I-Ad/d mice (positive selecting background) was larger than that in I-Ab/d mice (negative selecting background). Markedly low but significant proportions of NK1.1+ Vα13/Vβ8.2+ cells were detected in the spleens from I-Ad/d and I-Ab/d mice. It was shown that the splenic NK1.1+ T cells of the I-Ab/d mice were anergized against stimulation through TCR. When (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice were given cOVA, extensive or intermediate elimination of NK1.1+α/βTCR+ thymocytes was induced in I-Ad/d or I-Ab/d mice, respectively. However, the clonal elimination was not as complete as that seen in the major NK1.1− thymocyte population. The present findings indicate that normal generation of NK1.1+α/βTCR+ thymocytes occurs in the absence of Vα14-Jα281 and that substantial negative selection operates on the NK1.1+α/βTCR+ cells. PMID:9653164

The activation threshold of CD4+ T cells is defined by TCR/peptide-MHC class II interactions in the thymic medulla.

PubMed

Stephen, Tom Li; Tikhonova, Anastasia; Riberdy, Janice M; Laufer, Terri M

2009-11-01

Immature thymocytes that are positively selected based upon their response to self-peptide-MHC complexes develop into mature T cells that are not overtly reactive to those same complexes. Developmental tuning is the active process through which TCR-associated signaling pathways of single-positive thymocytes are attenuated to respond appropriately to the peptide-MHC molecules that will be encountered in the periphery. In this study, we explore the mechanisms that regulate the tuning of CD4(+) single-positive T cells to MHC class II encountered in the thymic medulla. Experiments with murine BM chimeras demonstrate that tuning can be mediated by MHC class II expressed by either thymic medullary epithelial cells or thymic dendritic cells. Tuning does not require the engagement of CD4 by MHC class II on stromal cells. Rather, it is mediated by interactions between MHC class II and the TCR. To understand the molecular changes that distinguish immature hyperactive T cells from tuned mature CD4(+) T cells, we compared their responses to TCR stimulation. The altered response of mature CD4 single-positive thymocytes is characterized by the inhibition of ERK activation by low-affinity self-ligands and increased expression of the inhibitory tyrosine phosphatase SHP-1. Thus, persistent TCR engagement by peptide-MHC class II on thymic medullary stroma inhibits reactivity to self-Ags and prevents autoreactivity in the mature repertoire.

Regulation of T-cell receptor signalling by membrane microdomains

PubMed Central

Razzaq, Tahir M; Ozegbe, Patricia; Jury, Elizabeth C; Sembi, Phupinder; Blackwell, Nathan M; Kabouridis, Panagiotis S

2004-01-01

There is now considerable evidence suggesting that the plasma membrane of mammalian cells is compartmentalized by functional lipid raft microdomains. These structures are assemblies of specialized lipids and proteins and have been implicated in diverse biological functions. Analysis of their protein content using proteomics and other methods revealed enrichment of signalling proteins, suggesting a role for these domains in intracellular signalling. In T lymphocytes, structure/function experiments and complementary pharmacological studies have shown that raft microdomains control the localization and function of proteins which are components of signalling pathways regulated by the T-cell antigen receptor (TCR). Based on these studies, a model for TCR phosphorylation in lipid rafts is presented. However, despite substantial progress in the field, critical questions remain. For example, it is unclear if membrane rafts represent a homogeneous population and if their structure is modified upon TCR stimulation. In the future, proteomics and the parallel development of complementary analytical methods will undoubtedly contribute in further delineating the role of lipid rafts in signal transduction mechanisms. PMID:15554919

Development of nanoscale structure in LAT-based signaling complexes

PubMed Central

2016-01-01

ABSTRACT The adapter molecule linker for activation of T cells (LAT) plays a crucial role in forming signaling complexes induced by stimulation of the T cell receptor (TCR). These multi-molecular complexes are dynamic structures that activate highly regulated signaling pathways. Previously, we have demonstrated nanoscale structure in LAT-based complexes where the adapter SLP-76 (also known as LCP2) localizes to the periphery of LAT clusters. In this study, we show that initially LAT and SLP-76 are randomly dispersed throughout the clusters that form upon TCR engagement. The segregation of LAT and SLP-76 develops near the end of the spreading process. The local concentration of LAT also increases at the same time. Both changes require TCR activation and an intact actin cytoskeleton. These results demonstrate that the nanoscale organization of LAT-based signaling complexes is dynamic and indicates that different kinds of LAT-based complexes appear at different times during T cell activation. PMID:27875277

Multi-Band Received Signal Strength Fingerprinting Based Indoor Location System

NASA Astrophysics Data System (ADS)

Sertthin, Chinnapat; Fujii, Takeo; Ohtsuki, Tomoaki; Nakagawa, Masao

This paper proposes a new multi-band received signal strength (MRSS) fingerprinting based indoor location system, which employs the frequency diversity on the conventional single-band received signal strength (RSS) fingerprinting based indoor location system. In the proposed system, the impacts of frequency diversity on the enhancements of positioning accuracy are analyzed. Effectiveness of the proposed system is proved by experimental approach, which was conducted in non line-of-sight (NLOS) environment under the area of 103m2 at Yagami Campus, Keio University. WLAN access points, which simultaneously transmit dual-band signal of 2.4 and 5.2GHz, are utilized as transmitters. Likewise, a dual-band WLAN receiver is utilized as a receiver. Signal distances calculated by both Manhattan and Euclidean were classified by K-Nearest Neighbor (KNN) classifier to illustrate the performance of the proposed system. The results confirmed that Frequency diversity attributions of multi-band signal provide accuracy improvement over 50% of the conventional single-band.

Calculating distance by wireless ethernet signal strength for global positioning method

NASA Astrophysics Data System (ADS)

Kim, Seung-Yong; Kim, Jeehong; Lee, Chang-goo

2005-12-01

This paper investigated mobile robot localization by using wireless Ethernet for global localization and INS for relative localization. For relative localization, the low-cost INS features self-contained was adopted. Low-cost MEMS-based INS has a short-period response and acceptable performance. Generally, variety sensor was used for mobile robot localization. In spite of precise modeling of the sensor, it leads inevitably to the accumulation of errors. The IEEE802.11b wireless Ethernet standard has been deployed in office building, museums, hospitals, shopping centers and other indoor environments. Many mobile robots already make use of wireless networking for communication. So location sensing with wireless Ethernet might be very useful for a low-cost robot. This research used wireless Ethernet card for compensation the accumulation of errors. So the mobile robot can use that for global localization through the installed many IEEE802.11b wireless Ethernets in indoor environments. The chief difficulty in localization with wireless Ethernet is predicting signal strength. As a sensor, RF signal strength measured indoors is non-linear with distance. So, there made the profiles of signal strength for points and used that. We wrote using function between signal strength profile and distance from the wireless Ethernet point.

Energetic and flexibility properties captured by long molecular dynamics simulations of a membrane-embedded pMHCII-TCR complex.

PubMed

Bello, Martiniano; Correa-Basurto, José

2016-04-01

Although crystallographic data have provided important molecular insight into the interactions in the pMHC-TCR complex, the inherent features of this structural approach cause it to only provide a static picture of the interactions. While unbiased molecular dynamics simulations (UMDSs) have provided important information about the dynamic structural behavior of the pMHC-TCR complex, most of them have modeled the pMHC-TCR complex as soluble, when in physiological conditions, this complex is membrane bound; therefore, following this latter UMDS protocol might hamper important dynamic results. In this contribution, we performed three independent 300 ns-long UMDSs of the pMHCII-TCR complex anchored in two opposing membranes to explore the structural and energetic properties of the recognition of pMHCII by the TCR. The conformational ensemble generated through UMDSs was subjected to clustering and Cartesian principal component analyses (cPCA) to explore the dynamical behavior of the pMHCII-TCR association. Furthermore, based on the conformational population sampled through UMDSs, the effective binding free energy, per-residue free energy decomposition, and alanine scanning mutations were explored for the native pMHCII-TCR complex, as well as for 12 mutations (p1-p12MHCII-TCR) introduced in the native peptide. Clustering analyses and cPCA provide insight into the rocking motion of the TCR onto pMHCII, together with the presence of new electrostatic interactions not observed through crystallographic methods. Energetic results provide evidence of the main contributors to the pMHC-TCR complex formation as well as the key residues involved in this molecular recognition process.

A novel T cell receptor single-chain signaling complex mediates antigen-specific T cell activity and tumor control

PubMed Central

Stone, Jennifer D.; Harris, Daniel T.; Soto, Carolina M.; Chervin, Adam S.; Aggen, David H.; Roy, Edward J.; Kranz, David M.

2014-01-01

Adoptive transfer of genetically modified T cells to treat cancer has shown promise in several clinical trials. Two main strategies have been applied to redirect T cells against cancer: 1) introduction of a full-length T cell receptor (TCR) specific for a tumor-associated peptide-MHC, or 2) introduction of a chimeric antigen receptor (CAR), including an antibody fragment specific for a tumor cell surface antigen, linked intracellularly to T cell signaling domains. Each strategy has advantages and disadvantages for clinical applications. Here, we present data on the in vitro and in vivo effectiveness of a single-chain signaling receptor incorporating a TCR variable fragment as the targeting element (referred to as TCR-SCS). This receptor contained a single-chain TCR (Vβ-linker-Vα) from a high-affinity TCR called m33, linked to the intracellular signaling domains of CD28 and CD3ζ. This format avoided mispairing with endogenous TCR chains, and mediated specific T cell activity when expressed in either CD4 or CD8 T cells. TCR-SCS-transduced CD8-negative cells showed an intriguing sensitivity, compared to full-length TCRs, to higher densities of less stable pepMHC targets. T cells that expressed this peptide-specific receptor persisted in vivo, and exhibited polyfunctional responses. Growth of metastatic antigen-positive tumors was significantly inhibited by T cells that expressed this receptor, and tumor cells that escaped were antigen loss variants. TCR-SCS receptors represent an alternative targeting receptor strategy that combines the advantages of single-chain expression, avoidance of TCR chain mispairing, and targeting of intracellular antigens presented in complex with MHC proteins. PMID:25082071

Generation of TCR-Expressing Innate Lymphoid-like Helper Cells that Induce Cytotoxic T Cell-Mediated Anti-leukemic Cell Response.

PubMed

Ueda, Norihiro; Uemura, Yasushi; Zhang, Rong; Kitayama, Shuichi; Iriguchi, Shoichi; Kawai, Yohei; Yasui, Yutaka; Tatsumi, Minako; Ueda, Tatsuki; Liu, Tian-Yi; Mizoro, Yasutaka; Okada, Chihiro; Watanabe, Akira; Nakanishi, Mahito; Senju, Satoru; Nishimura, Yasuharu; Kuzushima, Kiyotaka; Kiyoi, Hitoshi; Naoe, Tomoki; Kaneko, Shin

2018-06-05

CD4 + T helper (Th) cell activation is essential for inducing cytotoxic T lymphocyte (CTL) responses against malignancy. We reprogrammed a Th clone specific for chronic myelogenous leukemia (CML)-derived b3a2 peptide to pluripotency and re-differentiated the cells into original TCR-expressing T-lineage cells (iPS-T cells) with gene expression patterns resembling those of group 1 innate lymphoid cells. CD4 gene transduction into iPS-T cells enhanced b3a2 peptide-specific responses via b3a2 peptide-specific TCR. iPS-T cells upregulated CD40 ligand (CD40L) expression in response to interleukin-2 and interleukin-15. In the presence of Wilms tumor 1 (WT1) peptide, antigen-specific dendritic cells (DCs) conditioned by CD4-modified CD40L high iPS-T cells stimulated WT1-specific CTL priming, which eliminated WT1 peptide-expressing CML cells in vitro and in vivo. Thus, CD4 modification of CD40L high iPS-T cells generates innate lymphoid helper-like cells inducing bcr-abl-specific TCR signaling that mediates effectiveanti-leukemic CTL responses via DC maturation, showing potential for adjuvant immunotherapy against leukemia. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Artificial Neural Network-Based Early-Age Concrete Strength Monitoring Using Dynamic Response Signals.

PubMed

Kim, Junkyeong; Lee, Chaggil; Park, Seunghee

2017-06-07

Concrete is one of the most common materials used to construct a variety of civil infrastructures. However, since concrete might be susceptible to brittle fracture, it is essential to confirm the strength of concrete at the early-age stage of the curing process to prevent unexpected collapse. To address this issue, this study proposes a novel method to estimate the early-age strength of concrete, by integrating an artificial neural network algorithm with a dynamic response measurement of the concrete material. The dynamic response signals of the concrete, including both electromechanical impedances and guided ultrasonic waves, are obtained from an embedded piezoelectric sensor module. The cross-correlation coefficient of the electromechanical impedance signals and the amplitude of the guided ultrasonic wave signals are selected to quantify the variation in dynamic responses according to the strength of the concrete. Furthermore, an artificial neural network algorithm is used to verify a relationship between the variation in dynamic response signals and concrete strength. The results of an experimental study confirm that the proposed approach can be effectively applied to estimate the strength of concrete material from the early-age stage of the curing process.

Artificial Neural Network-Based Early-Age Concrete Strength Monitoring Using Dynamic Response Signals

PubMed Central

Kim, Junkyeong; Lee, Chaggil; Park, Seunghee

2017-01-01

Concrete is one of the most common materials used to construct a variety of civil infrastructures. However, since concrete might be susceptible to brittle fracture, it is essential to confirm the strength of concrete at the early-age stage of the curing process to prevent unexpected collapse. To address this issue, this study proposes a novel method to estimate the early-age strength of concrete, by integrating an artificial neural network algorithm with a dynamic response measurement of the concrete material. The dynamic response signals of the concrete, including both electromechanical impedances and guided ultrasonic waves, are obtained from an embedded piezoelectric sensor module. The cross-correlation coefficient of the electromechanical impedance signals and the amplitude of the guided ultrasonic wave signals are selected to quantify the variation in dynamic responses according to the strength of the concrete. Furthermore, an artificial neural network algorithm is used to verify a relationship between the variation in dynamic response signals and concrete strength. The results of an experimental study confirm that the proposed approach can be effectively applied to estimate the strength of concrete material from the early-age stage of the curing process. PMID:28590456

Activated PLC-γ1 is Catalytically Induced at LAT but Activated PLC-γ1 is Localized at both LAT- and TCR-Containing Complexes

PubMed Central

Cruz-Orcutt, Noemi; Vacaflores, Aldo; Connolly, Sean F.; Bunnell, Stephen C.; Houtman, Jon C.D.

2014-01-01

Phospholipase C-γ1 (PLC-γ1) is a key regulator of T cell receptor (TCR)-induced signaling. Activation of the TCR enhances PLC-γ1 enzymatic function, resulting in calcium influx and the activation of PKC family members and RasGRP. The current model is that phosphorylation of LAT tyrosine 132 facilitates the recruitment of PLC-γ1, leading to its activation and function at the LAT complex. In this study, we examined the phosphorylation kinetics of LAT and PLC-γ1 and the cellular localization of activated PLC-γ1. We observed that commencement of the phosphorylation of LAT tyrosine 132 and PLC-γ1 tyrosine 783 occurred simultaneously, supporting the current model. However, once begun, PLC-γ1 activation occurred more rapidly than LAT tyrosine 132. The association of LAT and PLC-γ1 was more transient than the interaction of LAT and Grb2 and a pool of activated PLC-γ1 translocated away from LAT to cellular structures containing the TCR. These studies demonstrate that LAT and PLC-γ1 form transient interactions that catalyze the activation of PLC-γ1, but that activated PLC-γ1 resides in both LAT and TCR clusters. Together, this work highlights that our current model is incomplete and the activation and function of PLC-γ1 in T cells is highly complex. PMID:24412752

Unique ζ-chain motifs mediate a direct TCR-actin linkage critical for immunological synapse formation and T-cell activation.

PubMed

Klieger, Yair; Almogi-Hazan, Osnat; Ish-Shalom, Eliran; Pato, Aviad; Pauker, Maor H; Barda-Saad, Mira; Wang, Lynn; Baniyash, Michal

2014-01-01

TCR-mediated activation induces receptor microclusters that evolve to a defined immune synapse (IS). Many studies showed that actin polymerization and remodeling, which create a scaffold critical to IS formation and stabilization, are TCR mediated. However, the mechanisms controlling simultaneous TCR and actin dynamic rearrangement in the IS are yet not fully understood. Herein, we identify two novel TCR ζ-chain motifs, mediating the TCR's direct interaction with actin and inducing actin bundling. While T cells expressing the ζ-chain mutated in these motifs lack cytoskeleton (actin) associated (cska)-TCRs, they express normal levels of non-cska and surface TCRs as cells expressing wild-type ζ-chain. However, such mutant cells are unable to display activation-dependent TCR clustering, IS formation, expression of CD25/CD69 activation markers, or produce/secrete cytokine, effects also seen in the corresponding APCs. We are the first to show a direct TCR-actin linkage, providing the missing gap linking between TCR-mediated Ag recognition, specific cytoskeleton orientation toward the T-cell-APC interacting pole and long-lived IS maintenance. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The pool of preactivated Lck in the initiation of T-cell signaling: a critical re-evaluation of the Lck standby model.

PubMed

Ballek, Ondřej; Valečka, Jan; Manning, Jasper; Filipp, Dominik

2015-04-01

The initiation of T-cell receptor (TCR) signaling, based on the cobinding of TCR and CD4-Lck heterodimer to a peptide-major histocompatibility complex II on antigen presenting cells, represents a classical model of T-cell signaling. What is less clear however, is the mechanism which translates TCR engagement to the phosphorylation of immunoreceptor tyrosine-based activation motifs on CD3 chains and how this event is coupled to the delivery of Lck function. Recently proposed 'standby model of Lck' posits that resting T-cells contain an abundant pool of constitutively active Lck (pY394(Lck)) required for TCR triggering, and this amount, upon TCR engagement, remains constant. Here, we show that although maintenance of the limited pool of pY394(Lck) is necessary for the generation of TCR proximal signals in a time-restricted fashion, the total amount of this pool, ~2%, is much smaller than previously reported (~40%). We provide evidence that this dramatic discrepancy in the content of pY394(Lck)is likely the consequence of spontaneous phosphorylation of Lck that occurred after cell solubilization. Additional discrepancies can be accounted for by the sensitivity of different pY394(Lck)-specific antibodies and the type of detergents used. These data suggest that reagents and conditions used for the quantification of signaling parameters must be carefully validated and interpreted. Thus, the limited size of pY394(Lck) pool in primary T-cells invites a discussion regarding the adjustment of the quantitative parameters of the standby model of Lck and reevaluation of the mechanism by which this pool contributes to the generation of proximal TCR signaling.

The discovery of a reciprocal relationship between tyrosine-kinase signaling and cullin neddylation.

PubMed

Friend, Samantha F; Peterson, Lisa K; Treacy, Eric; Stefanski, Adrianne L; Sosinowski, Tomasz; Pennock, Nathan D; Berger, Allison J; Winn, Virginia D; Dragone, Leonard L

2013-01-01

While neddylation is known to activate cullin (CUL)-RING ubiquitin ligases (CRLs), its role in regulating T cell signaling is poorly understood. Using the investigational NEDD8 activating enzyme (NAE) inhibitor, MLN4924, we found that neddylation negatively regulates T cell receptor (TCR) signaling, as its inhibition increases IL-2 production, T cell proliferation and Treg development in vitro. We also discovered that loss of CUL neddylation occurs upon TCR signaling, and CRLs negatively regulate IL-2 production. Additionally, we found that tyrosine kinase signaling leads to CUL deneddylation in multiple cell types. These studies indicate that CUL neddylation is a global regulatory mechanism for tyrosine kinase signaling.

Update on Staphylococcal Superantigen-Induced Signaling Pathways and Therapeutic Interventions

PubMed Central

Krakauer, Teresa

2013-01-01

Staphylococcal enterotoxin B (SEB) and related bacterial toxins cause diseases in humans and laboratory animals ranging from food poisoning, acute lung injury to toxic shock. These superantigens bind directly to the major histocompatibility complex class II molecules on antigen-presenting cells and specific Vβ regions of T-cell receptors (TCR), resulting in rapid hyper-activation of the host immune system. In addition to TCR and co-stimulatory signals, proinflammatory mediators activate signaling pathways culminating in cell-stress response, activation of NFκB and mammalian target of rapamycin (mTOR). This article presents a concise review of superantigen-activated signaling pathways and focuses on the therapeutic challenges against bacterial superantigens. PMID:24064719

Signal integration and cross-talk during thymocyte migration and emigration

PubMed Central

Love, Paul E.; Bhandoola, Avinash

2013-01-01

The thymus produces self-tolerant functionally competent T cells. This occurs by the import of multipotent hematopoietic progenitors that are signalled to adopt the T cell fate. Expression of T cell specific genes, including those encoding the T cell receptor (TCR), is followed by positive and negative selection and the eventual export of mature T cells. Significant progress has been made in elucidating the signals that direct progenitor cell trafficking to, within and out of the thymus. These advances are the subject of this Review, with a particular focus on the role of reciprocal cooperative and regulatory interactions between TCR and chemokine receptor-mediated signalling. PMID:21701522

SLAP, a regulator of immunoreceptor ubiquitination, signaling, and trafficking.

PubMed

Dragone, Leonard L; Shaw, Laura A; Myers, Margaret D; Weiss, Arthur

2009-11-01

Src-like adapter proteins (SLAP and SLAP-2) constitute a family of proteins that are expressed in a variety of cell types but are studied most extensively in lymphocytes. They have been shown to associate with proximal components of the T-cell receptor (TCR) and B-cell receptor (BCR) signaling complexes. An interaction of SLAP with c-Cbl leads to the ubiquitination and degradation of phosphorylated components of the TCR- and BCR-signaling complexes. The absence of this process in immature SLAP-deficient T and B cells leads to increased immunoreceptor levels due to decreased intracellular retention and degradation. We propose a model in which SLAP-dependent regulation of immunoreceptor levels allows for finer control of immunoreceptor signaling. Thus, SLAP functions to dampen immunoreceptor signaling, thereby influencing lymphocyte development and repertoire selection.

A Jurkat 76 based triple parameter reporter system to evaluate TCR functions and adoptive T cell strategies.

PubMed

Rosskopf, Sandra; Leitner, Judith; Paster, Wolfgang; Morton, Laura T; Hagedoorn, Renate S; Steinberger, Peter; Heemskerk, Mirjam H M

2018-04-03

Adoptive T cell therapy using TCR transgenic autologous T cells has shown great potential for the treatment of tumor patients. Thorough characterization of genetically reprogrammed T cells is necessary to optimize treatment success. Here, we describe the generation of triple parameter reporter T cells based on the Jurkat 76 T cell line for the evaluation of TCR and chimeric antigen receptor functions as well as adoptive T cell strategies. This Jurkat subline is devoid of endogenous TCR alpha and TCR beta chains, thereby circumventing the problem of TCR miss-pairing and unexpected specificities. The resultant reporter cells allow simultaneous determination of the activity of the transcription factors NF-κB, NFAT and AP-1 that play key roles in T cell activation. Human TCRs directed against tumor and virus antigens were introduced and reporter responses were determined using tumor cell lines endogenously expressing the antigens of interest or via addition of antigenic peptides. Finally, we demonstrate that coexpression of adhesion molecules like CD2 and CD226 as well as CD28 chimeric receptors represents an effective strategy to augment the response of TCR-transgenic reporters to cells presenting cognate antigens.

Social control of unreliable signals of strength in male but not female crayfish, Cherax destructor.

PubMed

Walter, Gregory M; van Uitregt, Vincent O; Wilson, Robbie S

2011-10-01

The maintenance of unreliable signals within animal populations remains a highly controversial subject in studies of animal communication. Crustaceans are an ideal group for studying unreliable signals of strength because their chela muscles are cryptically concealed beneath an exoskeleton, making it difficult for competitors to visually assess an opponent's strength. In this study, we examined the importance of social avenues for mediating the possible advantages gained by unreliable signals of strength in crustaceans. To do this, we investigated the factors that determine social dominance and the relative importance of signalling and fighting during aggressive encounters in male and female freshwater crayfish, Cherax destructor. Like other species of crayfish, we expected substantial variation in weapon force for a given weapon size, making the assessment of actual fighting ability of an opponent difficult from signalling alone. In addition, we expected fighting would be used to ensure that individuals that are weak for their signal (i.e. chela) size would not achieve higher than expected dominance. For both male and female C. destructor, we found large variation in the actual force of their chela for any given weapon size, indicating that it is difficult for competitors to accurately assess an opponent's force on signal size alone. For males, these unreliable signals of strength were controlled socially through increased levels of fighting and a decreased reliance on signalling, thus directly limiting the benefits accrued to individuals employing high-quality signals (large chelae) with only low resource holding potential. However, in contrast to our predictions, we found that females primarily relied on signalling to settle disputes, resulting in unreliable signals of strength being routinely used to establish dominance. The reliance by females on unreliable signals to determine dominance highlights our poor current understanding of the prevalence and

Integration of T Cell Receptor, Notch and Cytokine Signals Programs in Mouse γδ T Cell Effector Differentiation.

PubMed

Zarin, Payam; In, Tracy S H; Chen, Edward L Y; Singh, Jastaranpreet; Wong, Gladys W; Mohtashami, Mahmood; Wiest, David L; Anderson, Michele K; Zúñiga-Pflücker, Juan Carlos

2018-05-13

γδ T-cells perform a wide range of tissue and disease specific functions that are dependent on the effector cytokines produced by these cells. However, the aggregate signals required for the development of interferon-γ (IFNγ) and interleukin-17 (IL-17) producing γδ T-cells remain unknown. Here, we define the cues involved in the functional programming of γδ T-cells, by examining the roles of T-cell receptor (TCR), Notch, and cytokine-receptor signaling. KN6 γδTCR-transduced Rag2 -/- T-cell progenitors were cultured on stromal cells variably expressing TCR and Notch ligands, supplemented with different cytokines. We found that distinct combinations of these signals are required to program IFNγ versus IL-17 producing γδ T cell subsets, with Notch and weak TCR ligands optimally enabling development of γδ17 cells in the presence of IL-1β, IL-21 and IL-23. Notably, these cytokines were also shown to be required for the intrathymic development of γδ17 cells. Together, this work provides a framework of how signals downstream of TCR, Notch and cytokine receptors integrate to program the effector function of IFNγ and IL-17 producing γδ T-cell subsets. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Analysis of the CDR3 length repertoire and the diversity of TCR alpha chain in human peripheral blood T lymphocytes.

PubMed

Yao, Xin Sheng; Diao, Ying; Sun, Wan Bang; Luo, Jun Min; Qin, Ming; Tang, Xian Ying

2007-06-01

Analysis of complementarity determining region 3 (CDR3) length of T lymphocyte receptors (TCRs) by immunoscope spectratyping technique has been used successfully to investigate the diversity of TCR in autoimmune diseases and infection diseases. In this study, we investigated the patterns of CDR3 length distribution for all 32 TCR AV gene families in human peripheral blood lymphocytes of four normal volunteers by the immunoscope spectratyping technique. It was found that PCR products exhibited an obscure band on 1.5% agarose gel electrophoresis. Each TCR AV family exhibited more than 8 bands on 6% sequencing gel electrophoresis. The CDR3 spectratyping of all TCR AV families showed a standard Gaussian distribution with different CDR3 length, and the expression frequency of CDR3 was similar among the gene families. Most of CDR3 in TCR AV family recombine in frame. However, some of the CDR3 showed out-of frame gene rearrangement. Additionally, we found that in some of TCR AV families there were 18 amino acid discrepancies between the longest CDR3 and shortest CDR3. These results may be helpful to further study the recombination mechanism of human TCR genes, the TCR CDR3 gene repertoire, and the repertoire drift in health people and disease state.

In-depth Characterization of a TCR-specific Tracer for Sensitive Detection of Tumor-directed Transgenic T Cells by Immuno-PET.

PubMed

Yusufi, Nahid; Mall, Sabine; Bianchi, Henrique de Oliveira; Steiger, Katja; Reder, Sybille; Klar, Richard; Audehm, Stefan; Mustafa, Mona; Nekolla, Stephan; Peschel, Christian; Schwaiger, Markus; Krackhardt, Angela M; D'Alessandria, Calogero

2017-01-01

A number of different technologies have been developed to monitor in vivo the distribution of gene-modified T cells used in immunotherapy. Nevertheless, in-depth characterization of novel approaches with respect to sensitivity and clinical applicability are so far missing. We have previously described a novel method to track engineered human T cells in tumors using 89 Zr-Df-aTCRmu-F(ab') 2 targeting the murinized part of the TCR beta domain (TCRmu) of a transgenic TCR. Here, we performed an in-depth in vitro characterization of the tracer in terms of antigen affinity, immunoreactivity, influence on T-cell functionality and stability in vitro and in vivo . Of particular interest, we have developed diverse experimental settings to quantify TCR-transgenic T cells in vivo . Local application of 89 Zr-Df-aTCRmu-F(ab') 2 -labeled T cells in a spot-assay revealed signal detection down to approximately 1.8x10 4 cells. In a more clinically relevant model, NSG mice were intravenously injected with different numbers of transgenic T cells, followed by injection of the 89 Zr-Df-aTCRmu-F(ab') 2 tracer, PET/CT imaging and subsequent ex vivo T-cell quantification in the tumor. Using this setting, we defined a comparable detection limit of 1.0x10 4 T cells. PET signals correlated well to total numbers of transgenic T cells detected ex vivo independently of the engraftment rates observed in different individual experiments. Thus, these findings confirm the high sensitivity of our novel PET/CT T-cell tracking method and provide critical information about the quantity of transgenic T cells in the tumor environment suggesting our technology being highly suitable for further clinical translation.

High-throughput sequencing of TCR repertoires in multiple sclerosis reveals intrathecal enrichment of EBV-reactive CD8+ T cells.

PubMed

Lossius, Andreas; Johansen, Jorunn N; Vartdal, Frode; Robins, Harlan; Jūratė Šaltytė, Benth; Holmøy, Trygve; Olweus, Johanna

2014-11-01

Epstein-Barr virus (EBV) has long been suggested as a pathogen in multiple sclerosis (MS). Here, we used high-throughput sequencing to determine the diversity, compartmentalization, persistence, and EBV-reactivity of the T-cell receptor (TCR) repertoires in MS. TCR-β genes were sequenced in paired samples of cerebrospinal fluid (CSF) and blood from patients with MS and controls with other inflammatory neurological diseases. The TCR repertoires were highly diverse in both compartments and patient groups. Expanded T-cell clones, represented by TCR-β sequences >0.1%, were of different identity in CSF and blood of MS patients, and persisted for more than a year. Reference TCR-β libraries generated from peripheral blood T cells reactive against autologous EBV-transformed B cells were highly enriched for public EBV-specific sequences and were used to quantify EBV-reactive TCR-β sequences in CSF. TCR-β sequences of EBV-reactive CD8+ T cells, including several public EBV-specific sequences, were intrathecally enriched in MS patients only, whereas those of EBV-reactive CD4+ T cells were also enriched in CSF of controls. These data provide evidence for a clonally diverse, yet compartmentalized and persistent, intrathecal T-cell response in MS. The presented strategy links TCR sequence to intrathecal T-cell specificity, demonstrating enrichment of EBV-reactive CD8+ T cells in MS. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Increased Peptide Contacts Govern High Affinity Binding of a Modified TCR Whilst Maintaining a Native pMHC Docking Mode

PubMed Central

Cole, David K.; Sami, Malkit; Scott, Daniel R.; Rizkallah, Pierre J.; Borbulevych, Oleg Y.; Todorov, Penio T.; Moysey, Ruth K.; Jakobsen, Bent K.; Boulter, Jonathan M.; Baker, Brian M.; Yi Li

2013-01-01

Natural T cell receptors (TCRs) generally bind to their cognate pMHC molecules with weak affinity and fast kinetics, limiting their use as therapeutic agents. Using phage display, we have engineered a high affinity version of the A6 wild-type TCR (A6wt), specific for the human leukocyte antigen (HLA-A∗0201) complexed with human T cell lymphotropic virus type 111–19 peptide (A2-Tax). Mutations in just 4 residues in the CDR3β loop region of the A6wt TCR were selected that improved binding to A2-Tax by nearly 1000-fold. Biophysical measurements of this mutant TCR (A6c134) demonstrated that the enhanced binding was derived through favorable enthalpy and a slower off-rate. The structure of the free A6c134 TCR and the A6c134/A2-Tax complex revealed a native binding mode, similar to the A6wt/A2-Tax complex. However, concordant with the more favorable binding enthalpy, the A6c134 TCR made increased contacts with the Tax peptide compared with the A6wt/A2-Tax complex, demonstrating a peptide-focused mechanism for the enhanced affinity that directly involved the mutated residues in the A6c134 TCR CDR3β loop. This peptide-focused enhanced TCR binding may represent an important approach for developing antigen specific high affinity TCR reagents for use in T cell based therapies. PMID:23805144

Selection of Fecal Enterococci Exhibiting tcrB-Mediated Copper Resistance in Pigs Fed Diets Supplemented with Copper † ▿

PubMed Central

Amachawadi, R. G.; Shelton, N. W.; Shi, X.; Vinasco, J.; Dritz, S. S.; Tokach, M. D.; Nelssen, J. L.; Scott, H. M.; Nagaraja, T. G.

2011-01-01

Copper, as copper sulfate, is increasingly used as an alternative to in-feed antibiotics for growth promotion in weaned piglets. Acquired copper resistance, conferred by a plasmid-borne, transferable copper resistance (tcrB) gene, has been reported in Enterococcus faecium and E. faecalis. A longitudinal field study was undertaken to determine the relationship between copper supplementation and the prevalence of tcrB-positive enterococci in piglets. The study was done with weaned piglets, housed in 10 pens with 6 piglets per pen, fed diets supplemented with a normal (16.5 ppm; control) or an elevated (125 ppm) level of copper. Fecal samples were randomly collected from three piglets per pen on days 0, 14, 28, and 42 and plated on M-Enterococcus agar, and three enterococcal isolates were obtained from each sample. The overall prevalence of tcrB-positive enterococci was 21.1% (38/180) in piglets fed elevated copper and 2.8% (5/180) in the control. Among the 43 tcrB-positive isolates, 35 were E. faecium and 8 were E. faecalis. The mean MICs of copper for tcrB-negative and tcrB-positive enterococci were 6.2 and 22.2 mM, respectively. The restriction digestion of the genomic DNA of E. faecium or E. faecalis with S1 nuclease yielded a band of ∼194-kbp size to which both tcrB and the erm(B) gene probes hybridized. A conjugation assay demonstrated cotransfer of tcrB and erm(B) genes between E. faecium and E. faecalis strains. The higher prevalence of tcrB-positive enterococci in piglets fed elevated copper compared to that in piglets fed normal copper suggests that supplementation of copper in swine diets selected for resistance. PMID:21705534

Measuring Radiofrequency and Microwave Radiation from Varying Signal Strengths

NASA Technical Reports Server (NTRS)

Davis, Bette; Gaul, W. C.

2007-01-01

This viewgraph presentation discusses the process of measuring radiofrequency and microwave radiation from various signal strengths. The topics include: 1) Limits and Guidelines; 2) Typical Variable Standard (IEEE) Frequency Dependent; 3) FCC Standard 47 CFR 1.1310; 4) Compliance Follows Unity Rule; 5) Multiple Sources Contribute; 6) Types of RF Signals; 7) Interfering Radiations; 8) Different Frequencies Different Powers; 9) Power Summing - Peak Power; 10) Contribution from Various Single Sources; 11) Total Power from Multiple Sources; 12) Are You Out of Compliance?; and 13) In Compliance.

Definition of APC presentation of phosphoantigen (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate to Vgamma2Vdelta 2 TCR.

PubMed

Wei, Huiyong; Huang, Dan; Lai, Xiaomin; Chen, Meiling; Zhong, Weihua; Wang, Richard; Chen, Zheng W

2008-10-01

Although microbial (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP) can activate primate Vgamma2Vdelta2 T cells, molecular mechanisms by which HMBPP interacts with Vgamma2Vdelta2 T cells remain poorly characterized. Here, we developed soluble, tetrameric Vgamma2Vdelta2 TCR of rhesus macaques to define HMBPP/APC interaction with Vgamma2Vdelta2 TCR. While exogenous HMBPP was associated with APC membrane in an appreciable affinity, the membrane-associated HMBPP readily bound to the Vgamma2Vdelta2 TCR tetramer. The Vgamma2Vdelta2 TCR tetramer was shown to bind stably to HMBPP presented on membrane by various APC cell lines from humans and nonhuman primates but not those from mouse, rat, or pig. The Vgamma2Vdelta2 TCR tetramer also bound to the membrane-associated HMBPP on primary monocytes, B cells and T cells. Consistently, endogenous phosphoantigen produced in Mycobacterium-infected dendritic cells was transported and presented on membrane, and bound stably to the Vgamma2Vdelta2 TCR tetramer. The capability of APC to present HMBPP for recognition by Vgamma2Vdelta2 TCR was diminished after protease treatment of APC. Thus, our studies elucidated an affinity HMBPP-APC association conferring stable binding to the Vgamma2Vdelta2 TCR tetramer and the protease-sensitive nature of phosphoantigen presentation. The findings defined APC presentation of phosphoantigen HMBPP to Vgamma2Vdelta2 TCR.

Phosphorylation of a Tyrosine Residue on Zap70 by Lck and Its Subsequent Binding via an SH2 Domain May Be a Key Gatekeeper of T Cell Receptor Signaling In Vivo.

PubMed

Thill, Peter A; Weiss, Arthur; Chakraborty, Arup K

2016-09-15

The initiation of signaling in T lymphocytes in response to the binding of the T cell receptor (TCR) to cognate ligands is a key step in the emergence of adaptive immune responses. Conventional models posit that TCR signaling is initiated by the phosphorylation of receptor-associated immune receptor activation motifs (ITAMs). The cytoplasmic tyrosine kinase Zap70 binds to phosphorylated ITAMs, is subsequently activated, and then propagates downstream signaling. While evidence for such models is provided by experiments with cell lines, in vivo, Zap70 is bound to phosphorylated ITAMs in resting T cells. However, Zap70 is activated only upon TCR binding to cognate ligand. We report the results of computational studies of a new model for the initiation of TCR signaling that incorporates these in vivo observations. Importantly, the new model is shown to allow better and faster TCR discrimination between self-ligands and foreign ligands. The new model is consistent with many past experimental observations, and experiments that could further test the model are proposed. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Interpreting T-Cell Cross-reactivity through Structure: Implications for TCR-Based Cancer Immunotherapy.

PubMed

Antunes, Dinler A; Rigo, Maurício M; Freitas, Martiela V; Mendes, Marcus F A; Sinigaglia, Marialva; Lizée, Gregory; Kavraki, Lydia E; Selin, Liisa K; Cornberg, Markus; Vieira, Gustavo F

2017-01-01

Immunotherapy has become one of the most promising avenues for cancer treatment, making use of the patient's own immune system to eliminate cancer cells. Clinical trials with T-cell-based immunotherapies have shown dramatic tumor regressions, being effective in multiple cancer types and for many different patients. Unfortunately, this progress was tempered by reports of serious (even fatal) side effects. Such therapies rely on the use of cytotoxic T-cell lymphocytes, an essential part of the adaptive immune system. Cytotoxic T-cells are regularly involved in surveillance and are capable of both eliminating diseased cells and generating protective immunological memory. The specificity of a given T-cell is determined through the structural interaction between the T-cell receptor (TCR) and a peptide-loaded major histocompatibility complex (MHC); i.e., an intracellular peptide-ligand displayed at the cell surface by an MHC molecule. However, a given TCR can recognize different peptide-MHC (pMHC) complexes, which can sometimes trigger an unwanted response that is referred to as T-cell cross-reactivity. This has become a major safety issue in TCR-based immunotherapies, following reports of melanoma-specific T-cells causing cytotoxic damage to healthy tissues (e.g., heart and nervous system). T-cell cross-reactivity has been extensively studied in the context of viral immunology and tissue transplantation. Growing evidence suggests that it is largely driven by structural similarities of seemingly unrelated pMHC complexes. Here, we review recent reports about the existence of pMHC "hot-spots" for cross-reactivity and propose the existence of a TCR interaction profile (i.e., a refinement of a more general TCR footprint in which some amino acid residues are more important than others in triggering T-cell cross-reactivity). We also make use of available structural data and pMHC models to interpret previously reported cross-reactivity patterns among virus-derived peptides. Our

Interpreting T-Cell Cross-reactivity through Structure: Implications for TCR-Based Cancer Immunotherapy

PubMed Central

Antunes, Dinler A.; Rigo, Maurício M.; Freitas, Martiela V.; Mendes, Marcus F. A.; Sinigaglia, Marialva; Lizée, Gregory; Kavraki, Lydia E.; Selin, Liisa K.; Cornberg, Markus; Vieira, Gustavo F.

2017-01-01

Immunotherapy has become one of the most promising avenues for cancer treatment, making use of the patient’s own immune system to eliminate cancer cells. Clinical trials with T-cell-based immunotherapies have shown dramatic tumor regressions, being effective in multiple cancer types and for many different patients. Unfortunately, this progress was tempered by reports of serious (even fatal) side effects. Such therapies rely on the use of cytotoxic T-cell lymphocytes, an essential part of the adaptive immune system. Cytotoxic T-cells are regularly involved in surveillance and are capable of both eliminating diseased cells and generating protective immunological memory. The specificity of a given T-cell is determined through the structural interaction between the T-cell receptor (TCR) and a peptide-loaded major histocompatibility complex (MHC); i.e., an intracellular peptide–ligand displayed at the cell surface by an MHC molecule. However, a given TCR can recognize different peptide–MHC (pMHC) complexes, which can sometimes trigger an unwanted response that is referred to as T-cell cross-reactivity. This has become a major safety issue in TCR-based immunotherapies, following reports of melanoma-specific T-cells causing cytotoxic damage to healthy tissues (e.g., heart and nervous system). T-cell cross-reactivity has been extensively studied in the context of viral immunology and tissue transplantation. Growing evidence suggests that it is largely driven by structural similarities of seemingly unrelated pMHC complexes. Here, we review recent reports about the existence of pMHC “hot-spots” for cross-reactivity and propose the existence of a TCR interaction profile (i.e., a refinement of a more general TCR footprint in which some amino acid residues are more important than others in triggering T-cell cross-reactivity). We also make use of available structural data and pMHC models to interpret previously reported cross-reactivity patterns among virus

Analysis of acoustic emission cumulative signal strength of steel fibre reinforced concrete (SFRC) beams strengthened with carbon fibre reinforced polymer (CFRP)

NASA Astrophysics Data System (ADS)

Abdul Hakeem, Z.; Noorsuhada, M. N.; Azmi, I.; Noor Syafeekha, M. S.; Soffian Noor, M. S.

2017-12-01

In this study, steel fibre reinforced concrete (SFRC) beams strengthened with carbon fibre reinforced polymer (CFRP) were investigated using acoustic emission (AE) technique. Three beams with dimension of 150 mm width, 200 mm depth and 1500 mm length were fabricated. The results generated from AE parameters were analysed as well as signal strength and cumulative signal strength. Three relationships were produced namely load versus deflection, signal strength versus time and cumulative signal strength with respect to time. Each relationship indicates significant physical behaviour as the crack propagated in the beams. It is found that an addition of steel fibre in the concrete mix and strengthening of CFRP increase the ultimate load of the beam and the activity of signal strength. Moreover, the highest signal strength generated can be identified. From the study, the occurrence of crack in the beam can be predicted using AE signal strength.

CD28 co-stimulation restores T cell responsiveness in NOD mice by overcoming deficiencies in Rac-1/p38 mitogen-activated protein kinase signaling and IL-2 and IL-4 gene transcription.

PubMed

Zhang, J; Salojin, K V; Delovitch, T L

2001-03-01

Previously, we reported that T cell hyporesponsiveness induced by TCR ligation is causal to autoimmune diabetes in NOD mice. Neonatal CD28 co-stimulation reverses T cell hyporesponsiveness and protects NOD mice from diabetes by an IL-4-mediated mechanism, indicating that a deficiency in TCR signaling may be overcome by CD28/B7-2 co-stimulation in NOD T cells. To investigate which co-stimulation-induced signaling events mediate this protection, we analyzed the activity of Ras, Rac-1, mitogen-activated protein kinases (MAPK) and several transcription factors in TCR-activated NOD T cells in the presence or absence of CD28 co-stimulation. We show that CD28 co-stimulation restores normal TCR-induced activation of Rac-1 and p38 MAPK in NOD T cells. Deficiencies in TCR-induced nuclear expression of activating protein (AP)-1 binding proteins as well as activation of AP-1 and NF-AT in the IL-2 and IL-4 P1 promoters are also corrected by CD28 co-stimulation. Thus, CD28 co-stimulation reverses NOD T cell hyporesponsiveness by restoring TCR signaling leading to the activation of AP-1 and NF-AT during IL-2 and IL-4 gene transcription. Our findings provide additional evidence that CD28 co-stimulation amplifies signals delivered by the TCR and further explain the mechanism by which CD28 co-stimulation may protect against autoimmune diabetes.

Strength of signal: a fundamental mechanism for cell fate specification.

PubMed

Hayes, Sandra M; Love, Paul E

2006-02-01

How equipotent cells develop into complex tissues containing many diverse cell types is still a mystery. However, evidence is accumulating from different tissue systems in multiple organisms that many of the specific receptor families known to regulate cell fate decisions target conserved signaling pathways. A mechanism for preserving specificity in the cellular response that has emerged from these studies is one in which quantitative differences in receptor signaling regulate the cell fate decision. A signal strength model has recently gained support as a means to explain alphabeta/gammadelta lineage commitment. In this review, we compare the alphabeta/gammadelta fate decision with other cell fate decisions that occur outside of the lymphoid system to attain a better picture of the quantitative signaling mechanism for cell fate specification.

Efficient T-cell receptor signaling requires a high-affinity interaction between the Gads C-SH3 domain and the SLP-76 RxxK motif.

PubMed

Seet, Bruce T; Berry, Donna M; Maltzman, Jonathan S; Shabason, Jacob; Raina, Monica; Koretzky, Gary A; McGlade, C Jane; Pawson, Tony

2007-02-07

The relationship between the binding affinity and specificity of modular interaction domains is potentially important in determining biological signaling responses. In signaling from the T-cell receptor (TCR), the Gads C-terminal SH3 domain binds a core RxxK sequence motif in the SLP-76 scaffold. We show that residues surrounding this motif are largely optimized for binding the Gads C-SH3 domain resulting in a high-affinity interaction (K(D)=8-20 nM) that is essential for efficient TCR signaling in Jurkat T cells, since Gads-mediated signaling declines with decreasing affinity. Furthermore, the SLP-76 RxxK motif has evolved a very high specificity for the Gads C-SH3 domain. However, TCR signaling in Jurkat cells is tolerant of potential SLP-76 crossreactivity, provided that very high-affinity binding to the Gads C-SH3 domain is maintained. These data provide a quantitative argument that the affinity of the Gads C-SH3 domain for SLP-76 is physiologically important and suggest that the integrity of TCR signaling in vivo is sustained both by strong selection of SLP-76 for the Gads C-SH3 domain and by a capacity to buffer intrinsic crossreactivity.

Aurora A drives early signalling and vesicle dynamics during T-cell activation

PubMed Central

Blas-Rus, Noelia; Bustos-Morán, Eugenio; Pérez de Castro, Ignacio; de Cárcer, Guillermo; Borroto, Aldo; Camafeita, Emilio; Jorge, Inmaculada; Vázquez, Jesús; Alarcón, Balbino; Malumbres, Marcos; Martín-Cófreces, Noa B.; Sánchez-Madrid, Francisco

2016-01-01

Aurora A is a serine/threonine kinase that contributes to the progression of mitosis by inducing microtubule nucleation. Here we have identified an unexpected role for Aurora A kinase in antigen-driven T-cell activation. We find that Aurora A is phosphorylated at the immunological synapse (IS) during TCR-driven cell contact. Inhibition of Aurora A with pharmacological agents or genetic deletion in human or mouse T cells severely disrupts the dynamics of microtubules and CD3ζ-bearing vesicles at the IS. The absence of Aurora A activity also impairs the activation of early signalling molecules downstream of the TCR and the expression of IL-2, CD25 and CD69. Aurora A inhibition causes delocalized clustering of Lck at the IS and decreases phosphorylation levels of tyrosine kinase Lck, thus indicating Aurora A is required for maintaining Lck active. These findings implicate Aurora A in the propagation of the TCR activation signal. PMID:27091106

Attenuation of T cell receptor signaling by serine phosphorylation-mediated lysine 30 ubiquitination of SLP-76 protein.

PubMed

Wang, Xiaohong; Li, Ju-Pi; Chiu, Li-Li; Lan, Joung-Liang; Chen, Der-Yuan; Boomer, Jonathan; Tan, Tse-Hua

2012-10-05

SLP-76 (SH2 domain-containing leukocyte protein of 76 kDa) is an adaptor protein that is essential for T cell development and T cell receptor (TCR) signaling activation. Previous studies have identified an important negative feedback regulation of SLP-76 by HPK1 (hematopoietic progenitor kinase 1; MAP4K1)-induced Ser-376 phosphorylation. Ser-376 phosphorylation of SLP-76 mediates 14-3-3 binding, resulting in the attenuation of SLP-76 activation and downstream signaling; however, the underlying mechanism of this action remains unknown. Here, we report that phosphorylated SLP-76 is ubiquitinated and targeted for proteasomal degradation during TCR signaling. SLP-76 ubiquitination is mediated by Ser-376 phosphorylation. Furthermore, Lys-30 is identified as a ubiquitination site of SLP-76. Loss of Lys-30 ubiquitination of SLP-76 results in enhanced anti-CD3 antibody-induced ERK and JNK activation. These results reveal a novel regulation mechanism of SLP-76 by ubiquitination and proteasomal degradation of activated SLP-76, which is mediated by Ser-376 phosphorylation, leading to down-regulation of TCR signaling.

Attenuation of T Cell Receptor Signaling by Serine Phosphorylation-mediated Lysine 30 Ubiquitination of SLP-76 Protein*

PubMed Central

Wang, Xiaohong; Li, Ju-Pi; Chiu, Li-Li; Lan, Joung-Liang; Chen, Der-Yuan; Boomer, Jonathan; Tan, Tse-Hua

2012-01-01

SLP-76 (SH2 domain-containing leukocyte protein of 76 kDa) is an adaptor protein that is essential for T cell development and T cell receptor (TCR) signaling activation. Previous studies have identified an important negative feedback regulation of SLP-76 by HPK1 (hematopoietic progenitor kinase 1; MAP4K1)-induced Ser-376 phosphorylation. Ser-376 phosphorylation of SLP-76 mediates 14-3-3 binding, resulting in the attenuation of SLP-76 activation and downstream signaling; however, the underlying mechanism of this action remains unknown. Here, we report that phosphorylated SLP-76 is ubiquitinated and targeted for proteasomal degradation during TCR signaling. SLP-76 ubiquitination is mediated by Ser-376 phosphorylation. Furthermore, Lys-30 is identified as a ubiquitination site of SLP-76. Loss of Lys-30 ubiquitination of SLP-76 results in enhanced anti-CD3 antibody-induced ERK and JNK activation. These results reveal a novel regulation mechanism of SLP-76 by ubiquitination and proteasomal degradation of activated SLP-76, which is mediated by Ser-376 phosphorylation, leading to down-regulation of TCR signaling. PMID:22902619

Nanostructured vanadium oxide thin film with high TCR at room temperature for microbolometer

NASA Astrophysics Data System (ADS)

Wang, Bin; Lai, Jianjun; Li, Hui; Hu, Haoming; Chen, Sihai

2013-03-01

In order to obtain high quality of thermal sensitive material, VOx thin film of high temperature coefficient of resistance (TCR) of 6.5%/K at room temperature has been deposited by reactive ion beam sputtering and post annealing method. AFM and XRD measurements indicate that the VOx thin film with nanostructured crystalline is composed of VO2 and V2O3. The nanostructured VOx microbolometer has been designed and fabricated. The measurement of the film system with TiN absorbing layer indicates that it has about 92% infrared absorption in the range of 8-14 μm. The performance of this bolometer, comparing with that of bolometer with common VOx, has a better result. At 20 Hz frequency and 10 μA bias current, the bolometer with high TCR has reached detectivity of 1.0 × 109 cm Hz1/2/W. It also indicates that this nanostructured VOx thin film has not only a higher TCR but also a lower noise than common VOx thin film without annealing.

Different TCR-induced T lymphocyte responses are potentiated by stiffness with variable sensitivity

PubMed Central

Saitakis, Michael; Dogniaux, Stéphanie; Goudot, Christel; Bufi, Nathalie; Asnacios, Sophie; Maurin, Mathieu; Randriamampita, Clotilde; Asnacios, Atef; Hivroz, Claire

2017-01-01

T cells are mechanosensitive but the effect of stiffness on their functions is still debated. We characterize herein how human primary CD4+ T cell functions are affected by stiffness within the physiological Young’s modulus range of 0.5 kPa to 100 kPa. Stiffness modulates T lymphocyte migration and morphological changes induced by TCR/CD3 triggering. Stiffness also increases TCR-induced immune system, metabolism and cell-cycle-related genes. Yet, upon TCR/CD3 stimulation, while cytokine production increases within a wide range of stiffness, from hundreds of Pa to hundreds of kPa, T cell metabolic properties and cell cycle progression are only increased by the highest stiffness tested (100 kPa). Finally, mechanical properties of adherent antigen-presenting cells modulate cytokine production by T cells. Together, these results reveal that T cells discriminate between the wide range of stiffness values found in the body and adapt their responses accordingly. DOI: http://dx.doi.org/10.7554/eLife.23190.001 PMID:28594327

Differences in TCR-Vβ Repertoire and Effector Phenotype between Tumor Infiltrating Lymphocytes and Peripheral Blood Lymphocytes Increase with Age

PubMed Central

Wang, Teng; Shen, Han; Wu, Fenglin; Zhang, Wenfeng; Tao, Changli; Yuan, Yin; Bo, Huaben; Wang, Hui; Huang, Shulin

2014-01-01

Tumor infiltrating lymphocytes (TIL) reflect the host's anti-tumor immune response, and can be a valuable predictor of prognosis. However, many properties of TIL are not fully understood. In the present study, TCR-Vβ repertoires of cancer patients were primarily analyzed by flow cytometry. Abnormally expressed TCR-Vβ subfamilies were generally found in both TIL and peripheral blood lymphocytes (PBL) of each patient. Of note, increased patient age was associated with increasingly biased TCR-Vβ repertoire in TIL but not in PBL, and the dispersion degree of the differences of TCR-Vβ subfamilies between TIL and PBL correlated positively with age (P = 0.007). Utilizing immunoscope analysis, we identified the age-related reduction in TCR-Vβ diversity, but polyclonal pattern was predominant in significantly expanded TCR-Vβ subfamilies. In addition, we found that older patients possessed a decreased ratio of CD8+CD62L+ non-effector cells in TIL compared to PBL, implying age-related increase of CD8+CD62L− effector cells in TIL. The colocalization analysis of CD8 and CD3, however, suggested the suppressed activity of these effector cells in tumor microenvironment. These findings further elucidate the properties of TIL, showing an increasing difference between TIL and PBL with age, which may provide insight for the development of effective immunotherapies for cancer patients of different ages. PMID:25019226

Differential regulation of peripheral CD4+ T cell tolerance induced by deletion and TCR revision.

PubMed

Ali, Mohamed; Weinreich, Michael; Balcaitis, Stephanie; Cooper, Cristine J; Fink, Pamela J

2003-12-01

In Vbeta5 transgenic mice, mature Vbeta5(+)CD4(+) T cells are tolerized upon recognition of a self Ag, encoded by a defective endogenous retrovirus, whose expression is confined to the lymphoid periphery. Cells are driven by the tolerogen to enter one of two tolerance pathways, deletion or TCR revision. CD4(+) T cells entering the former pathway are rendered anergic and then eliminated. In contrast, TCR revision drives gene rearrangement at the endogenous TCR beta locus and results in the appearance of Vbeta5(-), endogenous Vbeta(+), CD4(+) T cells that are both self-tolerant and functional. An analysis of the molecules that influence each of these pathways was conducted to understand better the nature of the interactions that control tolerance induction in the lymphoid periphery. These studies reveal that deletion is efficient in reconstituted radiation chimeras and is B cell, CD28, inducible costimulatory molecule, Fas, CD4, and CD8 independent. In contrast, TCR revision is radiosensitive, B cell, CD28, and inducible costimulatory molecule dependent, Fas and CD4 influenced, and CD8 independent. Our data demonstrate the differential regulation of these two divergent tolerance pathways, despite the fact that they are both driven by the same tolerogen and restricted to mature CD4(+) T cells.

Src-like adaptor protein down-regulates T cell receptor (TCR)-CD3 expression by targeting TCRzeta for degradation.

PubMed

Myers, Margaret D; Dragone, Leonard L; Weiss, Arthur

2005-07-18

Src-like adaptor protein (SLAP) down-regulates expression of the T cell receptor (TCR)-CD3 complex during a specific stage of thymocyte development when the TCR repertoire is selected. Consequently, SLAP-/- thymocytes display alterations in thymocyte development. Here, we have studied the mechanism of SLAP function. We demonstrate that SLAP-deficient thymocytes have increased TCRzeta chain expression as a result of a defect in TCRzeta degradation. Failure to degrade TCRzeta leads to an increased pool of fully assembled TCR-CD3 complexes that are capable of recycling back to the cell surface. We also provide evidence that SLAP functions in a pathway that requires the phosphorylated TCRzeta chain and the Src family kinase Lck, but not ZAP-70 (zeta-associated protein of 70 kD). These studies reveal a unique mechanism by which SLAP contributes to the regulation of TCR expression during a distinct stage of thymocyte development.

T-cell Receptor Signaling Activates an ITK/NF-κB/GATA-3 axis in T-cell Lymphomas Facilitating Resistance to Chemotherapy.

PubMed

Wang, Tianjiao; Lu, Ye; Polk, Avery; Chowdhury, Pinki; Zamalloa, Carlos Murga; Fujiwara, Hiroshi; Suemori, Koichiro; Beyersdorf, Niklas; Hristov, Alexandra C; Lim, Megan S; Bailey, Nathanael G; Wilcox, Ryan A

2017-05-15

Purpose: T-cell lymphomas are a molecularly heterogeneous group of non-Hodgkin lymphomas (NHL) that account for a disproportionate number of NHL disease-related deaths due to their inherent and acquired resistance to standard multiagent chemotherapy regimens. Despite their molecular heterogeneity and frequent loss of various T cell-specific receptors, the T-cell antigen receptor is retained in the majority of these lymphomas. As T-cell receptor (TCR) engagement activates a number of signaling pathways and transcription factors that regulate T-cell growth and survival, we examined the TCR's role in mediating resistance to chemotherapy. Experimental Design: Genetic and pharmacologic strategies were utilized to determine the contribution of tyrosine kinases and transcription factors activated in conventional T cells following TCR engagement in acquired chemotherapy resistance in primary T-cell lymphoma cells and patient-derived cell lines. Results: Here, we report that TCR signaling activates a signaling axis that includes ITK, NF-κB, and GATA-3 and promotes chemotherapy resistance. Conclusions: These observations have significant therapeutic implications, as pharmacologic inhibition of ITK prevented the activation of this signaling axis and overcame chemotherapy resistance. Clin Cancer Res; 23(10); 2506-15. ©2016 AACR . ©2016 American Association for Cancer Research.

Effect of injection current and temperature on signal strength in a laser diode optical feedback interferometer.

PubMed

Al Roumy, Jalal; Perchoux, Julien; Lim, Yah Leng; Taimre, Thomas; Rakić, Aleksandar D; Bosch, Thierry

2015-01-10

We present a simple analytical model that describes the injection current and temperature dependence of optical feedback interferometry signal strength for a single-mode laser diode. The model is derived from the Lang and Kobayashi rate equations, and is developed both for signals acquired from the monitoring photodiode (proportional to the variations in optical power) and for those obtained by amplification of the corresponding variations in laser voltage. The model shows that both the photodiode and the voltage signal strengths are dependent on the laser slope efficiency, which itself is a function of the injection current and the temperature. Moreover, the model predicts that the photodiode and voltage signal strengths depend differently on injection current and temperature. This important model prediction was proven experimentally for a near-infrared distributed feedback laser by measuring both types of signals over a wide range of injection currents and temperatures. Therefore, this simple model provides important insight into the radically different biasing strategies required to achieve optimal sensor sensitivity for both interferometric signal acquisition schemes.

The effect of inhibitory signals on the priming of drug-hapten-specific T-cells that express distinct Vβ receptors

PubMed Central

Gibson, Andrew; Faulkner, Lee; Lichtenfels, Maike; Ogese, Monday; Al-Attar, Zaid; Alfirevic, Ana; Esser, Philipp R.; Martin, Stefan F.; Pirmohamed, Munir; Park, B. Kevin; Naisbitt, Dean J.

2017-01-01

Drug hypersensitivity involves the activation of T-cells in an HLA allele-restricted manner. Since the majority of individuals who carry HLA risk alleles do not develop hypersensitivity, other parameters must control development of the drug-specific T-cell response. Thus, we have utilized a T-cell priming assay and nitroso sulfamethoxazole (SMX-NO) as a model antigen to investigate (1) the activation of specific T-cell receptor (TCR)Vβ subtypes, (2) the impact of PD-1, CTLA4 and TIM-3 co-inhibitory signalling on activation of naïve and memory T-cells and (3) the ability of Tregs to prevent responses. An expansion of the TCR repertoire was observed for nine different Vβ subtypes, while spectratyping revealed that SMX-NO-specific T-cell responses are controlled by public TCRs present in all individuals alongside private TCR repertoires specific to each individual. We proceeded to evaluate the extent to which the activation of these TCR Vβ-restricted antigen-specific T-cell responses is governed by regulatory signals. Blockade of PDL-1/CTLA4 signalling dampened activation of SMX-NO-specific naïve and memory T-cells, while blockade of TIM-3 produced no effect. PD-1, CTLA4, and TIM-3 displayed discrete expression profiles during drug-induced T-cell activation and expression of each receptor was enhanced on dividing T-cells. As these receptors are also expressed on Tregs, Treg-mediated suppression of SMX-NO-induced T-cell activation was investigated. Tregs significantly dampened the priming of T-cells. In conclusion, our findings demonstrate that distinct TCR Vβ subtypes, dysregulation of co-inhibitory signalling pathways and dysfunctional Tregs may influence predisposition to hypersensitivity. PMID:28687658

Direct molecular mimicry enables off-target cardiovascular toxicity by an enhanced affinity TCR designed for cancer immunotherapy.

PubMed

Raman, Marine C C; Rizkallah, Pierre J; Simmons, Ruth; Donnellan, Zoe; Dukes, Joseph; Bossi, Giovanna; Le Provost, Gabrielle S; Todorov, Penio; Baston, Emma; Hickman, Emma; Mahon, Tara; Hassan, Namir; Vuidepot, Annelise; Sami, Malkit; Cole, David K; Jakobsen, Bent K

2016-01-13

Natural T-cell responses generally lack the potency to eradicate cancer. Enhanced affinity T-cell receptors (TCRs) provide an ideal approach to target cancer cells, with emerging clinical data showing significant promise. Nevertheless, the risk of off target reactivity remains a key concern, as exemplified in a recent clinical report describing fatal cardiac toxicity, following administration of MAGE-A3 specific TCR-engineered T-cells, mediated through cross-reactivity with an unrelated epitope from the Titin protein presented on cardiac tissue. Here, we investigated the structural mechanism enabling TCR cross-recognition of MAGE-A3 and Titin, and applied the resulting data to rationally design mutants with improved antigen discrimination, providing a proof-of-concept strategy for altering the fine specificity of a TCR towards an intended target antigen. This study represents the first example of direct molecular mimicry leading to clinically relevant fatal toxicity, mediated by a modified enhanced affinity TCR designed for cancer immunotherapy. Furthermore, these data demonstrate that self-antigens that are expressed at high levels on healthy tissue should be treated with extreme caution when designing immuno-therapeutics.

Propensity of a single-walled carbon nanotube-peptide to mimic a KK10 peptide in an HLA-TCR complex

NASA Astrophysics Data System (ADS)

Feng, Mei; Bell, David R.; Zhou, Ruhong

2017-12-01

The application of nanotechnology to improve disease diagnosis, treatment, monitoring, and prevention is the goal of nanomedicine. We report here a theoretical study of a functionalized single-walled carbon nanotube (CNT) mimic binding to a human leukocyte antigen-T cell receptor (HLA-TCR) immune complex as a first attempt of a potential nanomedicine for human immunodeficiency virus (HIV) vaccine development. The carbon nanotube was coated with three arginine residues to imitate the HIV type 1 immunodominant viral peptide KK10 (gag 263-272: KRWIILGLNK), named CNT-peptide hereafter. Through molecular dynamics simulations, we explore the CNT-peptide and KK10 binding to an important HLA-TCR complex. Our results suggest that the CNT-peptide and KK10 bind comparably to the HLA-TCR complex, but the CNT-peptide forms stronger interactions with the TCR. Desorption simulations highlight the innate flexibility of KK10 over the CNT-peptide, resulting in a slightly higher desorption energy required for KK10 over the CNT-peptide. Our findings indicate that the designed CNT-peptide mimic has favorable propensity to activate TCR pathways and should be further explored to understand therapeutic potential.

Intermittent IL-7 Signaling Essential for T cell Homeostasis | Center for Cancer Research

Cancer.gov

In order for the immune system to mount an appropriate response to foreign antigens throughout a person’s life, the body must maintain a sufficient population of circulating mature, naïve T cells, a process known as T cell homeostasis. Previous studies revealed that this process depends upon signaling from the cytokine interleukin-7 (IL-7) as well as from the T cell antigen receptor (TCR). Intriguingly, signals from each pathway affect the other and lead to their alternating activation: IL-7 binding to its receptor leads to increasing expression of the TCR co-receptor CD8; sufficient CD8 expression allows TCRs to signal when bound to self-ligands, blocking IL-7 signaling; suppressed IL-7 signals lead to down-regulation of CD8 and ligand disengagement, which allows T cells to again respond to IL-7. Alfred Singer, M.D., and his colleagues in CCR’s Experimental Immunology Branch set out to understand how this intricate pathway promotes T cell survival.

Direct molecular mimicry enables off-target cardiovascular toxicity by an enhanced affinity TCR designed for cancer immunotherapy

PubMed Central

Raman, Marine C C; Rizkallah, Pierre J; Simmons, Ruth; Donnellan, Zoe; Dukes, Joseph; Bossi, Giovanna; Le Provost, Gabrielle S; Todorov, Penio; Baston, Emma; Hickman, Emma; Mahon, Tara; Hassan, Namir; Vuidepot, Annelise; Sami, Malkit; Cole, David K; Jakobsen, Bent K.

2016-01-01

Natural T-cell responses generally lack the potency to eradicate cancer. Enhanced affinity T-cell receptors (TCRs) provide an ideal approach to target cancer cells, with emerging clinical data showing significant promise. Nevertheless, the risk of off target reactivity remains a key concern, as exemplified in a recent clinical report describing fatal cardiac toxicity, following administration of MAGE-A3 specific TCR-engineered T-cells, mediated through cross-reactivity with an unrelated epitope from the Titin protein presented on cardiac tissue. Here, we investigated the structural mechanism enabling TCR cross-recognition of MAGE-A3 and Titin, and applied the resulting data to rationally design mutants with improved antigen discrimination, providing a proof-of-concept strategy for altering the fine specificity of a TCR towards an intended target antigen. This study represents the first example of direct molecular mimicry leading to clinically relevant fatal toxicity, mediated by a modified enhanced affinity TCR designed for cancer immunotherapy. Furthermore, these data demonstrate that self-antigens that are expressed at high levels on healthy tissue should be treated with extreme caution when designing immuno-therapeutics. PMID:26758806

Evolutionarily conserved TCR binding sites, identification of T cells in primary lymphoid tissues, and surprising trans-rearrangements in nurse shark.

PubMed

Criscitiello, Michael F; Ohta, Yuko; Saltis, Mark; McKinney, E Churchill; Flajnik, Martin F

2010-06-15

Cartilaginous fish are the oldest animals that generate RAG-based Ag receptor diversity. We have analyzed the genes and expressed transcripts of the four TCR chains for the first time in a cartilaginous fish, the nurse shark (Ginglymostoma cirratum). Northern blotting found TCR mRNA expression predominantly in lymphoid and mucosal tissues. Southern blotting suggested translocon-type loci encoding all four chains. Based on diversity of V and J segments, the expressed combinatorial diversity for gamma is similar to that of human, alpha and beta may be slightly lower, and delta diversity is the highest of any organism studied to date. Nurse shark TCRdelta have long CDR3 loops compared with the other three chains, creating binding site topologies comparable to those of mammalian TCR in basic paratope structure; additionally, nurse shark TCRdelta CDR3 are more similar to IgH CDR3 in length and heterogeneity than to other TCR chains. Most interestingly, several cDNAs were isolated that contained IgM or IgW V segments rearranged to other gene segments of TCRdelta and alpha. Finally, in situ hybridization experiments demonstrate a conservation of both alpha/beta and gamma/delta T cell localization in the thymus across 450 million years of vertebrate evolution, with gamma/delta TCR expression especially high in the subcapsular region. Collectively, these data make the first cellular identification of TCR-expressing lymphocytes in a cartilaginous fish.

KIAA1530 Protein Is Recruited by Cockayne Syndrome Complementation Group Protein A (CSA) to Participate in Transcription-coupled Repair (TCR)

PubMed Central

Fei, Jia; Chen, Junjie

2012-01-01

Transcription-coupled repair (TCR) is the major pathway involved in the removal of UV-induced photolesions from the transcribed strand of active genes. Two Cockayne syndrome (CS) complementation group proteins, CSA and CSB, are important for TCR repair. The molecular mechanisms by which CS proteins regulate TCR remain elusive. Here, we report the characterization of KIAA1530, an evolutionarily conserved protein that participates in this pathway through its interaction with CSA and the TFIIH complex. We found that UV irradiation led to the recruitment of KIAA1530 onto chromatin in a CSA-dependent manner. Cells lacking KIAA1530 were highly sensitive to UV irradiation and displayed deficiency in TCR. In addition, KIAA1530 depletion abrogated stability of the CSB protein following UV irradiation. More excitingly, we found that a unique CSA mutant (W361C), which was previously identified in a patient with UVsS syndrome, showed defective KIAA1530 binding and resulted in a failure of recruiting KIAA1530 and stabilizing CSB after UV treatment. Together, our data not only reveal that KIAA1530 is an important player in TCR but also lead to a better understanding of the molecular mechanism underlying UVsS syndrome. PMID:22902626

Evenness indices measure the signal strength of biweight site chronologies

Treesearch

Kurt H. Riitters

1990-01-01

The signal strength of a biweight site chronology is properly viewed as an outcome of analysis rather than as a property of the forest-climate system. It can be estimated by the evenness of the empirical weights that are assigned to individual trees. The approach is demonstrated for a 45-year biweight chronology obtained from 40 jack pine (Pinus banksiana Lamb.) trees...

Occurrence of the Transferable Copper Resistance Gene tcrB among Fecal Enterococci of U.S. Feedlot Cattle Fed Copper-Supplemented Diets

PubMed Central

Amachawadi, R. G.; Alvarado, C. A.; Mainini, T. R.; Vinasco, J.; Drouillard, J. S.; Nagaraja, T. G.

2013-01-01

Copper, an essential micronutrient, is supplemented in the diet at elevated levels to reduce morbidity and mortality and to promote growth in feedlot cattle. Gut bacteria exposed to copper can acquire resistance, which among enterococci is conferred by a transferable copper resistance gene (tcrB) borne on a plasmid. The present study was undertaken to investigate whether the feeding of copper at levels sufficient to promote growth increases the prevalence of the tcrB gene among the fecal enterococci of feedlot cattle. The study was performed with 261 crossbred yearling heifers housed in 24 pens, with pens assigned randomly to a 2×2 factorial arrangement of treatments consisting of dietary copper and a commercial linseed meal-based energy protein supplement. A total of 22 isolates, each identified as Enterococcus faecium, were positive for tcrB with an overall prevalence of 3.8% (22/576). The prevalence was higher among the cattle fed diets supplemented with copper (6.9%) compared to normal copper levels (0.7%). The tcrB-positive isolates always contained both erm(B) and tet(M) genes. Median copper MICs for tcrB-positive and tcrB-negative enterococci were 22 and 4 mM, respectively. The transferability of the tcrB gene was demonstrated via a filter-mating assay. Multilocus variable number tandem repeat analysis revealed a genetically diverse population of enterococci. The finding of a strong association between the copper resistance gene and other antibiotic (tetracycline and tylosin) resistance determinants is significant because enterococci remain potential pathogens and have the propensity to transfer resistance genes to other bacteria in the gut. PMID:23666328

The WAVE2 complex regulates T cell receptor signaling to integrins via Abl- and CrkL-C3G-mediated activation of Rap1.

PubMed

Nolz, Jeffrey C; Nacusi, Lucas P; Segovis, Colin M; Medeiros, Ricardo B; Mitchell, Jason S; Shimizu, Yoji; Billadeau, Daniel D

2008-09-22

WAVE2 regulates T cell receptor (TCR)-stimulated actin cytoskeletal dynamics leading to both integrin clustering and affinity maturation. Although WAVE2 mediates integrin affinity maturation by recruiting vinculin and talin to the immunological synapse in an Arp2/3-dependent manner, the mechanism by which it regulates integrin clustering is unclear. We show that the Abl tyrosine kinase associates with the WAVE2 complex and TCR ligation induces WAVE2-dependent membrane recruitment of Abl. Furthermore, we show that WAVE2 regulates TCR-mediated activation of the integrin regulatory guanosine triphosphatase Rap1 via the recruitment and activation of the CrkL-C3G exchange complex. Moreover, we demonstrate that although Abl does not regulate the recruitment of CrkL-C3G into the membrane, it does affect the tyrosine phosphorylation of C3G, which is required for its guanine nucleotide exchange factor activity toward Rap1. This signaling node regulates not only TCR-stimulated integrin clustering but also affinity maturation. These findings identify a previously unknown mechanism by which the WAVE2 complex regulates TCR signaling to Rap1 and integrin activation.

The WAVE2 complex regulates T cell receptor signaling to integrins via Abl- and CrkL–C3G-mediated activation of Rap1

PubMed Central

Nolz, Jeffrey C.; Nacusi, Lucas P.; Segovis, Colin M.; Medeiros, Ricardo B.; Mitchell, Jason S.; Shimizu, Yoji; Billadeau, Daniel D.

2008-01-01

WAVE2 regulates T cell receptor (TCR)–stimulated actin cytoskeletal dynamics leading to both integrin clustering and affinity maturation. Although WAVE2 mediates integrin affinity maturation by recruiting vinculin and talin to the immunological synapse in an Arp2/3-dependent manner, the mechanism by which it regulates integrin clustering is unclear. We show that the Abl tyrosine kinase associates with the WAVE2 complex and TCR ligation induces WAVE2-dependent membrane recruitment of Abl. Furthermore, we show that WAVE2 regulates TCR-mediated activation of the integrin regulatory guanosine triphosphatase Rap1 via the recruitment and activation of the CrkL–C3G exchange complex. Moreover, we demonstrate that although Abl does not regulate the recruitment of CrkL–C3G into the membrane, it does affect the tyrosine phosphorylation of C3G, which is required for its guanine nucleotide exchange factor activity toward Rap1. This signaling node regulates not only TCR-stimulated integrin clustering but also affinity maturation. These findings identify a previously unknown mechanism by which the WAVE2 complex regulates TCR signaling to Rap1 and integrin activation. PMID:18809728

How many TCR clonotypes does a body maintain?

PubMed Central

Lythe, Grant; Callard, Robin E.; Hoare, Rollo L.; Molina-París, Carmen

2016-01-01

We consider the lifetime of a T cell clonotype, the set of T cells with the same T cell receptor, from its thymic origin to its extinction in a multiclonal repertoire. Using published estimates of total cell numbers and thymic production rates, we calculate the mean number of cells per TCR clonotype, and the total number of clonotypes, in mice and humans. When there is little peripheral division, as in a mouse, the number of cells per clonotype is small and governed by the number of cells with identical TCR that exit the thymus. In humans, peripheral division is important and a clonotype may survive for decades, during which it expands to comprise many cells. We therefore devise and analyse a computational model of homeostasis of a multiclonal population. Each T cell in the model competes for self pMHC stimuli, cells of any one clonotype only recognising a small fraction of the many subsets of stimuli. A constant mean total number of cells is maintained by a balance between cell division and death, and a stable number of clonotypes by a balance between thymic production of new clonotypes and extinction of existing ones. The number of distinct clonotypes in a human body may be smaller than the total number of naive T cells by only one order of magnitude. PMID:26546971

T cell receptor (TCR) structure of autologous melanoma-reactive cytotoxic T lymphocyte (CTL) clones: tumor-infiltrating lymphocytes overexpress in vivo the TCR beta chain sequence used by an HLA-A2- restricted and melanocyte-lineage-specific CTL clone

PubMed Central

1993-01-01

HLA-A2+ melanomas express common melanoma-associated antigens (Ags) recognized in vitro by autologous cytotoxic T lymphocytes (CTL). However, it is not known whether tumor Ags can drive in vivo a selective accumulation/expansion of Ag-specific, tumor-infiltrating T lymphocytes (TIL). Therefore, to evaluate this possibility, 39 CTL clones isolated from several independent mixed lymphocyte tumor cultures (MLTC) of TIL and peripheral blood lymphocytes (PBL) of an HLA- A2+ melanoma patient and selected for T cell receptor (TCR)-dependent, HLA-restricted tumor lysis, were used for analysis of TCR alpha and beta chain structure by the cDNA polymerase chain reaction (PCR) technique with variable gene-specific primers followed by sequencing. Despite absence of oligoclonality in fresh TIL and PBL, as well as in T cells of day 28 MLTC (day of cloning), sequence analysis of TCR alpha and beta chains of TIL clones revealed a dominance of a major category of melanoma-specific, HLA-A2-restricted T cells expressing a V alpha 8.2/J alpha AP511/C alpha and V beta 2.1/D beta 1/J beta 1.1/C beta 1 TCR. The same TCR was also found in 2 out of 14 PBL clones. The other PBL clones employed a V alpha 2.1 gene segment associated with either V beta 13.2, 14, or w22. Clones A81 (V alpha 2.1/J alpha IGRJ alpha 04/C alpha and V beta 14/D beta 1/J beta 1.2/C beta 1) and A21 (V alpha 8.2/J alpha AP511/C alpha and V beta 2.1/D beta 1/J beta 1.1/C beta 1), representative of the two most frequent TCR of PBL and TIL, respectively, expressed different lytic patterns, but both were HLA-A2 restricted and lysed only HLA-A2+ melanomas and normal melanocytes, thus indicating recognition of two distinct HLA-A2-associated and tissue-related Ags. Finally, by the inverse PCR technique, the specific TCR beta chain (V beta 2.1/D beta 1/J beta 1.1/C beta 1) expressed by the dominant TIL clone was found to represent 19 and 18.4% of all V beta 2 sequences expressed in the fresh tumor sample and in the purified TIL

Bacterial superantigens bypass Lck-dependent T cell receptor signaling by activating a Galpha11-dependent, PLC-beta-mediated pathway.

PubMed

Bueno, Clara; Lemke, Caitlin D; Criado, Gabriel; Baroja, Miren L; Ferguson, Stephen S G; Rahman, A K M Nur-Ur; Tsoukas, Constantine D; McCormick, John K; Madrenas, Joaquin

2006-07-01

The paradigm to explain antigen-dependent T cell receptor (TCR) signaling is based on the activation of the CD4 or CD8 coreceptor-associated kinase Lck. It is widely assumed that this paradigm is also applicable to signaling by bacterial superantigens. However, these bacterial toxins can activate human T cells lacking Lck, suggesting the existence of an additional pathway of TCR signaling. Here we showed that this alternative pathway operates in the absence of Lck-dependent tyrosine-phosphorylation events and was initiated by the TCR-dependent activation of raft-enriched heterotrimeric Galpha11 proteins. This event, in turn, activated a phospholipase C-beta and protein kinase C-mediated cascade that turned on the mitogen-activated protein kinases ERK-1 and ERK-2, triggered Ca(2+) influx, and translocated the transcription factors NF-AT and NF-kappaB to the nucleus, ultimately inducing the production of interleukin-2 in Lck-deficient T cells. The triggering of this alternative pathway by superantigens suggests that these toxins use a G protein-coupled receptor as a coreceptor on T cells.

Quantitative reduction of the TCR adapter protein SLP-76 unbalances immunity and immune regulation.

PubMed

Siggs, Owen M; Miosge, Lisa A; Daley, Stephen R; Asquith, Kelly; Foster, Paul S; Liston, Adrian; Goodnow, Christopher C

2015-03-15

Gene variants that disrupt TCR signaling can cause severe immune deficiency, yet less disruptive variants are sometimes associated with immune pathology. Null mutations of the gene encoding the scaffold protein Src homology 2 domain-containing leukocyte protein of 76 kDa (SLP-76), for example, cause an arrest of T cell positive selection, whereas a synthetic membrane-targeted allele allows limited positive selection but is associated with proinflammatory cytokine production and autoantibodies. Whether these and other enigmatic outcomes are due to a biochemical uncoupling of tolerogenic signaling, or simply a quantitative reduction of protein activity, remains to be determined. In this study we describe a splice variant of Lcp2 that reduced the amount of wild-type SLP-76 protein by ~90%, disrupting immunogenic and tolerogenic pathways to different degrees. Mutant mice produced excessive amounts of proinflammatory cytokines, autoantibodies, and IgE, revealing that simple quantitative reductions of SLP-76 were sufficient to trigger immune dysregulation. This allele reveals a dose-sensitive threshold for SLP-76 in the balance of immunity and immune dysregulation, a common disturbance of atypical clinical immune deficiencies. Copyright © 2015 by The American Association of Immunologists, Inc.

Narrowing the Gap: Preserving Repertoire Diversity Despite Clonal Selection during the CD4 T Cell Response.

PubMed

Merkenschlager, Julia; Kassiotis, George

2015-01-01

T cell immunity relies on the generation and maintenance of a diverse repertoire of T cell antigen receptors (TCRs). The strength of signaling emanating from the TCR dictates the fate of T cells during development, as well as during the immune response. Whereas development of new T cells in the thymus increases the available TCR repertoire, clonal selection during the immune response narrows TCR diversity through the outgrowth of clonotypes with the fittest TCR. To ensure maintenance of TCR diversity in the antigen-selected repertoire, specific mechanisms can be envisaged that facilitate the participation of T cell clonotypes with less than best fit TCRs. Here, we summarize the evidence for the existence of such mechanisms that can prevent the loss of diversity. A number of T cell-autonomous or extrinsic factors can reverse clonotypic hierarchies set by TCR affinity for given antigen. Although not yet complete, understanding of these factors and their mechanism of action will be critical in interventional attempts to mold the antigen-selected TCR repertoire.

T Cell Receptor Signaling in the Control of Regulatory T Cell Differentiation and Function

PubMed Central

Li, Ming O.; Rudensky, Alexander Y.

2016-01-01

Regulatory T cells (TReg cells), a specialized T cell lineage, have a pivotal function in the control of self-tolerance and inflammatory responses. Recent studies have revealed a discrete mode of TCR signaling that regulates Treg cell differentiation, maintenance and function and that impacts on gene expression, metabolism, cell adhesion and migration of these cells. Here, we discuss the emerging understanding of TCR-guided differentiation of Treg cells in the context of their function in health and disease. PMID:27026074

A signal strength priority based position estimation for mobile platforms

NASA Astrophysics Data System (ADS)

Kalgikar, Bhargav; Akopian, David; Chen, Philip

2010-01-01

Global Positioning System (GPS) products help to navigate while driving, hiking, boating, and flying. GPS uses a combination of orbiting satellites to determine position coordinates. This works great in most outdoor areas, but the satellite signals are not strong enough to penetrate inside most indoor environments. As a result, a new strain of indoor positioning technologies that make use of 802.11 wireless LANs (WLAN) is beginning to appear on the market. In WLAN positioning the system either monitors propagation delays between wireless access points and wireless device users to apply trilateration techniques or it maintains the database of location-specific signal fingerprints which is used to identify the most likely match of incoming signal data with those preliminary surveyed and saved in the database. In this paper we investigate the issue of deploying WLAN positioning software on mobile platforms with typically limited computational resources. We suggest a novel received signal strength rank order based location estimation system to reduce computational loads with a robust performance. The proposed system performance is compared to conventional approaches.

c-MPL provides tumor-targeted T-cell receptor-transgenic T cells with costimulation and cytokine signals.

PubMed

Nishimura, Christopher D; Brenner, Daniel A; Mukherjee, Malini; Hirsch, Rachel A; Ott, Leah; Wu, Meng-Fen; Liu, Hao; Dakhova, Olga; Orange, Jordan S; Brenner, Malcolm K; Lin, Charles Y; Arber, Caroline

2017-12-21

Adoptively transferred T-cell receptor (TCR)-engineered T cells depend on host-derived costimulation and cytokine signals for their full and sustained activation. However, in patients with cancer, both signals are frequently impaired. Hence, we developed a novel strategy that combines both essential signals in 1 transgene by expressing the nonlymphoid hematopoietic growth factor receptor c-MPL (myeloproliferative leukemia), the receptor for thrombopoietin (TPO), in T cells. c-MPL signaling activates pathways shared with conventional costimulatory and cytokine receptor signaling. Thus, we hypothesized that host-derived TPO, present in the tumor microenvironment, or pharmacological c-MPL agonists approved by the US Food and Drug Administration could deliver both signals to c-MPL-engineered TCR-transgenic T cells. We found that c-MPL + polyclonal T cells expand and proliferate in response to TPO, and persist longer after adoptive transfer in immunodeficient human TPO-transgenic mice. In TCR-transgenic T cells, c-MPL activation enhances antitumor function, T-cell expansion, and cytokine production and preserves a central memory phenotype. c-MPL signaling also enables sequential tumor cell killing, enhances the formation of effective immune synapses, and improves antileukemic activity in vivo in a leukemia xenograft model. We identify the type 1 interferon pathway as a molecular mechanism by which c-MPL mediates immune stimulation in T cells. In conclusion, we present a novel immunotherapeutic strategy using c-MPL-enhanced transgenic T cells responding to either endogenously produced TPO (a microenvironment factor in hematologic malignancies) or c-MPL-targeted pharmacological agents. © 2017 by The American Society of Hematology.

Evolution and Function of the TCR Vgamma9 Chain Repertoire: It’s Good to be Public

PubMed Central

Pauza, C. David; Cairo, Cristiana

2015-01-01

Lymphocytes expressing a T cell receptor (TCR) composed of Vgamma9 and Vdelta2 chains represent a minor fraction of human thymocytes. Extrathymic selection throughout post-natal life causes the proportion of cells with a Vgamma9-JP rearrangement to increase and elevates the capacity for responding to non-peptidic phosphoantigens. Extrathymic selection is so powerful that phosphoantigen-reactive cells comprise about 1 in 40 circulating memory T cells from healthy adults and the subset can be expanded rapidly upon infection or in response to malignancy. Skewing of the gamma delta TCR repertoire is accompanied by selection for public gamma chain sequences such that many unrelated individuals overlap extensive in their circulating repertoire. This type of selection implies the presence of a monomorphic antigen-presenting molecule that is an object of current research but remains incompletely defined. While selection on a monomorphic presenting molecule may seem unusual, similar mechanisms shape the alpha beta T cell repertoire including the extreme examples of NKT or mucosal-associated invariant T cells (MAIT) and the less dramatic amplification of public Vbeta chain rearrangements driven by individual MHC molecules and associated with resistance to viral pathogens. Selecting and amplifying public T cell receptors whether alpha beta or gamma delta, are important steps in developing an anticipatory TCR repertoire. Cell clones expressing public TCR can accelerate the kinetics of response to pathogens and impact host survival. PMID:25769734

T-cell receptor signaling activates an ITK/NF-κB/GATA-3 axis in T-cell lymphomas facilitating resistance to chemotherapy

PubMed Central

Wang, Tianjiao; Lu, Ye; Polk, Avery; Chowdhury, Pinki; Zamalloa, Carlos Murga; Fujiwara, Hiroshi; Suemori, Koichiro; Beyersdorf, Niklas; Hristov, Alexandra C.; Lim, Megan S.; Bailey, Nathanael G.; Wilcox, Ryan A.

2016-01-01

Purpose T-cell lymphomas are a molecularly heterogeneous group of non-Hodgkin lymphomas (NHL) that account for a disproportionate number of NHL disease-related deaths due to their inherent and acquired resistance to standard multiagent chemotherapy regimens. Despite their molecular heterogeneity and frequent loss of various T-cell specific receptors, the T-cell antigen receptor is retained in the majority of these lymphomas. As T-cell receptor (TCR) engagement activates a number of signaling pathways and transcription factors that regulate T-cell growth and survival, we examined the TCR’s role in mediating resistance to chemotherapy. Experimental Design Genetic and pharmacologic strategies were utilized to determine the contribution of tyrosine kinases and transcription factors activated in conventional T cells following T-cell receptor (TCR) engagement in acquired chemotherapy resistance in primary T-cell lymphoma cells and patient-derived cell lines. Results Here we report that TCR signaling activates a signaling axis that includes ITK, NF-κB, and GATA-3, and promotes chemotherapy resistance. Conclusions These observations have significant therapeutic implications, as pharmacologic inhibition of ITK prevented activation of this signaling axis and overcame chemotherapy resistance. PMID:27780854

Hippocampal Mismatch Signals Are Modulated by the Strength of Neural Predictions and Their Similarity to Outcomes.

PubMed

Long, Nicole M; Lee, Hongmi; Kuhl, Brice A

2016-12-14

The hippocampus is thought to compare predicted events with current perceptual input, generating a mismatch signal when predictions are violated. However, most prior studies have only inferred when predictions occur without measuring them directly. Moreover, an important but unresolved question is whether hippocampal mismatch signals are modulated by the degree to which predictions differ from outcomes. Here, we conducted a human fMRI study in which subjects repeatedly studied various word-picture pairs, learning to predict particular pictures (outcomes) from the words (cues). After initial learning, a subset of cues was paired with a novel, unexpected outcome, whereas other cues continued to predict the same outcome. Critically, when outcomes changed, the new outcome was either "near" to the predicted outcome (same visual category as the predicted picture) or "far" from the predicted outcome (different visual category). Using multivoxel pattern analysis, we indexed cue-evoked reactivation (prediction) within neocortical areas and related these trial-by-trial measures of prediction strength to univariate hippocampal responses to the outcomes. We found that prediction strength positively modulated hippocampal responses to unexpected outcomes, particularly when unexpected outcomes were close, but not identical, to the prediction. Hippocampal responses to unexpected outcomes were also associated with a tradeoff in performance during a subsequent memory test: relatively faster retrieval of new (updated) associations, but relatively slower retrieval of the original (older) associations. Together, these results indicate that hippocampal mismatch signals reflect a comparison between active predictions and current outcomes and that these signals are most robust when predictions are similar, but not identical, to outcomes. Although the hippocampus is widely thought to signal "mismatches" between memory-based predictions and outcomes, previous research has not linked

Conferring indirect allospecificity on CD4+CD25+ Tregs by TCR gene transfer favors transplantation tolerance in mice

PubMed Central

Tsang, Julia Yuen-Shan; Tanriver, Yakup; Jiang, Shuiping; Xue, Shao-An; Ratnasothy, Kulachelvy; Chen, Daxin; Stauss, Hans J.; Bucy, R. Pat; Lombardi, Giovanna; Lechler, Robert

2008-01-01

T cell responses to MHC-mismatched transplants can be mediated via direct recognition of allogeneic MHC molecules on the cells of the transplant or via recognition of allogeneic peptides presented on the surface of recipient APCs in recipient MHC molecules — a process known as indirect recognition. As CD4+CD25+ Tregs play an important role in regulating alloresponses, we investigated whether mouse Tregs specific for allogeneic MHC molecules could be generated in vitro and could promote transplantation tolerance in immunocompetent recipient mice. Tregs able to directly recognize allogeneic MHC class II molecules (dTregs) were obtained by stimulating CD4+CD25+ cells from C57BL/6 mice (H-2b) with allogeneic DCs from BALB/c mice (H-2d). To generate Tregs that indirectly recognized allogeneic MHC class II molecules, dTregs were retrovirally transduced with TCR genes conferring specificity for H-2Kd presented by H-2Ab MHC class II molecules. The dual direct and indirect allospecificity of the TCR-transduced Tregs was confirmed in vitro. In mice, TCR-transduced Tregs, but not dTregs, induced long-term survival of partially MHC-mismatched heart grafts when combined with short-term adjunctive immunosuppression. Further, although dTregs were only slightly less effective than TCR-transduced Tregs at inducing long-term survival of fully MHC-mismatched heart grafts, histologic analysis of long-surviving hearts demonstrated marked superiority of the TCR-transduced Tregs. Thus, Tregs specific for allogeneic MHC class II molecules are effective in promoting transplantation tolerance in mice, which suggests that such cells have clinical potential. PMID:18846251

T-cell receptor signaling enhances transcriptional elongation from latent HIV proviruses by activating P-TEFb through an ERK-dependent pathway.

PubMed

Kim, Young Kyeung; Mbonye, Uri; Hokello, Joseph; Karn, Jonathan

2011-07-29

Latent human immunodeficiency virus (HIV) proviruses are thought to be primarily reactivated in vivo through stimulation of the T-cell receptor (TCR). Activation of the TCR induces multiple signal transduction pathways, leading to the ordered nuclear migration of the HIV transcription initiation factors NF-κB (nuclear factor κB) and NFAT (nuclear factor of activated T-cells), as well as potential effects on HIV transcriptional elongation. We have monitored the kinetics of proviral reactivation using chromatin immunoprecipitation assays to measure changes in the distribution of RNA polymerase II in the HIV provirus. Surprisingly, in contrast to TNF-α (tumor necrosis factor α) activation, where early transcription elongation is highly restricted due to rate-limiting concentrations of Tat, efficient and sustained HIV elongation and positive transcription elongation factor b (P-TEFb) recruitment are detected immediately after the activation of latent proviruses through the TCR. Inhibition of NFAT activation by cyclosporine had no effect on either HIV transcription initiation or elongation. However, examination of P-TEFb complexes by gel-filtration chromatography showed that TCR signaling led to the rapid dissociation of the large inactive P-TEFb:7SK RNP (small nuclear RNA 7SK ribonucleoprotein) complex and the release of active low-molecular-weight P-TEFb complexes. Both P-TEFb recruitment to the HIV long terminal repeat and enhanced HIV processivity were blocked by the ERK (extracellular-signal-regulated kinase) inhibitor U0126, but not by AKT (serine/threonine protein kinase Akt) and PI3K (phosphatidylinositol 3-kinase) inhibitors. In contrast to treatment with HMBA (hexamethylene bisacetamide) and DRB (5,6-dichlorobenzimidazole 1-β-ribofuranoside), which disrupt the large 7SK RNP complex but do not stimulate early HIV elongation, TCR signaling provides the first example of a physiological pathway that can shift the balance between the inactive P-TEFb pool and

An alternative mode of CD43 signal transduction activates pro-survival pathways of T lymphocytes.

PubMed

Bravo-Adame, Maria Elena; Vera-Estrella, Rosario; Barkla, Bronwyn J; Martínez-Campos, Cecilia; Flores-Alcantar, Angel; Ocelotl-Oviedo, Jose Pablo; Pedraza-Alva, Gustavo; Rosenstein, Yvonne

2017-01-01

CD43 is one of the most abundant co-stimulatory molecules on a T-cell surface; it transduces activation signals through its cytoplasmic domain, contributing to modulation of the outcome of T-cell responses. The aim of this study was to uncover new signalling pathways regulated by this sialomucin. Analysis of changes in protein abundance allowed us to identify pyruvate kinase isozyme M2 (PKM2), an enzyme of the glycolytic pathway, as an element potentially participating in the signalling cascade resulting from the engagement of CD43 and the T-cell receptor (TCR). We found that the glycolytic activity of this enzyme was not significantly increased in response to TCR+CD43 co-stimulation, but that PKM2 was tyrosine phosphorylated, suggesting that it was performing moonlight functions. We report that phosphorylation of both Y 105 of PKM2 and of Y 705 of signal transducer and activator of transcription 3 was induced in response to TCR+CD43 co-stimulation, resulting in activation of the mitogen-activated protein kinase kinase 5/extracellular signal-regulated kinase 5 (MEK5/ERK5) pathway. ERK5 and the cAMP response element binding protein (CREB) were activated, and c-Myc and nuclear factor-κB (p65) nuclear localization, as well as Bad phosphorylation, were augmented. Consistent with this, expression of human CD43 in a murine T-cell hybridoma favoured cell survival. Altogether, our data highlight novel signalling pathways for the CD43 molecule in T lymphocytes, and underscore a role for CD43 in promoting cell survival through non-glycolytic functions of metabolic enzymes. © 2016 John Wiley & Sons Ltd.

T cell receptor for antigen induces linker for activation of T cell–dependent activation of a negative signaling complex involving Dok-2, SHIP-1, and Grb-2

PubMed Central

Dong, Shen; Corre, Béatrice; Foulon, Eliane; Dufour, Evelyne; Veillette, André; Acuto, Oreste; Michel, Frédérique

2006-01-01

Adaptor proteins positively or negatively regulate the T cell receptor for antigen (TCR) signaling cascade. We report that after TCR stimulation, the inhibitory adaptor downstream of kinase (Dok)-2 and its homologue Dok-1 are involved in a multimolecular complex including the lipid phosphatase Src homology 2 domain–containing inositol polyphosphate 5′-phosphatase (SHIP)-1 and Grb-2 which interacts with the membrane signaling scaffold linker for activation of T cells (LAT). Knockdown of LAT and SHIP-1 expression indicated that SHIP-1 favored recruitment of Dok-2 to LAT. Knockdown of Dok-2 and Dok-1 revealed their negative control on Akt and, unexpectedly, on Zap-70 activation. Our findings support the view that Dok-1 and -2 are critical elements of a LAT-dependent negative feedback loop that attenuates early TCR signal. Dok-1 and -2 may therefore exert a critical role in shaping the immune response and as gatekeepers for T cell tolerance. PMID:17043143

Interaction between Sam68 and Src family tyrosine kinases, Fyn and Lck, in T cell receptor signaling.

PubMed

Fusaki, N; Iwamatsu, A; Iwashima, M; Fujisawa, J i

1997-03-07

The Src family protein-tyrosine kinase, Fyn, is associated with the T cell receptor (TCR) and plays an important role in TCR-mediated signaling. We found that a human T cell leukemia virus type 1-infected T cell line, Hayai, overexpressed Fyn. To identify the molecules downstream of Fyn, we analyzed the tyrosine phosphorylation of cellular proteins in the cells. In Hayai, a 68-kDa protein was constitutively tyrosine-phosphorylated. The 68-kDa protein was coimmunoprecipitated with various signaling proteins such as phospholipase C gamma1, the phosphatidylinositol 3-kinase p85 subunit, Grb2, SHP-1, Cbl, and Jak3, implying that the protein might function as an adapter. Purification and microsequencing of this protein revealed that it was the RNA-binding protein, Sam68 (Src associated in mitosis, 68 kDa). Sam68 was associated with the Src homology 2 and 3 domains of Fyn and also those of another Src family kinase, Lck. CD3 cross-linking induced tyrosine phosphorylation of Sam68 in uninfected T cells. These data suggest that Sam68 participates in the signal transduction pathway downstream of TCR-coupled Src family kinases Fyn and Lck in lymphocytes, that is not only in the mitotic pathway downstream of c-Src in fibroblasts.

Proline substitution independently enhances H-2D(b) complex stabilization and TCR recognition of melanoma-associated peptides.

PubMed

Uchtenhagen, Hannes; Abualrous, Esam T; Stahl, Evi; Allerbring, Eva B; Sluijter, Marjolein; Zacharias, Martin; Sandalova, Tatyana; van Hall, Thorbald; Springer, Sebastian; Nygren, Per-Åke; Achour, Adnane

2013-11-01

The immunogenicity of H-2D(b) (D(b)) restricted epitopes can be significantly increased by substituting peptide position 3 to a proline (p3P). The p3P modification enhances MHC stability without altering the conformation of the modified epitope allowing for T-cell cross-reactivity with the native peptide. The present study reveals how specific interactions between p3P and the highly conserved MHC heavy chain residue Y159 increase the stability of D(b) in complex with an optimized version of the melanoma-associated epitope gp10025-33 . Furthermore, the p3P modification directly increased the affinity of the D(b)/gp10025-33 -specific T-cell receptor (TCR) pMel. Surprisingly, the enhanced TCR binding was independent from the observed increased stability of the optimized D(b)/gp10025-33 complex and from the interactions formed between p3P and Y159, indicating a direct effect of the p3P modification on TCR recognition. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A Discrete Ubiquitin-Mediated Network Regulates the Strength of NOD2 Signaling

PubMed Central

Tigno-Aranjuez, Justine T.; Bai, Xiaodong

2013-01-01

Dysregulation of NOD2 signaling is implicated in the pathology of various inflammatory diseases, including Crohn's disease, asthma, and sarcoidosis, making signaling proteins downstream of NOD2 potential therapeutic targets. Inhibitor-of-apoptosis (IAP) proteins, particularly cIAP1, are essential mediators of NOD2 signaling, and in this work, we describe a molecular mechanism for cIAP1's regulation in the NOD2 signaling pathway. While cIAP1 promotes RIP2's tyrosine phosphorylation and subsequent NOD2 signaling, this positive regulation is countered by another E3 ubiquitin ligase, ITCH, through direct ubiquitination of cIAP1. This ITCH-mediated ubiquitination leads to cIAP1's lysosomal degradation. Pharmacologic inhibition of cIAP1 expression in ITCH−/− macrophages attenuates heightened ITCH−/− macrophage muramyl dipeptide-induced responses. Transcriptome analysis, combined with pharmacologic inhibition of cIAP1, further defines specific pathways within the NOD2 signaling pathway that are targeted by cIAP1. This information provides genetic signatures that may be useful in repurposing cIAP1-targeted therapies to correct NOD2-hyperactive states and identifies a ubiquitin-regulated signaling network centered on ITCH and cIAP1 that controls the strength of NOD2 signaling. PMID:23109427

Targeting of HPV-16+ Epithelial Cancer Cells by TCR Gene Engineered T Cells Directed against E6.

PubMed

Draper, Lindsey M; Kwong, Mei Li M; Gros, Alena; Stevanović, Sanja; Tran, Eric; Kerkar, Sid; Raffeld, Mark; Rosenberg, Steven A; Hinrichs, Christian S

2015-10-01

The E6 and E7 oncoproteins of HPV-associated epithelial cancers are in principle ideal immunotherapeutic targets, but evidence that T cells specific for these antigens can recognize and kill HPV(+) tumor cells is limited. We sought to determine whether TCR gene engineered T cells directed against an HPV oncoprotein can successfully target HPV(+) tumor cells. T-cell responses against the HPV-16 oncoproteins were investigated in a patient with an ongoing 22-month disease-free interval after her second resection of distant metastatic anal cancer. T cells genetically engineered to express an oncoprotein-specific TCR from this patient's tumor-infiltrating T cells were tested for specific reactivity against HPV(+) epithelial tumor cells. We identified, from an excised metastatic anal cancer tumor, T cells that recognized an HLA-A*02:01-restricted epitope of HPV-16 E6. The frequency of the dominant T-cell clonotype from these cells was approximately 400-fold greater in the patient's tumor than in her peripheral blood. T cells genetically engineered to express the TCR from this clonotype displayed high avidity for an HLA-A*02:01-restricted epitope of HPV-16, and they showed specific recognition and killing of HPV-16(+) cervical, and head and neck cancer cell lines. These findings demonstrate that HPV-16(+) tumors can be targeted by E6-specific TCR gene engineered T cells, and they provide the foundation for a novel cellular therapy directed against HPV-16(+) malignancies, including cervical, oropharyngeal, anal, vulvar, vaginal, and penile cancers. ©2015 American Association for Cancer Research.

The phosphatase JKAP/DUSP22 inhibits T-cell receptor signalling and autoimmunity by inactivating Lck.

PubMed

Li, Ju-Pi; Yang, Chia-Yu; Chuang, Huai-Chia; Lan, Joung-Liang; Chen, Der-Yuan; Chen, Yi-Ming; Wang, Xiaohong; Chen, Alice J; Belmont, John W; Tan, Tse-Hua

2014-04-09

JNK pathway-associated phosphatase (JKAP, also known as DUSP22 or JSP-1) is a JNK activator. The in vivo role of JKAP in immune regulation remains unclear. Here we report that JKAP directly inactivates Lck by dephosphorylating tyrosine-394 residue during T-cell receptor (TCR) signalling. JKAP-knockout T cells display enhanced cell proliferation and cytokine production. JKAP-knockout mice show enhanced T-cell-mediated immune responses and are more susceptible to experimental autoimmune encephalomyelitis (EAE). In addition, the recipient mice that are adoptively transferred with JKAP-knockout T cells show exacerbated EAE symptoms. Aged JKAP-knockout mice spontaneously develop inflammation and autoimmunity. Thus, our results indicate that JKAP is an important phosphatase that inactivates Lck in the TCR signalling turn-off stage, leading to suppression of T-cell-mediated immunity and autoimmunity.

TCR-engineered, customized, antitumor T cells for cancer immunotherapy: advantages and limitations.

PubMed

Chhabra, Arvind

2011-01-05

The clinical outcome of the traditional adoptive cancer immunotherapy approaches involving the administration of donor-derived immune effectors, expanded ex vivo, has not met expectations. This could be attributed, in part, to the lack of sufficient high-avidity antitumor T-cell precursors in most cancer patients, poor immunogenicity of cancer cells, and the technological limitations to generate a sufficiently large number of tumor antigen-specific T cells. In addition, the host immune regulatory mechanisms and immune homeostasis mechanisms, such as activation-induced cell death (AICD), could further limit the clinical efficacy of the adoptively administered antitumor T cells. Since generation of a sufficiently large number of potent antitumor immune effectors for adoptive administration is critical for the clinical success of this approach, recent advances towards generating customized donor-specific antitumor-effector T cells by engrafting human peripheral blood-derived T cells with a tumor-associated antigen-specific transgenic T-cell receptor (TCR) are quite interesting. This manuscript provides a brief overview of the TCR engineering-based cancer immunotherapy approach, its advantages, and the current limitations.

The complex and specific pMHC interactions with diverse HIV-1 TCR clonotypes reveal a structural basis for alterations in CTL function

PubMed Central

Xia, Zhen; Chen, Huabiao; Kang, Seung-gu; Huynh, Tien; Fang, Justin W.; Lamothe, Pedro A.; Walker, Bruce D.; Zhou, Ruhong

2014-01-01

Immune control of viral infections is modulated by diverse T cell receptor (TCR) clonotypes engaging peptide-MHC class I complexes on infected cells, but the relationship between TCR structure and antiviral function is unclear. Here we apply in silico molecular modeling with in vivo mutagenesis studies to investigate TCR-pMHC interactions from multiple CTL clonotypes specific for a well-defined HIV-1 epitope. Our molecular dynamics simulations of viral peptide-HLA-TCR complexes, based on two independent co-crystal structure templates, reveal that effective and ineffective clonotypes bind to the terminal portions of the peptide-MHC through similar salt bridges, but their hydrophobic side-chain packings can be very different, which accounts for the major part of the differences among these clonotypes. Non-specific hydrogen bonding to viral peptide also accommodates greater epitope variants. Furthermore, free energy perturbation calculations for point mutations on the viral peptide KK10 show excellent agreement with in vivo mutagenesis assays, with new predictions confirmed by additional experiments. These findings indicate a direct structural basis for heterogeneous CTL antiviral function. PMID:24522437

The complex and specific pMHC interactions with diverse HIV-1 TCR clonotypes reveal a structural basis for alterations in CTL function

NASA Astrophysics Data System (ADS)

Xia, Zhen; Chen, Huabiao; Kang, Seung-Gu; Huynh, Tien; Fang, Justin W.; Lamothe, Pedro A.; Walker, Bruce D.; Zhou, Ruhong

2014-02-01

Immune control of viral infections is modulated by diverse T cell receptor (TCR) clonotypes engaging peptide-MHC class I complexes on infected cells, but the relationship between TCR structure and antiviral function is unclear. Here we apply in silico molecular modeling with in vivo mutagenesis studies to investigate TCR-pMHC interactions from multiple CTL clonotypes specific for a well-defined HIV-1 epitope. Our molecular dynamics simulations of viral peptide-HLA-TCR complexes, based on two independent co-crystal structure templates, reveal that effective and ineffective clonotypes bind to the terminal portions of the peptide-MHC through similar salt bridges, but their hydrophobic side-chain packings can be very different, which accounts for the major part of the differences among these clonotypes. Non-specific hydrogen bonding to viral peptide also accommodates greater epitope variants. Furthermore, free energy perturbation calculations for point mutations on the viral peptide KK10 show excellent agreement with in vivo mutagenesis assays, with new predictions confirmed by additional experiments. These findings indicate a direct structural basis for heterogeneous CTL antiviral function.

Human and Murine Clonal CD8+ T Cell Expansions Arise during Tuberculosis Because of TCR Selection

PubMed Central

Nunes-Alves, Cláudio; Booty, Matthew G.; Carpenter, Stephen M.; Rothchild, Alissa C.; Martin, Constance J.; Desjardins, Danielle; Steblenko, Katherine; Kløverpris, Henrik N.; Madansein, Rajhmun; Ramsuran, Duran; Leslie, Alasdair; Correia-Neves, Margarida; Behar, Samuel M.

2015-01-01

The immune system can recognize virtually any antigen, yet T cell responses against several pathogens, including Mycobacterium tuberculosis, are restricted to a limited number of immunodominant epitopes. The host factors that affect immunodominance are incompletely understood. Whether immunodominant epitopes elicit protective CD8+ T cell responses or instead act as decoys to subvert immunity and allow pathogens to establish chronic infection is unknown. Here we show that anatomically distinct human granulomas contain clonally expanded CD8+ T cells with overlapping T cell receptor (TCR) repertoires. Similarly, the murine CD8+ T cell response against M. tuberculosis is dominated by TB10.44-11-specific T cells with extreme TCRβ bias. Using a retrogenic model of TB10.44-11-specific CD8+ T cells, we show that TCR dominance can arise because of competition between clonotypes driven by differences in affinity. Finally, we demonstrate that TB10.4-specific CD8+ T cells mediate protection against tuberculosis, which requires interferon-γ production and TAP1-dependent antigen presentation in vivo. Our study of how immunodominance, biased TCR repertoires, and protection are inter-related, provides a new way to measure the quality of T cell immunity, which if applied to vaccine evaluation, could enhance our understanding of how to elicit protective T cell immunity. PMID:25945999

Human and Murine Clonal CD8+ T Cell Expansions Arise during Tuberculosis Because of TCR Selection.

PubMed

Nunes-Alves, Cláudio; Booty, Matthew G; Carpenter, Stephen M; Rothchild, Alissa C; Martin, Constance J; Desjardins, Danielle; Steblenko, Katherine; Kløverpris, Henrik N; Madansein, Rajhmun; Ramsuran, Duran; Leslie, Alasdair; Correia-Neves, Margarida; Behar, Samuel M

2015-05-01

The immune system can recognize virtually any antigen, yet T cell responses against several pathogens, including Mycobacterium tuberculosis, are restricted to a limited number of immunodominant epitopes. The host factors that affect immunodominance are incompletely understood. Whether immunodominant epitopes elicit protective CD8+ T cell responses or instead act as decoys to subvert immunity and allow pathogens to establish chronic infection is unknown. Here we show that anatomically distinct human granulomas contain clonally expanded CD8+ T cells with overlapping T cell receptor (TCR) repertoires. Similarly, the murine CD8+ T cell response against M. tuberculosis is dominated by TB10.44-11-specific T cells with extreme TCRβ bias. Using a retro genic model of TB10.44-11-specific CD8+ Tcells, we show that TCR dominance can arise because of competition between clonotypes driven by differences in affinity. Finally, we demonstrate that TB10.4-specific CD8+ T cells mediate protection against tuberculosis, which requires interferon-γ production and TAP1-dependent antigen presentation in vivo. Our study of how immunodominance, biased TCR repertoires, and protection are inter-related, provides a new way to measure the quality of T cell immunity, which if applied to vaccine evaluation, could enhance our understanding of how to elicit protective T cell immunity.

Normalization of neuronal responses in cortical area MT across signal strengths and motion directions

PubMed Central

Xiao, Jianbo; Niu, Yu-Qiong; Wiesner, Steven

2014-01-01

Multiple visual stimuli are common in natural scenes, yet it remains unclear how multiple stimuli interact to influence neuronal responses. We investigated this question by manipulating relative signal strengths of two stimuli moving simultaneously within the receptive fields (RFs) of neurons in the extrastriate middle temporal (MT) cortex. Visual stimuli were overlapping random-dot patterns moving in two directions separated by 90°. We first varied the motion coherence of each random-dot pattern and characterized, across the direction tuning curve, the relationship between neuronal responses elicited by bidirectional stimuli and by the constituent motion components. The tuning curve for bidirectional stimuli showed response normalization and can be accounted for by a weighted sum of the responses to the motion components. Allowing nonlinear, multiplicative interaction between the two component responses significantly improved the data fit for some neurons, and the interaction mainly had a suppressive effect on the neuronal response. The weighting of the component responses was not fixed but dependent on relative signal strengths. When two stimulus components moved at different coherence levels, the response weight for the higher-coherence component was significantly greater than that for the lower-coherence component. We also varied relative luminance levels of two coherently moving stimuli and found that MT response weight for the higher-luminance component was also greater. These results suggest that competition between multiple stimuli within a neuron's RF depends on relative signal strengths of the stimuli and that multiplicative nonlinearity may play an important role in shaping the response tuning for multiple stimuli. PMID:24899674

The chemokine CXCL12 generates costimulatory signals in T cells to enhance phosphorylation and clustering of the adaptor protein SLP-76.

PubMed

Smith, Xin; Schneider, Helga; Köhler, Karsten; Liu, Hebin; Lu, Yuning; Rudd, Christopher E

2013-07-30

The CXC chemokine CXCL12 mediates the chemoattraction of T cells and enhances the stimulation of T cells through the T cell receptor (TCR). The adaptor SLP-76 [Src homology 2 (SH2) domain-containing leukocyte protein of 76 kD] has two key tyrosine residues, Tyr(113) and Tyr(128), that mediate signaling downstream of the TCR. We investigated the effect of CXCL12 on SLP-76 phosphorylation and the TCR-dependent formation of SLP-76 microclusters. Although CXCL12 alone failed to induce SLP-76 cluster formation, it enhanced the number, stability, and phosphorylation of SLP-76 microclusters formed in response to stimulation of the TCR by an activating antibody against CD3, a component of the TCR complex. Addition of CXCL12 to anti-CD3-stimulated cells resulted in F-actin polymerization that stabilized SLP-76 microclusters in the cells' periphery at the interface with antibody-coated coverslips and increased the interaction between SLP-76 clusters and those containing ZAP-70, the TCR-associated kinase that phosphorylates SLP-76, as well as increased TCR-dependent gene expression. Costimulation with CXCL12 and anti-CD3 increased the extent of phosphorylation of SLP-76 at Tyr(113) and Tyr(128), but not that of other TCR-proximal components, and mutation of either one of these residues impaired the CXCL12-dependent effect on SLP-76 microcluster formation, F-actin polymerization, and TCR-dependent gene expression. The effects of CXCL12 on SLP-76 microcluster formation were dependent on the coupling of its receptor CXCR4 to G(i)-family G proteins (heterotrimeric guanine nucleotide-binding proteins). Thus, we identified a costimulatory mechanism by which CXCL12 and antigen converge at SLP-76 microcluster formation to enhance T cell responses.

Recovery of strength is dependent on mTORC1 signaling after eccentric muscle injury.

PubMed

Baumann, Cory Walter; Rogers, Russell George; Otis, Jeffrey Scott; Ingalls, Christopher Paul

2016-11-01

Eccentric contractions may cause immediate and long-term reductions in muscle strength that can be recovered through increased protein synthesis rates. The purpose of this study was to determine whether the mechanistic target-of-rapamycin complex 1 (mTORC1), a vital controller of protein synthesis rates, is required for return of muscle strength after injury. Isometric muscle strength was assessed before, immediately after, and then 3, 7, and 14 days after a single bout of 150 eccentric contractions in mice that received daily injections of saline or rapamycin. The bout of eccentric contractions increased the phosphorylation of mTORC1 (1.8-fold) and p70s6k1 (13.8-fold), mTORC1's downstream effector, 3 days post-injury. Rapamycin blocked mTORC1 and p70s6k1 phosphorylation and attenuated recovery of muscle strength (∼20%) at 7 and 14 days. mTORC1 signaling is instrumental in the return of muscle strength after a single bout of eccentric contractions in mice. Muscle Nerve 54: 914-924, 2016. © 2016 Wiley Periodicals, Inc.

Combining in silico evolution and nonlinear dimensionality reduction to redesign responses of signaling networks

NASA Astrophysics Data System (ADS)

Prescott, Aaron M.; Abel, Steven M.

2016-12-01

The rational design of network behavior is a central goal of synthetic biology. Here, we combine in silico evolution with nonlinear dimensionality reduction to redesign the responses of fixed-topology signaling networks and to characterize sets of kinetic parameters that underlie various input-output relations. We first consider the earliest part of the T cell receptor (TCR) signaling network and demonstrate that it can produce a variety of input-output relations (quantified as the level of TCR phosphorylation as a function of the characteristic TCR binding time). We utilize an evolutionary algorithm (EA) to identify sets of kinetic parameters that give rise to: (i) sigmoidal responses with the activation threshold varied over 6 orders of magnitude, (ii) a graded response, and (iii) an inverted response in which short TCR binding times lead to activation. We also consider a network with both positive and negative feedback and use the EA to evolve oscillatory responses with different periods in response to a change in input. For each targeted input-output relation, we conduct many independent runs of the EA and use nonlinear dimensionality reduction to embed the resulting data for each network in two dimensions. We then partition the results into groups and characterize constraints placed on the parameters by the different targeted response curves. Our approach provides a way (i) to guide the design of kinetic parameters of fixed-topology networks to generate novel input-output relations and (ii) to constrain ranges of biological parameters using experimental data. In the cases considered, the network topologies exhibit significant flexibility in generating alternative responses, with distinct patterns of kinetic rates emerging for different targeted responses.

Investigating Strength and Frequency Effects in Recognition Memory Using Type-2 Signal Detection Theory

ERIC Educational Resources Information Center

Higham, Philip A.; Perfect, Timothy J.; Bruno, Davide

2009-01-01

Criterion- versus distribution-shift accounts of frequency and strength effects in recognition memory were investigated with Type-2 signal detection receiver operating characteristic (ROC) analysis, which provides a measure of metacognitive monitoring. Experiment 1 demonstrated a frequency-based mirror effect, with a higher hit rate and lower…

BCL11B enhances TCR/CD28-triggered NF-kappaB activation through up-regulation of Cot kinase gene expression in T-lymphocytes.

PubMed

Cismasiu, Valeriu B; Duque, Javier; Paskaleva, Elena; Califano, Danielle; Ghanta, Sailaja; Young, Howard A; Avram, Dorina

2009-01-15

BCL11B is a transcriptional regulator with an important role in T-cell development and leukaemogenesis. We demonstrated recently that BCL11B controls expression from the IL (interleukin)-2 promoter through direct binding to the US1 (upstream site 1). In the present study, we provide evidence that BCL11B also participates in the activation of IL-2 gene expression by enhancing NF-kappaB (nuclear factor kappaB) activity in the context of TCR (T-cell receptor)/CD28-triggered T-cell activation. Enhanced NF-kappaB activation is not a consequence of BCL11B binding to the NF-kappaB response elements or association with the NF-kappaB-DNA complexes, but rather the result of higher translocation of NF-kappaB to the nucleus caused by enhanced degradation of IkappaB (inhibitor of NF-kappaB). The enhanced IkappaB degradation in cells with increased levels of BCL11B was specific for T-cells activated through the TCR, but not for cells activated through TNFalpha (tumour necrosis factor alpha) or UV light, and was caused by increased activity of IkappaB kinase, as indicated by its increase in phosphorylation. As BCL11B is a transcription factor, we investigated whether the expression of genes upstream of IkappaB kinase in the TCR/CD28 signalling pathway was affected by increased BCL11B expression, and found that Cot (cancer Osaka thyroid oncogene) kinase mRNA levels were elevated. Cot kinase is known to promote enhanced IkappaB kinase activity, which results in the phosphorylation and degradation of IkappaB and activation of NF-kappaB. The implied involvement of Cot kinase in BCL11B-mediated NF-kappaB activation in response to TCR activation is supported by the fact that a Cot kinase dominant-negative mutant or Cot kinase siRNA (small interfering RNA) knockdown blocked BCL11B-mediated NF-kappaB activation. In support of our observations, in the present study we report that BCL11B enhances the expression of several other NF-kappaB target genes, in addition to IL-2. In addition, we

Platypus TCRμ provides insight into the origins and evolution of a uniquely mammalian TCR locus.

PubMed

Wang, Xinxin; Parra, Zuly E; Miller, Robert D

2011-11-15

TCRμ is an unconventional TCR that was first discovered in marsupials and appears to be absent from placental mammals and nonmammals. In this study, we show that TCRμ is also present in the duckbill platypus, an egg-laying monotreme, consistent with TCRμ being ancient and present in the last common ancestor of all extant mammals. As in marsupials, platypus TCRμ is expressed in a form containing double V domains. These V domains more closely resemble Ab V than that of conventional TCR. Platypus TCRμ differs from its marsupial homolog by requiring two rounds of somatic DNA recombination to assemble both V exons and has a genomic organization resembling the likely ancestral form of the receptor genes. These results demonstrate that the ancestors of placental mammals would have had TCRμ but it has been lost from this lineage.

TCR gamma chain diversity in the spleen of the duckbill platypus (Ornithorhynchus anatinus).

PubMed

Parra, Zuly E; Arnold, Tamara; Nowak, Melissa A; Hellman, Lars; Miller, Robert D

2006-01-01

TCR gamma (TRG) chain diversity in splenic gammadelta T cells was determined for an egg-laying mammal (or monotreme), the duckbill platypus. Three distinct V subgroups were found in the expressed TRG chains and these three subgroups are members of a clade not found so far in eutherian mammals or birds. Each subgroup contains approximately five V gene segments, and their overall divergence is much less than is found in eutherians and birds, consistent with their recent evolution from an ancestral V gene segment. The platypus TRG locus also contains three C region genes and many of the residues involved in TCR function, such as interactions with CD3, were conserved in the monotreme C regions. All non-eutherian mammals (monotremes and marsupials) lacked the second cysteine residue necessary to form the intradomain disulfide bond in the C region, a loss apparently due to independent mutations in marsupials and monotremes. Monotreme TRGC regions also had among the most variation in the length of the connecting peptide region described for any species due to repeated motifs.

Antigen sensitivity of CD22-specific chimeric T cell receptors is modulated by target epitope distance from the cell membrane

PubMed Central

James, Scott E.; Greenberg, Philip D.; Jensen, Michael C.; Lin, Yukang; Wang, Jinjuan; Till, Brian G.; Raubitschek, Andrew A.; Forman, Stephen J.; Press, Oliver W.

2008-01-01

We have targeted CD22 as a novel tumor-associated antigen for recognition by human CTL genetically modified to express chimeric T cell receptors (cTCR) recognizing this surface molecule. CD22-specifc cTCR targeting different epitopes of the CD22 molecule promoted efficient lysis of target cells expressing high levels of CD22 with a maximum lytic potential that appeared to decrease as the distance of the target epitope from the target cell membrane increased. Targeting membrane-distal CD22 epitopes with cTCR+ CTL revealed defects in both degranulation and lytic granule targeting. CD22-specific cTCR+ CTL exhibited lower levels of maximum lysis and lower antigen sensitivity than CTL targeting CD20, which has a shorter extracellular domain than CD22. This diminished sensitivity was not a result of reduced avidity of antigen engagement, but instead reflected weaker signaling per triggered cTCR molecule when targeting membrane-distal epitopes of CD22. Both of these parameters were restored by targeting a ligand expressing the same epitope but constructed as a truncated CD22 molecule to approximate the length of a TCR:pMHC complex. The reduced sensitivity of CD22-specific cTCR+ CTL for antigen-induced triggering of effector functions has potential therapeutic applications, as such cells selectively lysed B cell lymphoma lines expressing high levels of CD22 but demonstrated minimal activity against autologous normal B cells, which express lower levels of CD22. Thus, our results demonstrate that cTCR signal strength – and consequently antigen sensitivity – can be modulated by differential choice of target epitopes with respect to distance from the cell membrane, allowing discrimination between targets with disparate antigen density. PMID:18453625

Can the Accuracy of Home Blood Glucose Monitors be affected by the Received Signal Strength of 900 MHz GSM Mobile Phones?

PubMed

Eslami, J; Ghafaripour, F; Mortazavi, S A R; Mortazavi, S M J; Shojaei-Fard, M B

2015-12-01

People who use home blood glucose monitors may use their mobile phones in the close vicinity of medical devices. This study is aimed at investigating the effect of the signal strength of 900 MHz GSM mobile phones on the accuracy of home blood glucose monitors. Sixty non-diabetic volunteer individuals aged 21 - 28 years participated in this study. Blood samples were analyzed for glucose level by using a common blood glucose monitoring system. Each blood sample was analyzed twice, within ten minutes in presence and absence of electromagnetic fields generated by a common GSM mobile phone during ringing. Blood samples were divided into 3 groups of 20 samples each. Group 1: exposure to mobile phone radiation with weak signal strength. Group2: exposure to mobile phone radiation with strong signal strength. Group3: exposure to a switched-on mobile phone with no signal strength. The magnitude of the changes in the first, second and third group between glucose levels of two measurements (׀ΔC׀) were 7.4±3.9 mg/dl, 10.2±4.5 mg/dl, 8.7±8.4 mg/dl respectively. The difference in the magnitude of the changes between the 1st and the 3rd groups was not statistically significant. Furthermore, the difference in the magnitude of the changes between the 2nd and the 3rd groups was not statistically significant. Findings of this study showed that the signal strength of 900 MHz GSM mobile phones cannot play a significant role in changing the accuracy of home blood glucose monitors.

Tonic LAT-HDAC7 Signals Sustain Nur77 and Irf4 Expression to Tune Naive CD4 T Cells.

PubMed

Myers, Darienne R; Lau, Tannia; Markegard, Evan; Lim, Hyung W; Kasler, Herbert; Zhu, Minghua; Barczak, Andrea; Huizar, John P; Zikherman, Julie; Erle, David J; Zhang, Weiguo; Verdin, Eric; Roose, Jeroen P

2017-05-23

CD4 + T cells differentiate into T helper cell subsets in feedforward manners with synergistic signals from the T cell receptor (TCR), cytokines, and lineage-specific transcription factors. Naive CD4 + T cells avoid spontaneous engagement of feedforward mechanisms but retain a prepared state. T cells lacking the adaptor molecule LAT demonstrate impaired TCR-induced signals yet cause a spontaneous lymphoproliferative T helper 2 (T H 2) cell syndrome in mice. Thus, LAT constitutes an unexplained maintenance cue. Here, we demonstrate that tonic signals through LAT constitutively export the repressor HDAC7 from the nucleus of CD4 + T cells. Without such tonic signals, HDAC7 target genes Nur77 and Irf4 are repressed. We reveal that Nur77 suppresses CD4 + T cell proliferation and uncover a suppressive role for Irf4 in T H 2 polarization; halving Irf4 gene-dosage leads to increases in GATA3 + and IL-4 + cells. Our studies reveal that naive CD4 + T cells are dynamically tuned by tonic LAT-HDAC7 signals. Published by Elsevier Inc.

Non-T cell activation linker (NTAL): a transmembrane adaptor protein involved in immunoreceptor signaling.

PubMed

Brdicka, Tomás; Imrich, Martin; Angelisová, Pavla; Brdicková, Nadezda; Horváth, Ondrej; Spicka, Jirí; Hilgert, Ivan; Lusková, Petra; Dráber, Petr; Novák, Petr; Engels, Niklas; Wienands, Jürgen; Simeoni, Luca; Osterreicher, Jan; Aguado, Enrique; Malissen, Marie; Schraven, Burkhart; Horejsí, Václav

2002-12-16

A key molecule necessary for activation of T lymphocytes through their antigen-specific T cell receptor (TCR) is the transmembrane adaptor protein LAT (linker for activation of T cells). Upon TCR engagement, LAT becomes rapidly tyrosine phosphorylated and then serves as a scaffold organizing a multicomponent complex that is indispensable for induction of further downstream steps of the signaling cascade. Here we describe the identification and preliminary characterization of a novel transmembrane adaptor protein that is structurally and evolutionarily related to LAT and is expressed in B lymphocytes, natural killer (NK) cells, monocytes, and mast cells but not in resting T lymphocytes. This novel transmembrane adaptor protein, termed NTAL (non-T cell activation linker) is the product of a previously identified WBSCR5 gene of so far unknown function. NTAL becomes rapidly tyrosine-phosphorylated upon cross-linking of the B cell receptor (BCR) or of high-affinity Fcgamma- and Fc epsilon -receptors of myeloid cells and then associates with the cytoplasmic signaling molecules Grb2, Sos1, Gab1, and c-Cbl. NTAL expressed in the LAT-deficient T cell line J.CaM2.5 becomes tyrosine phosphorylated and rescues activation of Erk1/2 and minimal transient elevation of cytoplasmic calcium level upon TCR/CD3 cross-linking. Thus, NTAL appears to be a structural and possibly also functional homologue of LAT in non-T cells.

Enhanced clinical-scale manufacturing of TCR transduced T-cells using closed culture system modules.

PubMed

Jin, Jianjian; Gkitsas, Nikolaos; Fellowes, Vicki S; Ren, Jiaqiang; Feldman, Steven A; Hinrichs, Christian S; Stroncek, David F; Highfill, Steven L

2018-01-24

Genetic engineering of T-cells to express specific T cell receptors (TCR) has emerged as a novel strategy to treat various malignancies. More widespread utilization of these types of therapies has been somewhat constrained by the lack of closed culture processes capable of expanding sufficient numbers of T-cells for clinical application. Here, we evaluate a process for robust clinical grade manufacturing of TCR gene engineered T-cells. TCRs that target human papillomavirus E6 and E7 were independently tested. A 21 day process was divided into a transduction phase (7 days) and a rapid expansion phase (14 days). This process was evaluated using two healthy donor samples and four samples obtained from patients with epithelial cancers. The process resulted in ~ 2000-fold increase in viable nucleated cells and high transduction efficiencies (64-92%). At the end of culture, functional assays demonstrated that these cells were potent and specific in their ability to kill tumor cells bearing target and secrete large quantities of interferon and tumor necrosis factor. Both phases of culture were contained within closed or semi-closed modules, which include automated density gradient separation and cell culture bags for the first phase and closed GREX culture devices and wash/concentrate systems for the second phase. Large-scale manufacturing using modular systems and semi-automated devices resulted in highly functional clinical-grade TCR transduced T-cells. This process is now in use in actively accruing clinical trials and the NIH Clinical Center and can be utilized at other cell therapy manufacturing sites that wish to scale-up and optimize their processing using closed systems.

Can the Accuracy of Home Blood Glucose Monitors be affected by the Received Signal Strength of 900 MHz GSM Mobile Phones?

PubMed Central

Eslami, J.; Ghafaripour, F.; Mortazavi, S.A.R.; Mortazavi, S.M.J.; Shojaei-fard, M.B.

2015-01-01

Background People who use home blood glucose monitors may use their mobile phones in the close vicinity of medical devices. This study is aimed at investigating the effect of the signal strength of 900 MHz GSM mobile phones on the accuracy of home blood glucose monitors. Methods Sixty non-diabetic volunteer individuals aged 21 - 28 years participated in this study. Blood samples were analyzed for glucose level by using a common blood glucose monitoring system. Each blood sample was analyzed twice, within ten minutes in presence and absence of electromagnetic fields generated by a common GSM mobile phone during ringing. Blood samples were divided into 3 groups of 20 samples each. Group 1: exposure to mobile phone radiation with weak signal strength. Group2: exposure to mobile phone radiation with strong signal strength. Group3: exposure to a switched–on mobile phone with no signal strength. Results The magnitude of the changes in the first, second and third group between glucose levels of two measurements (׀ΔC׀) were 7.4±3.9 mg/dl, 10.2±4.5 mg/dl, 8.7±8.4 mg/dl respectively. The difference in the magnitude of the changes between the 1st and the 3rd groups was not statistically significant. Furthermore, the difference in the magnitude of the changes between the 2nd and the 3rd groups was not statistically significant. Conclusion Findings of this study showed that the signal strength of 900 MHz GSM mobile phones cannot play a significant role in changing the accuracy of home blood glucose monitors. PMID:26688798

Platypus TCRμ provides insight into the origins and evolution of a uniquely mammalian TCR locus1

PubMed Central

Wang, Xinxin; Parra, Zuly E.; Miller, Robert D.

2011-01-01

TCRμ is an unconventional TCR that was first discovered in marsupials and appears to be absent from placental mammals and non-mammals. Here we show that TCRμ is also present in the duckbill platypus, an egg-laying monotreme, consistent with TCRμ being ancient and present in the last common ancestor of all extant mammals. As in marsupials, platypus TCRμ is expressed in a form containing double V domains. These V domains more closely resemble antibody V than that of conventional TCR. Platypus TCRμ differs from its marsupial homologue by requiring two rounds of somatic DNA recombination to assemble both V exons and has a genomic organization resembling the likely ancestral form of the receptor genes. These results demonstrate that the ancestors of placental mammals would have had TCRμ but it has been lost from this lineage. PMID:21976776

Signal Transduction in T Cell Activation and Tolerance

DTIC Science & Technology

1993-01-01

chains and ’ chains may transduce different signals in intact T cells. These studies demonstrate that while c- deficient and c-containing TCR complexes...three independently derived pairs of CD45- and CD45+ murine T cell lymphomas, the CD45- expressing cells were consistently deficient in...D.B., Larsen, A. and Wilson, C.B. (1986) Reduced interferon-gamma mRNA levels in human neonates: Evidence for an intrinsic T cell deficiency yi 114

Ablation of SLP-76 signaling after T cell priming generates memory CD4 T cells impaired in steady-state and cytokine-driven homeostasis.

PubMed

Bushar, Nicholas D; Corbo, Evann; Schmidt, Michelle; Maltzman, Jonathan S; Farber, Donna L

2010-01-12

The intracellular signaling mechanisms regulating the generation and long-term persistence of memory T cells in vivo remain unclear. In this study, we used mouse models with conditional deletion of the key T cell receptor (TCR)-coupled adaptor molecule SH2-domain-containing phosphoprotein of 76 kDa (SLP-76), to analyze signaling mechanisms for memory CD4 T cell generation, maintenance, and homeostasis. We found that ablation of SLP-76 expression after T cell priming did not inhibit generation of phenotypic effector or memory CD4 T cells; however, the resultant SLP-76-deficient memory CD4 T cells could not produce recall cytokines in response to TCR-mediated stimulation and showed decreased persistence in vivo. In addition, SLP-76-deficient memory CD4 T cells exhibited reduced steady-state homeostasis and were impaired in their ability to homeostatically expand in vivo in response to the gamma(c) cytokine IL-7, despite intact proximal signaling through the IL-7R-coupled JAK3/STAT5 pathway. Direct in vivo deletion of SLP-76 in polyclonal memory CD4 T cells likewise led to impaired steady-state homeostasis as well as impaired homeostatic responses to IL-7. Our findings demonstrate a dominant role for SLP-76-dependent TCR signals in regulating turnover and perpetuation of memory CD4 T cells and their responses to homeostatic cytokines, with implications for the selective survival of memory CD4 T cells following pathogen exposure, vaccination, and aging.

Activation of Extracellular Signal-Regulated Kinase but Not of p38 Mitogen-Activated Protein Kinase Pathways in Lymphocytes Requires Allosteric Activation of SOS

PubMed Central

Jun, Jesse E.; Yang, Ming; Chen, Hang; Chakraborty, Arup K.

2013-01-01

Thymocytes convert graded T cell receptor (TCR) signals into positive selection or deletion, and activation of extracellular signal-related kinase (ERK), p38, and Jun N-terminal protein kinase (JNK) mitogen-activated protein kinases (MAPKs) has been postulated to play a discriminatory role. Two families of Ras guanine nucleotide exchange factors (RasGEFs), SOS and RasGRP, activate Ras and the downstream RAF-MEK-ERK pathway. The pathways leading to lymphocyte p38 and JNK activation are less well defined. We previously described how RasGRP alone induces analog Ras-ERK activation while SOS and RasGRP cooperate to establish bimodal ERK activation. Here we employed computational modeling and biochemical experiments with model cell lines and thymocytes to show that TCR-induced ERK activation grows exponentially in thymocytes and that a W729E allosteric pocket mutant, SOS1, can only reconstitute analog ERK signaling. In agreement with RasGRP allosterically priming SOS, exponential ERK activation is severely decreased by pharmacological or genetic perturbation of the phospholipase Cγ (PLCγ)-diacylglycerol-RasGRP1 pathway. In contrast, p38 activation is not sharply thresholded and requires high-level TCR signal input. Rac and p38 activation depends on SOS1 expression but not allosteric activation. Based on computational predictions and experiments exploring whether SOS functions as a RacGEF or adaptor in Rac-p38 activation, we established that the presence of SOS1, but not its enzymatic activity, is critical for p38 activation. PMID:23589333

Received Signal Strength Database Interpolation by Kriging for a Wi-Fi Indoor Positioning System.

PubMed

Jan, Shau-Shiun; Yeh, Shuo-Ju; Liu, Ya-Wen

2015-08-28

The main approach for a Wi-Fi indoor positioning system is based on the received signal strength (RSS) measurements, and the fingerprinting method is utilized to determine the user position by matching the RSS values with the pre-surveyed RSS database. To build a RSS fingerprint database is essential for an RSS based indoor positioning system, and building such a RSS fingerprint database requires lots of time and effort. As the range of the indoor environment becomes larger, labor is increased. To provide better indoor positioning services and to reduce the labor required for the establishment of the positioning system at the same time, an indoor positioning system with an appropriate spatial interpolation method is needed. In addition, the advantage of the RSS approach is that the signal strength decays as the transmission distance increases, and this signal propagation characteristic is applied to an interpolated database with the Kriging algorithm in this paper. Using the distribution of reference points (RPs) at measured points, the signal propagation model of the Wi-Fi access point (AP) in the building can be built and expressed as a function. The function, as the spatial structure of the environment, can create the RSS database quickly in different indoor environments. Thus, in this paper, a Wi-Fi indoor positioning system based on the Kriging fingerprinting method is developed. As shown in the experiment results, with a 72.2% probability, the error of the extended RSS database with Kriging is less than 3 dBm compared to the surveyed RSS database. Importantly, the positioning error of the developed Wi-Fi indoor positioning system with Kriging is reduced by 17.9% in average than that without Kriging.

The phosphatase JKAP/DUSP22 inhibits t-cell receptor signalling and autoimmunity by inactivating Lck

USDA-ARS?s Scientific Manuscript database

JNK pathway-associated phosphatase (JKAP, also known as DUSP22 or JSP-1) is a JNK activator. The in vivo role of JKAP in immune regulation remains unclear. Here we report that JKAP directly inactivates Lck by dephosphorylating tyrosine-394 residue during T-cell receptor (TCR) signalling. JKAP-knocko...

Retinoic Acid Modulates Interferon-γ Production by Hepatic Natural Killer T Cells via Phosphatase 2A and the Extracellular Signal-Regulated Kinase Pathway

PubMed Central

Chang, Heng-Kwei

2015-01-01

Retinoic acid (RA), an active metabolite converted from vitamin A, plays an active role in immune function, such as defending against infections and immune regulation. Although RA affects various types of immune cells, including antigen-presenting cells, B lymphocytes, and T lymphocytes, whether it affects natural killer T (NKT) cells remain unknown. In this study, we found that RA decreased interferon (IFN)-γ production by activated NKT cells through T-cell receptor (TCR) and CD28. We also found that RA reduced extracellular signal-regulated kinase (ERK) phosphorylation, but increased phosphatase 2A (PP2A) activity in TCR/CD28-stimulated NKT cells. The increased PP2A activity, at least partly, contributed to the reduction of ERK phosphorylation. Since inhibition of ERK activation decreases IFN-γ production by TCR/CD28-stimulated NKT cells, RA may downregulate IFN-γ production by TCR/CD28-stimulated NKT cells through the PP2A-ERK pathway. Our results demonstrated a novel function of RA in modulating the IFN-γ expression by activated NKT cells. PMID:25343668

Lack of T-cell receptor-induced signaling is crucial for CD95 ligand up-regulation and protects cutaneous T-cell lymphoma cells from activation-induced cell death.

PubMed

Klemke, Claus-Detlev; Brenner, Dirk; Weiss, Eva-Maria; Schmidt, Marc; Leverkus, Martin; Gülow, Karsten; Krammer, Peter H

2009-05-15

Restimulation of previously activated T cells via the T-cell receptor (TCR) leads to activation-induced cell death (AICD), which is, at least in part, dependent on the death receptor CD95 (APO-1, FAS) and its natural ligand (CD95L). Here, we characterize cutaneous T-cell lymphoma (CTCL) cells (CTCL tumor cell lines and primary CTCL tumor cells from CTCL patients) as AICD resistant. We show that CTCL cells have elevated levels of the CD95-inhibitory protein cFLIP. However, cFLIP is not responsible for CTCL AICD resistance. Instead, our data suggest that reduced TCR-proximal signaling in CTCL cells is responsible for the observed AICD resistance. CTCL cells exhibit no PLC-gamma1 activity, resulting in an impaired Ca(2+)release and reduced generation of reactive oxygen species upon TCR stimulation. Ca(2+) and ROS production are crucial for up-regulation of CD95L and reconstitution of both signals resulted in AICD sensitivity of CTCL cells. In accordance with these data, CTCL tumor cells from patients with Sézary syndrome do not up-regulate CD95L upon TCR-stimulation and are therefore resistant to AICD. These results show a novel mechanism of AICD resistance in CTCL that could have future therapeutic implications to overcome apoptosis resistance in CTCL patients.

Accelerated Loss of TCR Repertoire Diversity in Common Variable Immunodeficiency

PubMed Central

Wong, Gabriel K.; Millar, David; Penny, Sarah; Heather, James M.; Mistry, Punam; Buettner, Nico; Bryon, Jane; Huissoon, Aarnoud P.

2016-01-01

Although common variable immunodeficiency (CVID) has long been considered as a group of primary Ab deficiencies, growing experimental data now suggest a global disruption of the entire adaptive immune response in a segment of patients. Oligoclonality of the TCR repertoire was previously demonstrated; however, the manner in which it relates to other B cell and T cell findings reported in CVID remains unclear. Using a combination approach of high-throughput TCRβ sequencing and multiparametric flow cytometry, we compared the TCR repertoire diversity between various subgroups of CVID patients according to their B cell immunophenotypes. Our data suggest that the reduction in repertoire diversity is predominantly restricted to those patients with severely reduced class-switched memory B cells and an elevated level of CD21lo B cells (Freiburg 1a), and may be driven by a reduced number of naive T cells unmasking underlying memory clonality. Moreover, our data indicate that this loss in repertoire diversity progresses with advancing age far exceeding the expected physiological rate. Radiological evidence supports the loss in thymic volume, correlating with the decrease in repertoire diversity. Evidence now suggests that primary thymic failure along with other well-described B cell abnormalities play an important role in the pathophysiology in Freiburg group 1a patients. Clinically, our findings emphasize the integration of combined B and T cell testing to identify those patients at the greatest risk for infection. Future work should focus on investigating the link between thymic failure and the severe reduction in class-switched memory B cells, while gathering longitudinal laboratory data to examine the progressive nature of the disease. PMID:27481850

Received Signal Strength Database Interpolation by Kriging for a Wi-Fi Indoor Positioning System

PubMed Central

Jan, Shau-Shiun; Yeh, Shuo-Ju; Liu, Ya-Wen

2015-01-01

The main approach for a Wi-Fi indoor positioning system is based on the received signal strength (RSS) measurements, and the fingerprinting method is utilized to determine the user position by matching the RSS values with the pre-surveyed RSS database. To build a RSS fingerprint database is essential for an RSS based indoor positioning system, and building such a RSS fingerprint database requires lots of time and effort. As the range of the indoor environment becomes larger, labor is increased. To provide better indoor positioning services and to reduce the labor required for the establishment of the positioning system at the same time, an indoor positioning system with an appropriate spatial interpolation method is needed. In addition, the advantage of the RSS approach is that the signal strength decays as the transmission distance increases, and this signal propagation characteristic is applied to an interpolated database with the Kriging algorithm in this paper. Using the distribution of reference points (RPs) at measured points, the signal propagation model of the Wi-Fi access point (AP) in the building can be built and expressed as a function. The function, as the spatial structure of the environment, can create the RSS database quickly in different indoor environments. Thus, in this paper, a Wi-Fi indoor positioning system based on the Kriging fingerprinting method is developed. As shown in the experiment results, with a 72.2% probability, the error of the extended RSS database with Kriging is less than 3 dBm compared to the surveyed RSS database. Importantly, the positioning error of the developed Wi-Fi indoor positioning system with Kriging is reduced by 17.9% in average than that without Kriging. PMID:26343673

Improved statistical signal detection in pharmacovigilance by combining multiple strength-of-evidence aspects in vigiRank.

PubMed

Caster, Ola; Juhlin, Kristina; Watson, Sarah; Norén, G Niklas

2014-08-01

Detection of unknown risks with marketed medicines is key to securing the optimal care of individual patients and to reducing the societal burden from adverse drug reactions. Large collections of individual case reports remain the primary source of information and require effective analytics to guide clinical assessors towards likely drug safety signals. Disproportionality analysis is based solely on aggregate numbers of reports and naively disregards report quality and content. However, these latter features are the very fundament of the ensuing clinical assessment. Our objective was to develop and evaluate a data-driven screening algorithm for emerging drug safety signals that accounts for report quality and content. vigiRank is a predictive model for emerging safety signals, here implemented with shrinkage logistic regression to identify predictive variables and estimate their respective contributions. The variables considered for inclusion capture different aspects of strength of evidence, including quality and clinical content of individual reports, as well as trends in time and geographic spread. A reference set of 264 positive controls (historical safety signals from 2003 to 2007) and 5,280 negative controls (pairs of drugs and adverse events not listed in the Summary of Product Characteristics of that drug in 2012) was used for model fitting and evaluation; the latter used fivefold cross-validation to protect against over-fitting. All analyses were performed on a reconstructed version of VigiBase(®) as of 31 December 2004, at around which time most safety signals in our reference set were emerging. The following aspects of strength of evidence were selected for inclusion into vigiRank: the numbers of informative and recent reports, respectively; disproportional reporting; the number of reports with free-text descriptions of the case; and the geographic spread of reporting. vigiRank offered a statistically significant improvement in area under the receiver

WC1 is a hybrid γδ TCR coreceptor and pattern recognition receptor for pathogenic bacteria.

PubMed

Hsu, Haoting; Chen, Chuang; Nenninger, Ariel; Holz, Lauren; Baldwin, Cynthia L; Telfer, Janice C

2015-03-01

WC1 proteins are uniquely expressed on γδ T cells and belong to the scavenger receptor cysteine-rich (SRCR) superfamily. While present in variable, and sometimes high, numbers in the genomes of mammals and birds, in cattle there are 13 distinct genes (WC1-1 to WC1-13). All bovine WC1 proteins can serve as coreceptors for the TCR in a tyrosine phosphorylation dependent manner, and some are required for the γδ T cell response to Leptospira. We hypothesized that individual WC1 receptors encode Ag specificity via coligation of bacteria with the γδ TCR. SRCR domain binding was directly correlated with γδ T cell response, as WC1-3 SRCR domains from Leptospira-responsive cells, but not WC1-4 SRCR domains from Leptospira-nonresponsive cells, bound to multiple serovars of two Leptospira species, L. borgpetersenii, and L. interrogans. Three to five of eleven WC1-3 SRCR domains, but none of the eleven WC1-4 SRCR domains, interacted with Leptospira spp. and Borrelia burgdorferi, but not with Escherichia coli or Staphylococcus aureus. Mutational analysis indicated that the active site for bacterial binding in one of the SRCR domains is composed of amino acids in three discontinuous regions. Recombinant WC1 SRCR domains with the ability to bind leptospires inhibited Leptospira growth. Our data suggest that WC1 gene arrays play a multifaceted role in the γδ T cell response to bacteria, including acting as hybrid pattern recognition receptors and TCR coreceptors, and they may function as antimicrobials. Copyright © 2015 by The American Association of Immunologists, Inc.

Indoor Location Sensing with Invariant Wi-Fi Received Signal Strength Fingerprinting.

PubMed

Husen, Mohd Nizam; Lee, Sukhan

2016-11-11

A method of location fingerprinting based on the Wi-Fi received signal strength (RSS) in an indoor environment is presented. The method aims to overcome the RSS instability due to varying channel disturbances in time by introducing the concept of invariant RSS statistics. The invariant RSS statistics represent here the RSS distributions collected at individual calibration locations under minimal random spatiotemporal disturbances in time. The invariant RSS statistics thus collected serve as the reference pattern classes for fingerprinting. Fingerprinting is carried out at an unknown location by identifying the reference pattern class that maximally supports the spontaneous RSS sensed from individual Wi-Fi sources. A design guideline is also presented as a rule of thumb for estimating the number of Wi-Fi signal sources required to be available for any given number of calibration locations under a certain level of random spatiotemporal disturbances. Experimental results show that the proposed method not only provides 17% higher success rate than conventional ones but also removes the need for recalibration. Furthermore, the resolution is shown finer by 40% with the execution time more than an order of magnitude faster than the conventional methods. These results are also backed up by theoretical analysis.

Shank3 Is Part of a Zinc-Sensitive Signaling System That Regulates Excitatory Synaptic Strength

PubMed Central

Arons, Magali H.; Lee, Kevin; Thynne, Charlotte J.; Kim, Sally A.; Schob, Claudia; Kindler, Stefan

2016-01-01

Shank3 is a multidomain scaffold protein localized to the postsynaptic density of excitatory synapses. Functional studies in vivo and in vitro support the concept that Shank3 is critical for synaptic plasticity and the trans-synaptic coupling between the reliability of presynaptic neurotransmitter release and postsynaptic responsiveness. However, how Shank3 regulates synaptic strength remains unclear. The C terminus of Shank3 contains a sterile alpha motif (SAM) domain that is essential for its postsynaptic localization and also binds zinc, thus raising the possibility that changing zinc levels modulate Shank3 function in dendritic spines. In support of this hypothesis, we find that zinc is a potent regulator of Shank3 activation and dynamics in rat hippocampal neurons. Moreover, we show that zinc modulation of synaptic transmission is Shank3 dependent. Interestingly, an autism spectrum disorder (ASD)-associated variant of Shank3 (Shank3R87C) retains its zinc sensitivity and supports zinc-dependent activation of AMPAR-mediated synaptic transmission. However, elevated zinc was unable to rescue defects in trans-synaptic signaling caused by the R87C mutation, implying that trans-synaptic increases in neurotransmitter release are not necessary for the postsynaptic effects of zinc. Together, these data suggest that Shank3 is a key component of a zinc-sensitive signaling system, regulating synaptic strength that may be impaired in ASD. SIGNIFICANCE STATEMENT Shank3 is a postsynaptic protein associated with neurodevelopmental disorders such as autism and schizophrenia. In this study, we show that Shank3 is a key component of a zinc-sensitive signaling system that regulates excitatory synaptic transmission. Intriguingly, an autism-associated mutation in Shank3 partially impairs this signaling system. Therefore, perturbation of zinc homeostasis may impair, not only synaptic functionality and plasticity, but also may lead to cognitive and behavioral abnormalities seen in

Activation of human naïve Th cells increases surface expression of GD3 and induces neoexpression of GD2 that colocalize with TCR clusters.

PubMed

Villanueva-Cabello, Tania M; Mollicone, Rosella; Cruz-Muñoz, Mario E; López-Guerrero, Delia V; Martínez-Duncker, Iván

2015-12-01

CD4+ T helper lymphocytes (Th) orchestrate the immune response after their activation by antigen-presenting cells. Activation of naïve Th cells is reported to generate the reduction in surface epitopes of sialic acid (Sia) in α2,3 and α2,6 linkages. In this work, we report that in spite of this glycophenotype, anti-CD3/anti-CD28-activated purified human naïve Th cells show a significant increase in surface Sia, as assessed by metabolic labeling, compared with resting naïve Th cells, suggesting an increased flux of Sia toward Siaα2,8 glycoconjugates. To understand this increase as a result of ganglioside up-regulation, we observed that very early after activation, human naïve Th cells show an increased expression in surface GD3 and neoexpression of surface GD2 gangliosides, the latter clustering with the T cell receptor (TCR). Also, we report that in contrast to GM2/GD2 synthase null mice, lentiviral vector-mediated silencing of the GM2/GD2 synthase in activated human naïve Th cells reduced efficient TCR clustering and downstream signaling, as assessed by proliferation assays and IL-2 and IL-2R expression, pointing to an important role of this enzyme in activation of human naive Th cells. © The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Signal Amplification in Field Effect-Based Sandwich Enzyme-Linked Immunosensing by Tuned Buffer Concentration with Ionic Strength Adjuster.

PubMed

Kumar, Satyendra; Kumar, Narendra; Panda, Siddhartha

2016-04-01

Miniaturization of the sandwich enzyme-based immunosensor has several advantages but could result in lower signal strength due to lower enzyme loading. Hence, technologies for amplification of the signal are needed. Signal amplification in a field effect-based electrochemical immunosensor utilizing chip-based ELISA is presented in this work. First, the molarities of phosphate buffer saline (PBS) and concentrations of KCl as ionic strength adjuster were optimized to maximize the GOx glucose-based enzymatic reactions in a beaker for signal amplification measured by change in the voltage shift with an EIS device (using 20 μl of solution) and validated with a commercial pH meter (using 3 ml of solution). The PBS molarity of 100 μM with 25 mM KCl provided the maximum voltage shift. These optimized buffer conditions were further verified for GOx immobilized on silicon chips, and similar trends with decreased PBS molarity were obtained; however, the voltage shift values obtained on chip reaction were lower as compared to the reactions occurring in the beaker. The decreased voltage shift with immobilized enzyme on chip could be attributed to the increased Km (Michaelis-Menten constant) values in the immobilized GOx. Finally, a more than sixfold signal enhancement (from 8 to 47 mV) for the chip-based sandwich immunoassay was obtained by altering the PBS molarity from 10 to 100 μM with 25 mM KCl.

Extraordinary TCR in Carbon Nanotube-Polymer Composites and Device Implications in Bolometric Infrared Detection

DTIC Science & Technology

2015-03-24

were made via filtration of 1 mL of 1 mg/L SWCNT solution (Nanointegris Inc.) through a 20 nm pore anodized aluminum oxide ( AAO ) membrane filter to...prepared by attaching a rectangular piece cut from the AAO membrane filter onto a glass substrate. Contacts were made with silver paste to the extremities...NUMBER(S) 14. ABSTRACT _ The development of high TCR materials, such as vanadium oxide (VOx), has enabled the introduction of bolometric infrared

The quantum chemical causality of pMHC-TCR biological avidity: Peptide atomic coordination data and the electronic state of agonist N termini.

PubMed

Antipas, Georgios S E; Germenis, Anastasios E

2015-06-01

The quantum state of functional avidity of the synapse formed between a peptide-Major Histocompatibility Complex (pMHC) and a T cell receptor (TCR) is a subject not previously touched upon. Here we present atomic pair correlation meta-data based on crystalized tertiary structures of the Tax (HTLV-1) peptide along with three artificially altered variants, all of which were presented by the (Class I) HLA-A201 protein in complexation with the human (CD8(+)) A6TCR. The meta-data reveal the existence of a direct relationship between pMHC-TCR functional avidity (agonist/antagonist) and peptide pair distribution function (PDF). In this context, antagonist peptides are consistently under-coordinated in respect to Tax. Moreover, Density Functional Theory (DFT) datasets in the BLYP/TZ2P level of theory resulting from relaxation of the H species on peptide tertiary structures reveal that the coordination requirement of agonist peptides is also expressed as a physical observable of the protonation state of their N termini: agonistic peptides are always found to retain a stable ammonium (NH3 (+)) terminal group while antagonist peptides are not.

Interplay between glucose and leptin signaling determines the strength of GABAergic synapses at POMC neurons

PubMed Central

Lee, Dong Kun; Jeong, Jae Hoon; Chun, Sung-Kun; Chua, Streamson; Jo, Young-Hwan

2015-01-01

Regulation of GABAergic inhibitory inputs and alterations in POMC neuron activity by nutrients and adiposity signals regulate energy and glucose homeostasis. Thus, understanding how POMC neurons integrate these two signal molecules at the synaptic level is important. Here we show that leptin’s action on GABA release to POMC neurons is influenced by glucose levels. Leptin stimulates the JAK2-PI3K pathway in both presynaptic GABAergic terminals and postsynaptic POMC neurons. Inhibition of AMPK activity in presynaptic terminals decreases GABA release at 10 mM glucose. However, postsynaptic TRPC channel opening by the PI3K-PLC signaling pathway in POMC neurons enhances spontaneous GABA release via activation of presynaptic MC3/4 and mGlu receptors at 2.5 mM glucose. High-fat feeding blunts AMPK-dependent presynaptic inhibition, whereas PLC-mediated GABAergic feedback inhibition remains responsive to leptin. Our data indicate that the interplay between glucose and leptin signaling in glutamatergic POMC neurons is critical for determining the strength of inhibitory tone towards POMC neurons. PMID:25808323

Interplay between glucose and leptin signalling determines the strength of GABAergic synapses at POMC neurons.

PubMed

Lee, Dong Kun; Jeong, Jae Hoon; Chun, Sung-Kun; Chua, Streamson; Jo, Young-Hwan

2015-03-26

Regulation of GABAergic inhibitory inputs and alterations in POMC neuron activity by nutrients and adiposity signals regulate energy and glucose homeostasis. Thus, understanding how POMC neurons integrate these two signal molecules at the synaptic level is important. Here we show that leptin's action on GABA release to POMC neurons is influenced by glucose levels. Leptin stimulates the JAK2-PI3K pathway in both presynaptic GABAergic terminals and postsynaptic POMC neurons. Inhibition of AMPK activity in presynaptic terminals decreases GABA release at 10 mM glucose. However, postsynaptic TRPC channel opening by the PI3K-PLC signalling pathway in POMC neurons enhances spontaneous GABA release via activation of presynaptic MC3/4 and mGlu receptors at 2.5 mM glucose. High-fat feeding blunts AMPK-dependent presynaptic inhibition, whereas PLC-mediated GABAergic feedback inhibition remains responsive to leptin. Our data indicate that the interplay between glucose and leptin signalling in glutamatergic POMC neurons is critical for determining the strength of inhibitory tone towards POMC neurons.

The Us3 Protein of Herpes Simplex Virus 1 Inhibits T Cell Signaling by Confining Linker for Activation of T Cells (LAT) Activation via TRAF6 Protein*

PubMed Central

Yang, Yin; Wu, Songfang; Wang, Yu; Pan, Shuang; Lan, Bei; Liu, Yaohui; Zhang, Liming; Leng, Qianli; Chen, Da; Zhang, Cuizhu; He, Bin; Cao, Youjia

2015-01-01

Herpes simplex virus 1 (HSV-1) is the most prevalent human virus and causes global morbidity because the virus is able to infect multiple cell types. Remarkably, HSV infection switches between lytic and latent cycles, where T cells play a critical role. However, the precise way of virus-host interactions is incompletely understood. Here we report that HSV-1 productively infected Jurkat T-cells and inhibited antigen-induced T cell receptor activation. We discovered that HSV-1-encoded Us3 protein interrupted TCR signaling and interleukin-2 production by inactivation of the linker for activation of T cells. This study unveils a mechanism by which HSV-1 intrudes into early events of TCR-mediated cell signaling and may provide novel insights into HSV infection, during which the virus escapes from host immune surveillance. PMID:25907557

Site preference, magnetism and lattice vibrations of intermetallics Lu₂Fe 17–xT x (T=Cr, Mn, Ru)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Jin-Chun; Qian, Ping, E-mail: qianpinghu@sohu.com; Zhang, Zhen-Feng

We present an atomistic study on the phase stability, site preference and lattice constants of the rare earth intermetallics Lu₂Fe 17–xT x (T=Cr, Mn, Ru). The calculated preferential occupation site of ternary element T is found to be the 4f site. The order of site preference is given as 4f, 12k, 12j and 6g for Lu₂Fe 17–xT x. The calculated lattice parameters are corresponding to the experimental results. We have calculated the magnetic moments of Lu₂Fe 17–xT x compounds. Results show that the calculated total magnetic moment of Lu₂Fe₁₇ compound is M=37.34 μ B/f.u. In addition, the total and partialmore » phonon densities of states are evaluated first for these complicated structures. - Graphical abstract: The vibrational modes are mostly excited by Fe atoms, Lu contributes to the lower frequencies modes, and the contribution of Ru atoms is the same as Fe atoms. Highlights: • There are no reports on lattice vibrations of Lu₂(Fe, T) 17–x (T=Cr, Mn, Ru) compounds. • The phase stability and site preference are evaluated first for the complex structures of Lu₂(Fe, T) 17–x (T=Cr, Mn, Ru) compounds. • The lattice inversion method to obtain the interatomic pair potential is the unique one.« less

CDR3 analysis of TCR Vβ repertoire of CD8⁺ T cells from chickens infected with Eimeria maxima.

PubMed

Ren, Chao; Yin, Guangwen; Qin, Mei; Suo, Jingxia; Lv, Qiyao; Xie, Li; Wang, Yunzhou; Huang, Xiaoxi; Chen, Yuchen; Liu, Xianyong; Suo, Xun

2014-08-01

CD8(+) T cells play a major role in the immune protection of host against the reinfection of Eimeria maxima, the most immunogenic species of eimerian parasites in chickens. To explore the dominant complementarity-determining regions 3 (CDR3) of CD8(+) T cell populations induced by the infection of this parasite, sequence analysis was performed in this study for CDR3 of CD8(+) T cells from E. maxima infected chickens. After 5 days post the third or forth infection, intraepithelial lymphocytes were isolated from the jejunum of bird. CD3(+)CD8(+) T cells were sorted and subjected to total RNA isolation and cDNA preparation. PCR amplification and cloning of the loci between Vβ1 and Cβ was conducted for the subsequent sequencing of CDR3 of T cell receptor (TCR). After the forth infection, 2 birds exhibited two same frequent TCR CDR3 sequences, i.e., AKQDWGTGGYSNMI and AGRVLNIQY; while the third bird showed two different frequent TCR CDR3 sequences, AKQGARGHTPLN and AKQDIEVRGPNTPLN. No frequent CDR3 sequence was detected from uninfected birds, though AGRVLNIQY was also found in two uninfected birds. Our result preliminarily demonstrates that frequent CDR3 sequences may exist in E. maxima immunized chickens, encouraging the mining of the immunodominant CD8(+) T cells against E. maxima infection. Copyright © 2014 Elsevier Inc. All rights reserved.

RasGRP1 regulates antigen-induced developmental programming by naive CD8 T cells.

PubMed

Priatel, John J; Chen, Xiaoxi; Huang, Yu-Hsuan; Chow, Michael T; Zenewicz, Lauren A; Coughlin, Jason J; Shen, Hao; Stone, James C; Tan, Rusung; Teh, Hung Sia

2010-01-15

Ag encounter by naive CD8 T cells initiates a developmental program consisting of cellular proliferation, changes in gene expression, and the formation of effector and memory T cells. The strength and duration of TCR signaling are known to be important parameters regulating the differentiation of naive CD8 T cells, although the molecular signals arbitrating these processes remain poorly defined. The Ras-guanyl nucleotide exchange factor RasGRP1 has been shown to transduce TCR-mediated signals critically required for the maturation of developing thymocytes. To elucidate the role of RasGRP1 in CD8 T cell differentiation, in vitro and in vivo experiments were performed with 2C TCR transgenic CD8 T cells lacking RasGRP1. In this study, we report that RasGRP1 regulates the threshold of T cell activation and Ag-induced expansion, at least in part, through the regulation of IL-2 production. Moreover, RasGRP1(-/-) 2C CD8 T cells exhibit an anergic phenotype in response to cognate Ag stimulation that is partially reversible upon the addition of exogenous IL-2. By contrast, the capacity of IL-2/IL-2R interactions to mediate Ras activation and CD8 T cell expansion and differentiation appears to be largely RasGRP1-independent. Collectively, our results demonstrate that RasGRP1 plays a selective role in T cell signaling, controlling the initiation and duration of CD8 T cell immune responses.

Na+ influx via Orai1 inhibits intracellular ATP-induced mTORC2 signaling to disrupt CD4 T cell gene expression and differentiation.

PubMed

Miao, Yong; Bhushan, Jaya; Dani, Adish; Vig, Monika

2017-05-11

T cell effector functions require sustained calcium influx. However, the signaling and phenotypic consequences of non-specific sodium permeation via calcium channels remain unknown. α-SNAP is a crucial component of Orai1 channels, and its depletion disrupts the functional assembly of Orai1 multimers. Here we show that α-SNAP hypomorph, hydrocephalus with hopping gait, Napa hyh/hyh mice harbor significant defects in CD4 T cell gene expression and Foxp3 regulatory T cell (Treg) differentiation. Mechanistically, TCR stimulation induced rapid sodium influx in Napa hyh/hyh CD4 T cells, which reduced intracellular ATP, [ATP] i . Depletion of [ATP] i inhibited mTORC2 dependent NFκB activation in Napa hyh/hyh cells but ablation of Orai1 restored it. Remarkably, TCR stimulation in the presence of monensin phenocopied the defects in Napa hyh/hyh signaling and Treg differentiation, but not IL-2 expression. Thus, non-specific sodium influx via bonafide calcium channels disrupts unexpected signaling nodes and may provide mechanistic insights into some divergent phenotypes associated with Orai1 function.

Na+ influx via Orai1 inhibits intracellular ATP-induced mTORC2 signaling to disrupt CD4 T cell gene expression and differentiation

PubMed Central

Miao, Yong; Bhushan, Jaya; Dani, Adish; Vig, Monika

2017-01-01

T cell effector functions require sustained calcium influx. However, the signaling and phenotypic consequences of non-specific sodium permeation via calcium channels remain unknown. α-SNAP is a crucial component of Orai1 channels, and its depletion disrupts the functional assembly of Orai1 multimers. Here we show that α-SNAP hypomorph, hydrocephalus with hopping gait, Napahyh/hyh mice harbor significant defects in CD4 T cell gene expression and Foxp3 regulatory T cell (Treg) differentiation. Mechanistically, TCR stimulation induced rapid sodium influx in Napahyh/hyh CD4 T cells, which reduced intracellular ATP, [ATP]i. Depletion of [ATP]i inhibited mTORC2 dependent NFκB activation in Napahyh/hyh cells but ablation of Orai1 restored it. Remarkably, TCR stimulation in the presence of monensin phenocopied the defects in Napahyh/hyh signaling and Treg differentiation, but not IL-2 expression. Thus, non-specific sodium influx via bonafide calcium channels disrupts unexpected signaling nodes and may provide mechanistic insights into some divergent phenotypes associated with Orai1 function. DOI: http://dx.doi.org/10.7554/eLife.25155.001 PMID:28492364

Yellow Fever Virus, but Not Zika Virus or Dengue Virus, Inhibits T-Cell Receptor-Mediated T-Cell Function by an RNA-Based Mechanism.

PubMed

McLinden, James H; Bhattarai, Nirjal; Stapleton, Jack T; Chang, Qing; Kaufman, Thomas M; Cassel, Suzanne L; Sutterwala, Fayyaz S; Haim, Hillel; Houtman, Jon C; Xiang, Jinhua

2017-11-27

The Flavivirus genus within the Flaviviridae family is comprised of many important human pathogens including yellow fever virus (YFV), dengue virus (DENV), and Zika virus (ZKV), all of which are global public health concerns. Although the related flaviviruses hepatitis C virus and human pegivirus (formerly named GBV-C) interfere with T-cell receptor (TCR) signaling by novel RNA and protein-based mechanisms, the effect of other flaviviruses on TCR signaling is unknown. Here, we studied the effect of YFV, DENV, and ZKV on TCR signaling. Both YFV and ZKV replicated in human T cells in vitro; however, only YFV inhibited TCR signaling. This effect was mediated at least in part by the YFV envelope (env) protein coding RNA. Deletion mutagenesis studies demonstrated that expression of a short, YFV env RNA motif (vsRNA) was required and sufficient to inhibit TCR signaling. Expression of this vsRNA and YFV infection of T cells reduced the expression of a Src-kinase regulatory phosphatase (PTPRE), while ZKV infection did not. YFV infection in mice resulted in impaired TCR signaling and PTPRE expression, with associated reduction in murine response to experimental ovalbumin vaccination. Together, these data suggest that viruses within the flavivirus genus inhibit TCR signaling in a species-dependent manner. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Indoor Location Sensing with Invariant Wi-Fi Received Signal Strength Fingerprinting

PubMed Central

Husen, Mohd Nizam; Lee, Sukhan

2016-01-01

A method of location fingerprinting based on the Wi-Fi received signal strength (RSS) in an indoor environment is presented. The method aims to overcome the RSS instability due to varying channel disturbances in time by introducing the concept of invariant RSS statistics. The invariant RSS statistics represent here the RSS distributions collected at individual calibration locations under minimal random spatiotemporal disturbances in time. The invariant RSS statistics thus collected serve as the reference pattern classes for fingerprinting. Fingerprinting is carried out at an unknown location by identifying the reference pattern class that maximally supports the spontaneous RSS sensed from individual Wi-Fi sources. A design guideline is also presented as a rule of thumb for estimating the number of Wi-Fi signal sources required to be available for any given number of calibration locations under a certain level of random spatiotemporal disturbances. Experimental results show that the proposed method not only provides 17% higher success rate than conventional ones but also removes the need for recalibration. Furthermore, the resolution is shown finer by 40% with the execution time more than an order of magnitude faster than the conventional methods. These results are also backed up by theoretical analysis. PMID:27845711

Secondary anchor polymorphism in the HA-1 minor histocompatibility antigen critically affects MHC stability and TCR recognition

PubMed Central

Nicholls, Sarah; Piper, Karen P.; Mohammed, Fiyaz; Dafforn, Timothy R.; Tenzer, Stefan; Salim, Mahboob; Mahendra, Premini; Craddock, Charles; van Endert, Peter; Schild, Hansjörg; Cobbold, Mark; Engelhard, Victor H.; Moss, Paul A. H.; Willcox, Benjamin E.

2009-01-01

T cell recognition of minor histocompatibility antigens (mHags) underlies allogeneic immune responses that mediate graft-versus-host disease and the graft-versus-leukemia effect following stem cell transplantation. Many mHags derive from single amino acid polymorphisms in MHC-restricted epitopes, but our understanding of the molecular mechanisms governing mHag immunogenicity and recognition is incomplete. Here we examined antigenic presentation and T-cell recognition of HA-1, a prototypic autosomal mHag derived from single nucleotide dimorphism (HA-1H versus HA-1R) in the HMHA1 gene. The HA-1H peptide is restricted by HLA-A2 and is immunogenic in HA-1R/R into HA-1H transplants, while HA-1R has been suggested to be a “null allele” in terms of T cell reactivity. We found that proteasomal cleavage and TAP transport of the 2 peptides is similar and that both variants can bind to MHC. However, the His>Arg change substantially decreases the stability and affinity of HLA-A2 association, consistent with the reduced immunogenicity of the HA-1R variant. To understand these findings, we determined the structure of an HLA-A2-HA-1H complex to 1.3Å resolution. Whereas His-3 is accommodated comfortably in the D pocket, incorporation of the lengthy Arg-3 is predicted to require local conformational changes. Moreover, a soluble TCR generated from HA-1H-specific T-cells bound HA-1H peptide with moderate affinity but failed to bind HA-1R, indicating complete discrimination of HA-1 variants at the level of TCR/MHC interaction. Our results define the molecular mechanisms governing immunogenicity of HA-1, and highlight how single amino acid polymorphisms in mHags can critically affect both MHC association and TCR recognition. PMID:19234124

WASp Family Verprolin-homologous Protein-2 (WAVE2) and Wiskott-Aldrich Syndrome Protein (WASp) Engage in Distinct Downstream Signaling Interactions at the T Cell Antigen Receptor Site*

PubMed Central

Pauker, Maor H.; Reicher, Barak; Joseph, Noah; Wortzel, Inbal; Jakubowicz, Shlomi; Noy, Elad; Perl, Orly; Barda-Saad, Mira

2014-01-01

T cell antigen receptor (TCR) engagement has been shown to activate pathways leading to actin cytoskeletal polymerization and reorganization, which are essential for lymphocyte activation and function. Several actin regulatory proteins were implicated in regulating the actin machinery, such as members of the Wiskott-Aldrich syndrome protein (WASp) family. These include WASp and the WASp family verprolin-homologous protein-2 (WAVE2). Although WASp and WAVE2 share several structural features, the precise regulatory mechanisms and potential redundancy between them have not been fully characterized. Specifically, unlike WASp, the dynamic molecular interactions that regulate WAVE2 recruitment to the cell membrane and specifically to the TCR signaling complex are largely unknown. Here, we identify the molecular mechanism that controls the recruitment of WAVE2 in comparison with WASp. Using fluorescence resonance energy transfer (FRET) and novel triple-color FRET (3FRET) technology, we demonstrate how WAVE2 signaling complexes are dynamically regulated during lymphocyte activation in vivo. We show that, similar to WASp, WAVE2 recruitment to the TCR site depends on protein-tyrosine kinase, ZAP-70, and the adaptors LAT, SLP-76, and Nck. However, in contrast to WASp, WAVE2 leaves this signaling complex and migrates peripherally together with vinculin to the membrane leading edge. Our experiments demonstrate that WASp and WAVE2 differ in their dynamics and their associated proteins. Thus, this study reveals the differential mechanisms regulating the function of these cytoskeletal proteins. PMID:25342748

Proteasome inhibition blocks NF-κB and ERK1/2 pathways, restores antigen expression and sensitizes resistant human melanoma to TCR-engineered CTLs

PubMed Central

Jazirehi, Ali R.; Economou, James S.

2012-01-01

Adoptive cell transfer (ACT) of ex vivo engineered autologous lymphocytes encoding high-affinity MART-1/HLA-A*0201-specific T-cell receptor (TCR) α/β chains (F5 CTL), densely infiltrate into sites of metastatic disease, mediating dramatic but partial clinical responses in melanoma patients. We hypothesized that MART-1 down-modulation in addition to aberrant apoptotic/survival signaling could confer resistance to death signals delivered by transgenic CTLs. To explore this hypothesis, we established an in vitro model of resistant (R) lines from MART-1+/HLA-A*0201+ F5 CTL-sensitive parental (P) lines under serial F5 CTL-selective pressure. We have recently reported that several melanoma R lines, while retaining MART-1 expression, exhibited constitutive NF-κB activation and over-expression of NF-κB-dependent resistance factors. Another established melanoma cell line M244, otherwise sensitive to F5 CTL, yielded R lines after serial F5 CTL selective pressure which had both reduced MART-1 expression levels, thus, could not be recognized, and were resistant to CTL-delivered apoptotic death signals. The proteasome inhibitor bortezomib blocked NF-κB activity, decreased phopspho-ERK1/2, increased phospho-JNK levels, reduced expression of resistance-factors, restored MART-1 expression to sufficient levels, which in combination allowed M244R lines be sensitized to F5 CTL-killing. These findings suggest that proteasome inhibition in immune resistant tumors can restore proapoptotic signaling and improve tumor antigen expression. PMID:22532603

CD3ζ-based chimeric antigen receptors mediate T cell activation via cis- and trans-signalling mechanisms: implications for optimization of receptor structure for adoptive cell therapy

PubMed Central

Bridgeman, J S; Ladell, K; Sheard, V E; Miners, K; Hawkins, R E; Price, D A; Gilham, D E

2014-01-01

Chimeric antigen receptors (CARs) can mediate redirected lysis of tumour cells in a major histocompatibility complex (MHC)-independent manner, thereby enabling autologous adoptive T cell therapy for a variety of malignant neoplasms. Currently, most CARs incorporate the T cell receptor (TCR) CD3ζ signalling chain; however, the precise mechanisms responsible for CAR-mediated T cell activation are unclear. In this study, we used a series of immunoreceptor tyrosine-based activation motif (ITAM)-mutant and transmembrane-modified receptors to demonstrate that CARs activate T cells both directly via the antigen-ligated signalling chain and indirectly via associated chains within the TCR complex. These observations allowed us to generate new receptors capable of eliciting polyfunctional responses in primary human T cells. This work increases our understanding of CAR function and identifies new avenues for the optimization of CAR-based therapeutic interventions. PMID:24116999

V(D)J recombination and allelic exclusion of a TCR beta-chain minilocus occurs in the absence of a functional promoter.

PubMed

Alvarez, J D; Anderson, S J; Loh, D Y

1995-08-01

Transcriptional activation of rearranging Ag receptor gene segments has been hypothesized to regulate their accessibility to V(D)J recombination. We analyzed the role of a functional promoter in the rearrangement of the murine TCR beta-chain locus using two transgenic minilocus constructs. These miniloci each contain an unrearranged V beta 8.3 gene. One has a wild-type V beta 8.3 gene, but the other has a V beta 8.3 gene with a promoter mutation that was previously shown to abrogate transcription in tissue culture. FACS analysis of thymus and lymph node cells from transgenic mouse lines showed that only the lines with the wild-type V beta 8.3 gene promoter express an 8.3 TCR beta-chain. Consistent with the protein expression data, V beta 8.3 gene transcripts were found only in the transgenic lines with the wild-type promoter. Using a quantitative PCR-based assay, it was shown that both types of transgenic lines recombine the V beta 8.3 gene at similar levels. Rearrangement of the transgenes was normal with respect to thymic development and junctional reading frame. Interestingly, both types of miniloci also underwent allelic exclusion in that recombination was blocked by the expression of a rearranged TCR beta-chain transgene. We conclude that a functional V beta gene promoter is not necessary for proper V(D)J recombination to occur.

Shank3 Is Part of a Zinc-Sensitive Signaling System That Regulates Excitatory Synaptic Strength.

PubMed

Arons, Magali H; Lee, Kevin; Thynne, Charlotte J; Kim, Sally A; Schob, Claudia; Kindler, Stefan; Montgomery, Johanna M; Garner, Craig C

2016-08-31

Shank3 is a multidomain scaffold protein localized to the postsynaptic density of excitatory synapses. Functional studies in vivo and in vitro support the concept that Shank3 is critical for synaptic plasticity and the trans-synaptic coupling between the reliability of presynaptic neurotransmitter release and postsynaptic responsiveness. However, how Shank3 regulates synaptic strength remains unclear. The C terminus of Shank3 contains a sterile alpha motif (SAM) domain that is essential for its postsynaptic localization and also binds zinc, thus raising the possibility that changing zinc levels modulate Shank3 function in dendritic spines. In support of this hypothesis, we find that zinc is a potent regulator of Shank3 activation and dynamics in rat hippocampal neurons. Moreover, we show that zinc modulation of synaptic transmission is Shank3 dependent. Interestingly, an autism spectrum disorder (ASD)-associated variant of Shank3 (Shank3(R87C)) retains its zinc sensitivity and supports zinc-dependent activation of AMPAR-mediated synaptic transmission. However, elevated zinc was unable to rescue defects in trans-synaptic signaling caused by the R87C mutation, implying that trans-synaptic increases in neurotransmitter release are not necessary for the postsynaptic effects of zinc. Together, these data suggest that Shank3 is a key component of a zinc-sensitive signaling system, regulating synaptic strength that may be impaired in ASD. Shank3 is a postsynaptic protein associated with neurodevelopmental disorders such as autism and schizophrenia. In this study, we show that Shank3 is a key component of a zinc-sensitive signaling system that regulates excitatory synaptic transmission. Intriguingly, an autism-associated mutation in Shank3 partially impairs this signaling system. Therefore, perturbation of zinc homeostasis may impair, not only synaptic functionality and plasticity, but also may lead to cognitive and behavioral abnormalities seen in patients with

Induction of activator protein (AP)-1 and nuclear factor-kappaB by CD28 stimulation involves both phosphatidylinositol 3-kinase and acidic sphingomyelinase signals.

PubMed

Edmead, C E; Patel, Y I; Wilson, A; Boulougouris, G; Hall, N D; Ward, S G; Sansom, D M

1996-10-15

A major obstacle in understanding the signaling events that follow CD28 receptor ligation arises from the fact that CD28 acts as a costimulus to TCR engagement, making it difficult to assess the relative contribution of CD28 signals as distinct from those of the TCR. To overcome this problem, we have exploited the observation that activated human T cell blasts can be stimulated via the CD28 surface molecule in the absence of antigenic challenge; thus, we have been able to observe the response of normal T cells to CD28 activation in isolation. Using this system, we observed that CD28 stimulation by B7-transfected CHO cells induced a proliferative response in T cells that was not accompanied by measurable IL-2 production. However, subsequent analysis of transcription factor generation revealed that B7 stimulation induced both activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB) complexes, but not NF-AT. In contrast, engagement of the TCR by class II MHC/superantigen, either with or without CD28 ligation, resulted in the induction of NF-AT, AP-1, and NF-kappaB as well as IL-2 production. Using selective inhibitors, we investigated the signaling pathways involved in the CD28-mediated induction of AP-1 and NF-kappaB. This revealed that NF-kappaB generation was sensitive to chloroquine, an inhibitor of acidic sphingomyelinase, but not to the phosphatidylinositol 3-kinase inhibitor, wortmannin. In contrast, AP-1 generation was inhibited by wortmannin and was also variably sensitive to chloroquine. These data suggest that in activated normal T cells, CD28-derived signals can stimulate proliferation at least in part via NF-kappaB and AP-1 generation, and that this response uses both acidic sphingomyelinase and phosphatidylinositol 3-kinase-linked pathways.

A set of enhanced green fluorescent protein concatemers for quantitative determination of nuclear localization signal strength.

PubMed

Böhm, Jennifer; Thavaraja, Ramya; Giehler, Susanne; Nalaskowski, Marcus M

2017-09-15

Regulated transport of proteins between nucleus and cytoplasm is an important process in the eukaryotic cell. In most cases, active nucleo-cytoplasmic protein transport is mediated by nuclear localization signal (NLS) and/or nuclear export signal (NES) motifs. In this study, we developed a set of vectors expressing enhanced GFP (EGFP) concatemers ranging from 2 to 12 subunits (2xEGFP to 12xEGFP) for analysis of NLS strength. As shown by in gel GFP fluorescence analysis and αGFP Western blotting, EGFP concatemers are expressed as fluorescent full-length proteins in eukaryotic cells. As expected, nuclear localization of concatemeric EGFPs decreases with increasing molecular weight. By oligonucleotide ligation this set of EGFP concatemers can be easily fused to NLS motifs. After determination of intracellular localization of EGFP concatemers alone and fused to different NLS motifs we calculated the size of a hypothetic EGFP concatemer showing a defined distribution of EGFP fluorescence between nucleus and cytoplasm (n/c ratio = 2). Clear differences of the size of the hypothetic EGFP concatemer depending on the fused NLS motif were observed. Therefore, we propose to use the size of this hypothetic concatemer as quantitative indicator for comparing strength of different NLS motifs. Copyright © 2017 Elsevier Inc. All rights reserved.

Constraints on the off-shell Higgs boson signal strength in the high-mass ZZ and WW final states with the ATLAS detector

NASA Astrophysics Data System (ADS)

Aad, G.; Abbott, B.; Abdallah, J.; Abdinov, O.; Aben, R.; Abolins, M.; AbouZeid, O. S.; Abramowicz, H.; Abreu, H.; Abreu, R.; Abulaiti, Y.; Acharya, B. S.; Adamczyk, L.; Adams, D. L.; Adelman, J.; Adomeit, S.; Adye, T.; Affolder, A. A.; Agatonovic-Jovin, T.; Aguilar-Saavedra, J. A.; Agustoni, M.; Ahlen, S. P.; Ahmadov, F.; Aielli, G.; Akerstedt, H.; Åkesson, T. P. A.; Akimoto, G.; Akimov, A. V.; Alberghi, G. L.; Albert, J.; Albrand, S.; Alconada Verzini, M. J.; Aleksa, M.; Aleksandrov, I. N.; Alexa, C.; Alexander, G.; Alexopoulos, T.; Alhroob, M.; Alimonti, G.; Alio, L.; Alison, J.; Alkire, S. P.; Allbrooke, B. M. M.; Allport, P. P.; Aloisio, A.; Alonso, A.; Alonso, F.; Alpigiani, C.; Altheimer, A.; Alvarez Gonzalez, B.; Piqueras, D. Álvarez; Alviggi, M. G.; Amako, K.; Amaral Coutinho, Y.; Amelung, C.; Amidei, D.; Amor Dos Santos, S. P.; Amorim, A.; Amoroso, S.; Amram, N.; Amundsen, G.; Anastopoulos, C.; Ancu, L. S.; Andari, N.; Andeen, T.; Anders, C. F.; Anders, G.; Anderson, K. 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A.; Ciocio, A.; Citron, Z. H.; Ciubancan, M.; Clark, A.; Clark, B. L.; Clark, P. J.; Clarke, R. N.; Cleland, W.; Clement, C.; Coadou, Y.; Cobal, M.; Coccaro, A.; Cochran, J.; Coffey, L.; Cogan, J. G.; Cole, B.; Cole, S.; Colijn, A. P.; Collot, J.; Colombo, T.; Compostella, G.; Conde Muiño, P.; Coniavitis, E.; Connell, S. H.; Connelly, I. A.; Consonni, S. M.; Consorti, V.; Constantinescu, S.; Conta, C.; Conti, G.; Conventi, F.; Cooke, M.; Cooper, B. D.; Cooper-Sarkar, A. M.; Copic, K.; Cornelissen, T.; Corradi, M.; Corriveau, F.; Corso-Radu, A.; Cortes-Gonzalez, A.; Cortiana, G.; Costa, G.; Costa, M. J.; Costanzo, D.; Côté, D.; Cottin, G.; Cowan, G.; Cox, B. E.; Cranmer, K.; Cree, G.; Crépé-Renaudin, S.; Crescioli, F.; Cribbs, W. A.; Crispin Ortuzar, M.; Cristinziani, M.; Croft, V.; Crosetti, G.; Cuhadar Donszelmann, T.; Cummings, J.; Curatolo, M.; Cuthbert, C.; Czirr, H.; Czodrowski, P.; D'Auria, S.; D'Onofrio, M.; Cunha Sargedas De Sousa, M. J. Da; Via, C. Da; Dabrowski, W.; Dafinca, A.; Dai, T.; Dale, O.; Dallaire, F.; Dallapiccola, C.; Dam, M.; Dandoy, J. R.; Daniells, A. C.; Danninger, M.; Dano Hoffmann, M.; Dao, V.; Darbo, G.; Darmora, S.; Dassoulas, J.; Dattagupta, A.; Davey, W.; David, C.; Davidek, T.; Davies, E.; Davies, M.; Davison, P.; Davygora, Y.; Dawe, E.; Dawson, I.; Daya-Ishmukhametova, R. K.; De, K.; de Asmundis, R.; De Castro, S.; De Cecco, S.; De Groot, N.; de Jong, P.; De la Torre, H.; De Lorenzi, F.; De Nooij, L.; De Pedis, D.; De Salvo, A.; De Sanctis, U.; De Santo, A.; De Vivie De Regie, J. B.; Dearnaley, W. J.; Debbe, R.; Debenedetti, C.; Dedovich, D. V.; Deigaard, I.; Del Peso, J.; Del Prete, T.; Delgove, D.; Deliot, F.; Delitzsch, C. M.; Deliyergiyev, M.; Dell'Acqua, A.; Dell'Asta, L.; Dell'Orso, M.; Della Pietra, M.; della Volpe, D.; Delmastro, M.; Delsart, P. A.; Deluca, C.; DeMarco, D. A.; Demers, S.; Demichev, M.; Demilly, A.; Denisov, S. P.; Derendarz, D.; Derkaoui, J. E.; Derue, F.; Dervan, P.; Desch, K.; Deterre, C.; Deviveiros, P. O.; Dewhurst, A.; Dhaliwal, S.; Di Ciaccio, A.; Di Ciaccio, L.; Di Domenico, A.; Di Donato, C.; Di Girolamo, A.; Di Girolamo, B.; Di Mattia, A.; Di Micco, B.; Di Nardo, R.; Di Simone, A.; Di Sipio, R.; Di Valentino, D.; Diaconu, C.; Diamond, M.; Dias, F. A.; Diaz, M. A.; Diehl, E. B.; Dietrich, J.; Diglio, S.; Dimitrievska, A.; Dingfelder, J.; Dita, P.; Dita, S.; Dittus, F.; Djama, F.; Djobava, T.; Djuvsland, J. I.; do Vale, M. A. B.; Dobos, D.; Dobre, M.; Doglioni, C.; Dohmae, T.; Dolejsi, J.; Dolezal, Z.; Dolgoshein, B. A.; Donadelli, M.; Donati, S.; Dondero, P.; Donini, J.; Dopke, J.; Doria, A.; Dova, M. T.; Doyle, A. T.; Drechsler, E.; Dris, M.; Dubreuil, E.; Duchovni, E.; Duckeck, G.; Ducu, O. A.; Duda, D.; Dudarev, A.; Duflot, L.; Duguid, L.; Dührssen, M.; Dunford, M.; Duran Yildiz, H.; Düren, M.; Durglishvili, A.; Duschinger, D.; Dwuznik, M.; Dyndal, M.; Eckardt, C.; Ecker, K. M.; Edson, W.; Edwards, N. C.; Ehrenfeld, W.; Eifert, T.; Eigen, G.; Einsweiler, K.; Ekelof, T.; El Kacimi, M.; Ellert, M.; Elles, S.; Ellinghaus, F.; Elliot, A. A.; Ellis, N.; Elmsheuser, J.; Elsing, M.; Emeliyanov, D.; Enari, Y.; Endner, O. C.; Endo, M.; Engelmann, R.; Erdmann, J.; Ereditato, A.; Ernis, G.; Ernst, J.; Ernst, M.; Errede, S.; Ertel, E.; Escalier, M.; Esch, H.; Escobar, C.; Esposito, B.; Etienvre, A. I.; Etzion, E.; Evans, H.; Ezhilov, A.; Fabbri, L.; Facini, G.; Fakhrutdinov, R. M.; Falciano, S.; Falla, R. J.; Faltova, J.; Fang, Y.; Fanti, M.; Farbin, A.; Farilla, A.; Farooque, T.; Farrell, S.; Farrington, S. M.; Farthouat, P.; Fassi, F.; Fassnacht, P.; Fassouliotis, D.; Favareto, A.; Fayard, L.; Federic, P.; Fedin, O. L.; Fedorko, W.; Feigl, S.; Feligioni, L.; Feng, C.; Feng, E. J.; Feng, H.; Fenyuk, A. B.; Martinez, P. Fernandez; Fernandez Perez, S.; Ferrag, S.; Ferrando, J.; Ferrari, A.; Ferrari, P.; Ferrari, R.; Ferreira de Lima, D. 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A.; Scheirich, D.; Schernau, M.; Schiavi, C.; Schillo, C.; Schioppa, M.; Schlenker, S.; Schmidt, E.; Schmieden, K.; Schmitt, C.; Schmitt, S.; Schmitt, S.; Schneider, B.; Schnellbach, Y. J.; Schnoor, U.; Schoeffel, L.; Schoening, A.; Schoenrock, B. D.; Schopf, E.; Schorlemmer, A. L. S.; Schott, M.; Schouten, D.; Schovancova, J.; Schramm, S.; Schreyer, M.; Schroeder, C.; Schuh, N.; Schultens, M. J.; Schultz-Coulon, H.-C.; Schulz, H.; Schumacher, M.; Schumm, B. A.; Schune, Ph.; Schwanenberger, C.; Schwartzman, A.; Schwarz, T. A.; Schwegler, Ph.; Schwemling, Ph.; Schwienhorst, R.; Schwindling, J.; Schwindt, T.; Schwoerer, M.; Sciacca, F. G.; Scifo, E.; Sciolla, G.; Scuri, F.; Scutti, F.; Searcy, J.; Sedov, G.; Sedykh, E.; Seema, P.; Seidel, S. C.; Seiden, A.; Seifert, F.; Seixas, J. M.; Sekhniaidze, G.; Sekula, S. J.; Selbach, K. E.; Seliverstov, D. M.; Semprini-Cesari, N.; Serfon, C.; Serin, L.; Serkin, L.; Serre, T.; Seuster, R.; Severini, H.; Sfiligoj, T.; Sforza, F.; Sfyrla, A.; Shabalina, E.; Shamim, M.; Shan, L. Y.; Shang, R.; Shank, J. T.; Shapiro, M.; Shatalov, P. B.; Shaw, K.; Shcherbakova, A.; Shehu, C. Y.; Sherwood, P.; Shi, L.; Shimizu, S.; Shimmin, C. O.; Shimojima, M.; Shiyakova, M.; Shmeleva, A.; Saadi, D. Shoaleh; Shochet, M. J.; Shojaii, S.; Shrestha, S.; Shulga, E.; Shupe, M. A.; Shushkevich, S.; Sicho, P.; Sidiropoulou, O.; Sidorov, D.; Sidoti, A.; Siegert, F.; Sijacki, Dj.; Silva, J.; Silver, Y.; Silverstein, S. B.; Simak, V.; Simard, O.; Simic, Lj.; Simion, S.; Simioni, E.; Simmons, B.; Simon, D.; Simoniello, R.; Sinervo, P.; Sinev, N. B.; Siragusa, G.; Sisakyan, A. N.; Sivoklokov, S. Yu.; Sjölin, J.; Sjursen, T. B.; Skinner, M. B.; Skottowe, H. P.; Skubic, P.; Slater, M.; Slavicek, T.; Slawinska, M.; Sliwa, K.; Smakhtin, V.; Smart, B. H.; Smestad, L.; Smirnov, S. Yu.; Smirnov, Y.; Smirnova, L. N.; Smirnova, O.; Smith, M. N. K.; Smizanska, M.; Smolek, K.; Snesarev, A. A.; Snidero, G.; Snyder, S.; Sobie, R.; Socher, F.; Soffer, A.; Soh, D. A.; Solans, C. A.; Solar, M.; Solc, J.; Soldatov, E. Yu.; Soldevila, U.; Solodkov, A. A.; Soloshenko, A.; Solovyanov, O. V.; Solovyev, V.; Sommer, P.; Song, H. Y.; Soni, N.; Sood, A.; Sopczak, A.; Sopko, B.; Sopko, V.; Sorin, V.; Sosa, D.; Sosebee, M.; Sotiropoulou, C. L.; Soualah, R.; Soueid, P.; Soukharev, A. M.; South, D.; Spagnolo, S.; Spalla, M.; Spanò, F.; Spearman, W. R.; Spettel, F.; Spighi, R.; Spigo, G.; Spiller, L. A.; Spousta, M.; Spreitzer, T.; Denis, R. D. St.; Staerz, S.; Stahlman, J.; Stamen, R.; Stamm, S.; Stanecka, E.; Stanescu, C.; Stanescu-Bellu, M.; Stanitzki, M. M.; Stapnes, S.; Starchenko, E. A.; Stark, J.; Staroba, P.; Starovoitov, P.; Staszewski, R.; Stavina, P.; Steinberg, P.; Stelzer, B.; Stelzer, H. J.; Stelzer-Chilton, O.; Stenzel, H.; Stern, S.; Stewart, G. A.; Stillings, J. A.; Stockton, M. C.; Stoebe, M.; Stoicea, G.; Stolte, P.; Stonjek, S.; Stradling, A. R.; Straessner, A.; Stramaglia, M. E.; Strandberg, J.; Strandberg, S.; Strandlie, A.; Strauss, E.; Strauss, M.; Strizenec, P.; Ströhmer, R.; Strom, D. M.; Stroynowski, R.; Strubig, A.; Stucci, S. A.; Stugu, B.; Styles, N. A.; Su, D.; Su, J.; Subramaniam, R.; Succurro, A.; Sugaya, Y.; Suhr, C.; Suk, M.; Sulin, V. V.; Sultansoy, S.; Sumida, T.; Sun, S.; Sun, X.; Sundermann, J. E.; Suruliz, K.; Susinno, G.; Sutton, M. R.; Suzuki, S.; Suzuki, Y.; Svatos, M.; Swedish, S.; Swiatlowski, M.; Sykora, I.; Sykora, T.; Ta, D.; Taccini, C.; Tackmann, K.; Taenzer, J.; Taffard, A.; Tafirout, R.; Taiblum, N.; Takai, H.; Takashima, R.; Takeda, H.; Takeshita, T.; Takubo, Y.; Talby, M.; Talyshev, A. A.; Tam, J. Y. C.; Tan, K. G.; Tanaka, J.; Tanaka, R.; Tanaka, S.; Tanaka, S.; Tannenwald, B. B.; Tannoury, N.; Tapprogge, S.; Tarem, S.; Tarrade, F.; Tartarelli, G. F.; Tas, P.; Tasevsky, M.; Tashiro, T.; Tassi, E.; Tavares Delgado, A.; Tayalati, Y.; Taylor, F. E.; Taylor, G. N.; Taylor, W.; Teischinger, F. A.; Teixeira Dias Castanheira, M.; Teixeira-Dias, P.; Temming, K. K.; Ten Kate, H.; Teng, P. K.; Teoh, J. J.; Tepel, F.; Terada, S.; Terashi, K.; Terron, J.; Terzo, S.; Testa, M.; Teuscher, R. J.; Therhaag, J.; Theveneaux-Pelzer, T.; Thomas, J. P.; Thomas-Wilsker, J.; Thompson, E. N.; Thompson, P. D.; Thompson, R. J.; Thompson, A. S.; Thomsen, L. A.; Thomson, E.; Thomson, M.; Thun, R. P.; Tibbetts, M. J.; Torres, R. E. Ticse; Tikhomirov, V. O.; Tikhonov, Yu. A.; Timoshenko, S.; Tiouchichine, E.; Tipton, P.; Tisserant, S.; Todorov, T.; Todorova-Nova, S.; Tojo, J.; Tokár, S.; Tokushuku, K.; Tollefson, K.; Tolley, E.; Tomlinson, L.; Tomoto, M.; Tompkins, L.; Toms, K.; Torrence, E.; Torres, H.; Torró Pastor, E.; Toth, J.; Touchard, F.; Tovey, D. R.; Trefzger, T.; Tremblet, L.; Tricoli, A.; Trigger, I. M.; Trincaz-Duvoid, S.; Tripiana, M. F.; Trischuk, W.; Trocmé, B.; Troncon, C.; Trottier-McDonald, M.; Trovatelli, M.; True, P.; Trzebinski, M.; Trzupek, A.; Tsarouchas, C.; Tseng, J. C.-L.; Tsiareshka, P. V.; Tsionou, D.; Tsipolitis, G.; Tsirintanis, N.; Tsiskaridze, S.; Tsiskaridze, V.; Tskhadadze, E. G.; Tsukerman, I. I.; Tsulaia, V.; Tsuno, S.; Tsybychev, D.; Tudorache, A.; Tudorache, V.; Tuna, A. N.; Tupputi, S. A.; Turchikhin, S.; Turecek, D.; Turra, R.; Turvey, A. J.; Tuts, P. M.; Tykhonov, A.; Tylmad, M.; Tyndel, M.; Ueda, I.; Ueno, R.; Ughetto, M.; Ugland, M.; Uhlenbrock, M.; Ukegawa, F.; Unal, G.; Undrus, A.; Unel, G.; Ungaro, F. C.; Unno, Y.; Unverdorben, C.; Urban, J.; Urquijo, P.; Urrejola, P.; Usai, G.; Usanova, A.; Vacavant, L.; Vacek, V.; Vachon, B.; Valderanis, C.; Valencic, N.; Valentinetti, S.; Valero, A.; Valery, L.; Valkar, S.; Valladolid Gallego, E.; Vallecorsa, S.; Valls Ferrer, J. A.; Van Den Wollenberg, W.; Van Der Deijl, P. C.; van der Geer, R.; van der Graaf, H.; Van Der Leeuw, R.; van Eldik, N.; van Gemmeren, P.; Van Nieuwkoop, J.; van Vulpen, I.; van Woerden, M. C.; Vanadia, M.; Vandelli, W.; Vanguri, R.; Vaniachine, A.; Vannucci, F.; Vardanyan, G.; Vari, R.; Varnes, E. W.; Varol, T.; Varouchas, D.; Vartapetian, A.; Varvell, K. E.; Vazeille, F.; Vazquez Schroeder, T.; Veatch, J.; Veloso, F.; Velz, T.; Veneziano, S.; Ventura, A.; Ventura, D.; Venturi, M.; Venturi, N.; Venturini, A.; Vercesi, V.; Verducci, M.; Verkerke, W.; Vermeulen, J. C.; Vest, A.; Vetterli, M. C.; Viazlo, O.; Vichou, I.; Vickey, T.; Vickey Boeriu, O. E.; Viehhauser, G. H. A.; Viel, S.; Vigne, R.; Villa, M.; Villaplana Perez, M.; Vilucchi, E.; Vincter, M. G.; Vinogradov, V. B.; Vivarelli, I.; Vives Vaque, F.; Vlachos, S.; Vladoiu, D.; Vlasak, M.; Vogel, M.; Vokac, P.; Volpi, G.; Volpi, M.; von der Schmitt, H.; von Radziewski, H.; von Toerne, E.; Vorobel, V.; Vorobev, K.; Vos, M.; Voss, R.; Vossebeld, J. H.; Vranjes, N.; Vranjes Milosavljevic, M.; Vrba, V.; Vreeswijk, M.; Vuillermet, R.; Vukotic, I.; Vykydal, Z.; Wagner, P.; Wagner, W.; Wahlberg, H.; Wahrmund, S.; Wakabayashi, J.; Walder, J.; Walker, R.; Walkowiak, W.; Wang, C.; Wang, F.; Wang, H.; Wang, H.; Wang, J.; Wang, J.; Wang, K.; Wang, R.; Wang, S. M.; Wang, T.; Wang, X.; Wanotayaroj, C.; Warburton, A.; Ward, C. P.; Wardrope, D. R.; Warsinsky, M.; Washbrook, A.; Wasicki, C.; Watkins, P. M.; Watson, A. T.; Watson, I. J.; Watson, M. F.; Watts, G.; Watts, S.; Waugh, B. M.; Webb, S.; Weber, M. S.; Weber, S. W.; Webster, J. S.; Weidberg, A. R.; Weinert, B.; Weingarten, J.; Weiser, C.; Weits, H.; Wells, P. S.; Wenaus, T.; Wengler, T.; Wenig, S.; Wermes, N.; Werner, M.; Werner, P.; Wessels, M.; Wetter, J.; Whalen, K.; Wharton, A. M.; White, A.; White, M. J.; White, R.; White, S.; Whiteson, D.; Wickens, F. J.; Wiedenmann, W.; Wielers, M.; Wienemann, P.; Wiglesworth, C.; Wiik-Fuchs, L. A. M.; Wildauer, A.; Wilkens, H. G.; Williams, H. H.; Williams, S.; Willis, C.; Willocq, S.; Wilson, A.; Wilson, J. A.; Wingerter-Seez, I.; Winklmeier, F.; Winter, B. T.; Wittgen, M.; Wittkowski, J.; Wollstadt, S. J.; Wolter, M. W.; Wolters, H.; Wosiek, B. K.; Wotschack, J.; Woudstra, M. J.; Wozniak, K. W.; Wu, M.; Wu, M.; Wu, S. L.; Wu, X.; Wu, Y.; Wyatt, T. R.; Wynne, B. M.; Xella, S.; Xu, D.; Xu, L.; Yabsley, B.; Yacoob, S.; Yakabe, R.; Yamada, M.; Yamaguchi, Y.; Yamamoto, A.; Yamamoto, S.; Yamanaka, T.; Yamauchi, K.; Yamazaki, Y.; Yan, Z.; Yang, H.; Yang, H.; Yang, Y.; Yao, L.; Yao, W.-M.; Yasu, Y.; Yatsenko, E.; Yau Wong, K. H.; Ye, J.; Ye, S.; Yeletskikh, I.; Yen, A. L.; Yildirim, E.; Yorita, K.; Yoshida, R.; Yoshihara, K.; Young, C.; Young, C. J. S.; Youssef, S.; Yu, D. R.; Yu, J.; Yu, J. M.; Yu, J.; Yuan, L.; Yurkewicz, A.; Yusuff, I.; Zabinski, B.; Zaidan, R.; Zaitsev, A. M.; Zalieckas, J.; Zaman, A.; Zambito, S.; Zanello, L.; Zanzi, D.; Zeitnitz, C.; Zeman, M.; Zemla, A.; Zengel, K.; Zenin, O.; Ženiš, T.; Zerwas, D.; Zhang, D.; Zhang, F.; Zhang, J.; Zhang, L.; Zhang, R.; Zhang, X.; Zhang, Z.; Zhao, X.; Zhao, Y.; Zhao, Z.; Zhemchugov, A.; Zhong, J.; Zhou, B.; Zhou, C.; Zhou, L.; Zhou, L.; Zhou, N.; Zhu, C. G.; Zhu, H.; Zhu, J.; Zhu, Y.; Zhuang, X.; Zhukov, K.; Zibell, A.; Zieminska, D.; Zimine, N. I.; Zimmermann, C.; Zimmermann, R.; Zimmermann, S.; Zinonos, Z.; Zinser, M.; Ziolkowski, M.; Živković, L.; Zobernig, G.; Zoccoli, A.; zur Nedden, M.; Zurzolo, G.; Zwalinski, L.

2015-07-01

Measurements of the ZZ and WW final states in the mass range above the and thresholds provide a unique opportunity to measure the off-shell coupling strength of the Higgs boson. This paper presents constraints on the off-shell Higgs boson event yields normalised to the Standard Model prediction (signal strength) in the , and final states. The result is based on pp collision data collected by the ATLAS experiment at the LHC, corresponding to an integrated luminosity of 20.3 fb at a collision energy of TeV. Using the method, the observed 95 confidence level (CL) upper limit on the off-shell signal strength is in the range 5.1-8.6, with an expected range of 6.7-11.0. In each case the range is determined by varying the unknown and background K-factor from higher-order quantum chromodynamics corrections between half and twice the value of the known signal K-factor. Assuming the relevant Higgs boson couplings are independent of the energy scale of the Higgs boson production, a combination with the on-shell measurements yields an observed (expected) 95 CL upper limit on in the range 4.5-7.5 (6.5-11.2) using the same variations of the background K-factor. Assuming that the unknown background K-factor is equal to the signal K-factor, this translates into an observed (expected) 95 CL upper limit on the Higgs boson total width of 22.7 (33.0) MeV.

Lipid rafts in T cell signalling and disease

PubMed Central

Jury, Elizabeth C.; Flores-Borja, Fabian; Kabouridis, Panagiotis S.

2007-01-01

Lipid rafts is a blanket term used to describe distinct areas in the plasma membrane rich in certain lipids and proteins and which are thought to perform diverse functions. A large number of studies report on lipid rafts having a key role in receptor signalling and activation of lymphocytes. In T cells, lipid raft involvement was demonstrated in the early steps during T cell receptor (TCR) stimulation. Interestingly, recent evidence has shown that signalling in these domains differs in T cells isolated from patients with autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Here, we discuss these findings and explore the potential of lipid rafts as targets for the development of a new class of agents to downmodulate immune responses and for the treatment of autoimmune diseases. PMID:17890113

Cytotoxic-T-lymphocyte antigen 4 receptor signaling for lymphocyte adhesion is mediated by C3G and Rap1.

PubMed

Kloog, Yoel; Mor, Adam

2014-03-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders.

Cytotoxic-T-Lymphocyte Antigen 4 Receptor Signaling for Lymphocyte Adhesion Is Mediated by C3G and Rap1

PubMed Central

Kloog, Yoel

2014-01-01

T-lymphocyte adhesion plays a critical role in both inflammatory and autoimmune responses. The small GTPase Rap1 is the key coordinator mediating T-cell adhesion to endothelial cells, antigen-presenting cells, and virus-infected cells. We describe a signaling pathway, downstream of the cytotoxic T-lymphocyte antigen 4 (CTLA-4) receptor, leading to Rap1-mediated adhesion. We identified a role for the Rap1 guanine nucleotide exchange factor C3G in the regulation of T-cell adhesion and showed that this factor is required for both T-cell receptor (TCR)-mediated and CTLA-4-mediated T-cell adhesion. Our data indicated that C3G translocates to the plasma membrane downstream of TCR signaling, where it regulates activation of Rap1. We also showed that CTLA-4 receptor signaling mediates tyrosine phosphorylation in the C3G protein, and that this is required for augmented activation of Rap1 and increased adhesion mediated by leukocyte function-associated antigen type 1 (LFA-1). Zap70 is required for C3G translocation to the plasma membrane, whereas the Src family member Hck facilitates C3G phosphorylation. These findings point to C3G and Hck as promising potential therapeutic targets for the treatment of T-cell-dependent autoimmune disorders. PMID:24396067

Assessment of Receiver Signal Strength Sensing for Location Estimation Based on Fisher Information

PubMed Central

Nielsen, John; Nielsen, Christopher

2016-01-01

Currently there is almost ubiquitous availability of wireless signaling for data communications within commercial building complexes resulting in receiver signal strength (RSS) observables that are typically sufficient for generating viable location estimates of mobile wireless devices. However, while RSS observables are generally plentiful, achieving an accurate estimation of location is difficult due to several factors affecting the electromagnetic coupling between the mobile antenna and the building access points that are not modeled and hence contribute to the overall estimation uncertainty. Such uncertainty is typically mitigated with a moderate redundancy of RSS sensor observations in combination with other constraints imposed on the mobile trajectory. In this paper, the Fisher Information (FI) of a set of RSS sensor observations in the context of variables related to the mobile location is developed. This provides a practical method of determining the potential location accuracy for the given set of wireless signals available. Furthermore, the information value of individual RSS measurements can be quantified and the RSS observables weighted accordingly in estimation combining algorithms. The practical utility of using FI in this context was demonstrated experimentally with an extensive set of RSS measurements recorded in an office complex. The resulting deviation of the mobile location estimation based on application of weighted likelihood processing to the experimental RSS data was shown to agree closely with the Cramer Rao bound determined from the FI analysis. PMID:27669262

A Higher Activation Threshold of Memory CD8+ T Cells Has a Fitness Cost That Is Modified by TCR Affinity during Tuberculosis.

PubMed

Carpenter, Stephen M; Nunes-Alves, Cláudio; Booty, Matthew G; Way, Sing Sing; Behar, Samuel M

2016-01-01

T cell vaccines against Mycobacterium tuberculosis (Mtb) and other pathogens are based on the principle that memory T cells rapidly generate effector responses upon challenge, leading to pathogen clearance. Despite eliciting a robust memory CD8+ T cell response to the immunodominant Mtb antigen TB10.4 (EsxH), we find the increased frequency of TB10.4-specific CD8+ T cells conferred by vaccination to be short-lived after Mtb challenge. To compare memory and naïve CD8+ T cell function during their response to Mtb, we track their expansions using TB10.4-specific retrogenic CD8+ T cells. We find that the primary (naïve) response outnumbers the secondary (memory) response during Mtb challenge, an effect moderated by increased TCR affinity. To determine whether the expansion of polyclonal memory T cells is restrained following Mtb challenge, we used TCRβ deep sequencing to track TB10.4-specific CD8+ T cells after vaccination and subsequent challenge in intact mice. Successful memory T cells, defined by their clonal expansion after Mtb challenge, express similar CDR3β sequences suggesting TCR selection by antigen. Thus, both TCR-dependent and -independent factors affect the fitness of memory CD8+ responses. The impaired expansion of the majority of memory T cell clonotypes may explain why some TB vaccines have not provided better protection.

A Higher Activation Threshold of Memory CD8+ T Cells Has a Fitness Cost That Is Modified by TCR Affinity during Tuberculosis

PubMed Central

Carpenter, Stephen M.; Nunes-Alves, Cláudio; Booty, Matthew G.; Way, Sing Sing; Behar, Samuel M.

2016-01-01

T cell vaccines against Mycobacterium tuberculosis (Mtb) and other pathogens are based on the principle that memory T cells rapidly generate effector responses upon challenge, leading to pathogen clearance. Despite eliciting a robust memory CD8+ T cell response to the immunodominant Mtb antigen TB10.4 (EsxH), we find the increased frequency of TB10.4-specific CD8+ T cells conferred by vaccination to be short-lived after Mtb challenge. To compare memory and naïve CD8+ T cell function during their response to Mtb, we track their expansions using TB10.4-specific retrogenic CD8+ T cells. We find that the primary (naïve) response outnumbers the secondary (memory) response during Mtb challenge, an effect moderated by increased TCR affinity. To determine whether the expansion of polyclonal memory T cells is restrained following Mtb challenge, we used TCRβ deep sequencing to track TB10.4-specific CD8+ T cells after vaccination and subsequent challenge in intact mice. Successful memory T cells, defined by their clonal expansion after Mtb challenge, express similar CDR3β sequences suggesting TCR selection by antigen. Thus, both TCR-dependent and -independent factors affect the fitness of memory CD8+ responses. The impaired expansion of the majority of memory T cell clonotypes may explain why some TB vaccines have not provided better protection. PMID:26745507

Where did my wifi go? Measuring soil moisture using wifi signal strength

NASA Astrophysics Data System (ADS)

Hut, Rolf; de Jeu, Richard

2015-04-01

Soil moisture is tricky to measure. Currently soil moisture is measured at small footprints using probes and other field devices, or at large footprints using satellites. Promising developments in measuring soil moisture are using fiber optic cables for measurements along a line, or using cosmos rays for field scale measurements. In this demonstration we present a low cost alternative to measure soil moisture at footprints of a few square meters. We use a wifi hotspot and a wifi dongle, both mounted in a cantenna for beam forming. We aim the hotspot on a piece of soil and put the dongle in the path of the reflection. By logging the signal strength of the wifi netwerk, we have a proxy for soil moisture. A first proof of concept is presented.

Estimating TCR using an integrated model-observation framework that accounts for spatio-temporal variability and pre-industrial radiative imbalances.

NASA Astrophysics Data System (ADS)

Haustein, K.; Schurer, A. P.; Venema, V.

2016-12-01

Apart from a few exceptions (e.g. Aldrin et al. 2012, Skeie et al. 2013) TCR estimates with EBMs are based on global data. Since these estimates don't represent the true spatial-temporal behaviour for observed temperature as well as external forcing (Marvel et al. 2015), we have developed a two-box EBM framework that accounts for these effects. In addition, external forcing from anthropogenic aerosol and GHGs tends to have different response times in comparison to volcanic stratospheric aerosols. Using PMIP3 and an extended ensemble of HadCM3 simulations (Euro500; Schurer et al. 2014) GCM simulations for the pre-industrial period, we obtain the fast and slow response time constants required in the EBM. With the most recent anthropogenic and natural forcing estimates, we test a range of TCR values against observations. The TCR/ECS ratio necessary to achieve that goal is taken from CMIP5 as observationally OHC-based estimates are notoriously unreliable. Given that observed and modelled OHC estimates are in agreement (Cheng et al. 2016), we argue that this should be the standard procedure the make inferences about ECS. Alternatively, it should be distinguished between equilibrium and effective climate sensitivity. The preliminary best estimate for TCR is 1.6K (1.1-2.2K) with an associated ECS value of 2.9K (2.0-4.0K). This is in good agreement with other D&A techniques that do use spatio-temporal patterns as well (e.g. Jones et al. 2016, Gillet et al. 2013). Correcting for natural ENSO variability and tas/tos-related inaccuracies (Richardson et al. 2016) further increases the robustness of the estimated sensitivity range. Our results also indicate that the small radiative imbalance caused by the period of very strong volcanic eruptions just before the CMIP5 historical period starts (1809-1840) has noteworthy implications for the response to later volcanic eruptions and the temperature evolution after 1850. Simply put, CMIP5-type simulations are slightly more sensitive

WASp family verprolin-homologous protein-2 (WAVE2) and Wiskott-Aldrich syndrome protein (WASp) engage in distinct downstream signaling interactions at the T cell antigen receptor site.

PubMed

Pauker, Maor H; Reicher, Barak; Joseph, Noah; Wortzel, Inbal; Jakubowicz, Shlomi; Noy, Elad; Perl, Orly; Barda-Saad, Mira

2014-12-12

T cell antigen receptor (TCR) engagement has been shown to activate pathways leading to actin cytoskeletal polymerization and reorganization, which are essential for lymphocyte activation and function. Several actin regulatory proteins were implicated in regulating the actin machinery, such as members of the Wiskott-Aldrich syndrome protein (WASp) family. These include WASp and the WASp family verprolin-homologous protein-2 (WAVE2). Although WASp and WAVE2 share several structural features, the precise regulatory mechanisms and potential redundancy between them have not been fully characterized. Specifically, unlike WASp, the dynamic molecular interactions that regulate WAVE2 recruitment to the cell membrane and specifically to the TCR signaling complex are largely unknown. Here, we identify the molecular mechanism that controls the recruitment of WAVE2 in comparison with WASp. Using fluorescence resonance energy transfer (FRET) and novel triple-color FRET (3FRET) technology, we demonstrate how WAVE2 signaling complexes are dynamically regulated during lymphocyte activation in vivo. We show that, similar to WASp, WAVE2 recruitment to the TCR site depends on protein-tyrosine kinase, ZAP-70, and the adaptors LAT, SLP-76, and Nck. However, in contrast to WASp, WAVE2 leaves this signaling complex and migrates peripherally together with vinculin to the membrane leading edge. Our experiments demonstrate that WASp and WAVE2 differ in their dynamics and their associated proteins. Thus, this study reveals the differential mechanisms regulating the function of these cytoskeletal proteins. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Constraints on the off-shell Higgs boson signal strength in the high-mass ZZ and WW final states with the ATLAS detector

DOE PAGES

Aad, G.

2015-07-17

The measurements of the ZZ and WW final states in the mass range above the \\(2m_Z\\) and \\(2m_W\\) thresholds provide a unique opportunity to measure the off-shell coupling strength of the Higgs boson. This paper presents constraints on the off-shell Higgs boson event yields normalised to the Standard Model prediction (signal strength) in the \\(ZZ \\rightarrow 4\\ell \\), \\(ZZ\\rightarrow 2\\ell 2\

SLAP deficiency enhances number and function of regulatory T cells preventing chronic autoimmune arthritis in SKG mice.

PubMed

Peterson, Lisa K; Shaw, Laura A; Joetham, Anthony; Sakaguchi, Shimon; Gelfand, Erwin W; Dragone, Leonard L

2011-02-15

To test if manipulating TCR complex-mediated signaling (TCR signaling) could treat autoimmune disease, we generated the double SKG Src-like adapter protein (SLAP) knockout (DSSKO) mouse model. The SKG mutation in ZAP70 and SLAP have opposing functions on the regulation of TCR signaling. The combination of these two mutations alters TCR signaling in the context of a defined genetic background, uniform environmental conditions, and a well-characterized signaling disruption. In contrast to SKG mice, DSSKO mice do not develop zymosan-induced chronic autoimmune arthritis. This arthritis prevention is not due to significant alterations in thymocyte development or repertoire selection but instead enhanced numbers of regulatory T cells (Tregs) and decreased numbers of Th17 cells skewing the ratio of Tregs to autoreactive effector T cells. Treg depletion and/or functional blockade led to the development of arthritis in DSSKO mice. In vitro suppression of effector T cell proliferation was also enhanced, demonstrating that DSSKO mice have increased numbers of Tregs with increased function. Understanding how TCR signals influence development, expansion, and function of Tregs in DSSKO mice could advance our ability to manipulate Treg biology to treat ultimately autoimmune disease.

SLAP Deficiency Enhances Number and Function of Regulatory T Cells Preventing Chronic Autoimmune Arthritis in SKG Mice

PubMed Central

Peterson, Lisa K.; Shaw, Laura A.; Joetham, Anthony; Sakaguchi, Shimon; Gelfand, Erwin W.; Dragone, Leonard L.

2011-01-01

To test if manipulating TCR complex-mediated signaling (TCR signaling) could treat autoimmune disease, we generated the double SKG Src-like adapter protein (SLAP) knockout (DSSKO) mouse model. The SKG mutation in ZAP70 and SLAP have opposing functions on the regulation of TCR signaling. The combination of these two mutations alters TCR signaling in the context of a defined genetic background, uniform environmental conditions, and a well-characterized signaling disruption. In contrast to SKG mice, DSSKO mice do not develop zymosan-induced chronic autoimmune arthritis. This arthritis prevention is not due to significant alterations in thymocyte development or repertoire selection but instead enhanced numbers of regulatory T cells (Tregs) and decreased numbers of Th17 cells skewing the ratio of Tregs to autoreactive effector T cells. Treg depletion and/or functional blockade led to the development of arthritis in DSSKO mice. In vitro suppression of effector T cell proliferation was also enhanced, demonstrating that DSSKO mice have increased numbers of Tregs with increased function. Understanding how TCR signals influence development, expansion, and function of Tregs in DSSKO mice could advance our ability to manipulate Treg biology to treat ultimately autoimmune disease. PMID:21248251

PRIMAL: Page Rank-Based Indoor Mapping and Localization Using Gene-Sequenced Unlabeled WLAN Received Signal Strength

PubMed Central

Zhou, Mu; Zhang, Qiao; Xu, Kunjie; Tian, Zengshan; Wang, Yanmeng; He, Wei

2015-01-01

Due to the wide deployment of wireless local area networks (WLAN), received signal strength (RSS)-based indoor WLAN localization has attracted considerable attention in both academia and industry. In this paper, we propose a novel page rank-based indoor mapping and localization (PRIMAL) by using the gene-sequenced unlabeled WLAN RSS for simultaneous localization and mapping (SLAM). Specifically, first of all, based on the observation of the motion patterns of the people in the target environment, we use the Allen logic to construct the mobility graph to characterize the connectivity among different areas of interest. Second, the concept of gene sequencing is utilized to assemble the sporadically-collected RSS sequences into a signal graph based on the transition relations among different RSS sequences. Third, we apply the graph drawing approach to exhibit both the mobility graph and signal graph in a more readable manner. Finally, the page rank (PR) algorithm is proposed to construct the mapping from the signal graph into the mobility graph. The experimental results show that the proposed approach achieves satisfactory localization accuracy and meanwhile avoids the intensive time and labor cost involved in the conventional location fingerprinting-based indoor WLAN localization. PMID:26404274

Decreased SAP expression in T cells from patients with SLE contributes to early signaling abnormalities and reduced IL-2 production

PubMed Central

Karampetsou, Maria P.; Comte, Denis; Kis-Toth, Katalin; Terhorst, Cox; Kyttaris, Vasileios C.; Tsokos, George C.

2016-01-01

T cells from patients with systemic lupus erythematosus (SLE) display a number of functions including increased early signaling events following engagement of the T cell receptor (TCR). Signaling lymphocytic activation molecule family (SLAMF) cell surface receptors and the X-chromosome-defined signaling lymphocytic activation molecule-associated protein (SAP) adaptor are important in the development of several immunocyte lineages and modulating immune response. Here we present evidence that SAP protein levels are decreased in T cells and in their main subsets isolated from 32 women and 3 men with SLE independently of disease activity. In SLE T cells the SAP protein is also subject to increased degradation by a caspase-3. Forced expression of SAP in SLE T cells simultaneously heightened IL-2 production, calcium (Ca2+) responses and tyrosine phosphorylation of a number of proteins. Exposure of normal T cells to SLE serum IgG, known to contain anti-CD3/TCR antibodies, resulted in SAP downregulation. We conclude that SLE T cells display reduced levels of the adaptor protein SAP probably as a result of continuous T cell activation and degradation by caspase-3. Restoration of SAP levels in SLE T cells corrects the overexcitable lupus T cell phenotype. PMID:27183584

Bayesian-based localization of wireless capsule endoscope using received signal strength.

PubMed

Nadimi, Esmaeil S; Blanes-Vidal, Victoria; Tarokh, Vahid; Johansen, Per Michael

2014-01-01

In wireless body area sensor networking (WBASN) applications such as gastrointestinal (GI) tract monitoring using wireless video capsule endoscopy (WCE), the performance of out-of-body wireless link propagating through different body media (i.e. blood, fat, muscle and bone) is still under investigation. Most of the localization algorithms are vulnerable to the variations of path-loss coefficient resulting in unreliable location estimation. In this paper, we propose a novel robust probabilistic Bayesian-based approach using received-signal-strength (RSS) measurements that accounts for Rayleigh fading, variable path-loss exponent and uncertainty in location information received from the neighboring nodes and anchors. The results of this study showed that the localization root mean square error of our Bayesian-based method was 1.6 mm which was very close to the optimum Cramer-Rao lower bound (CRLB) and significantly smaller than that of other existing localization approaches (i.e. classical MDS (64.2mm), dwMDS (32.2mm), MLE (36.3mm) and POCS (2.3mm)).

The opposite role of two UBA-UBX containing proteins, p47 and SAKS1 in the degradation of a single ERAD substrate, α-TCR.

PubMed

Park, Eun Sil; Yoo, Yung Joon; Elangovan, Muthukumar

2017-01-01

The UBA-UBX domain-containing proteins can interact with ubiquitinated substrates and p97 during endoplasmic reticulum-associated degradation (ERAD). Here, we found that the expressions of all UBA-UBX genes p47, SAKS1, UBXD8, FAF1, and UBXD7 were elevated upon ER stress, albeit with different levels. Of which p47, SAKS1, and UBXD8 are 'immediate' respondents whereas FAF1 and UBXD7 were 'late' respondents to ER stress. Interestingly, the expression of specific UBA-UBX genes were altered in cells stably expressing three different ERAD substrates such as α-TCR, α1-antitrypsin, and δCD3. We first found that p47 and UBXD8 expression levels were increased in α-TCR and α1-antitrypsin stable cell lines, respectively, whereas SAKS1 expression level was reduced in all the three ERAD substrates tested. Of note, we also found p47 promotes, whereas SASK1 delays the degradation of a single ERAD substrate, α-TCR. Additionally, we found that SAKS1 selectively inhibits the degradation of ERAD substrates without affecting cytosolic proteasomal substrates. Taken together, our results identified that UBA-UBX proteins possess substrate selectivity and opposite role of two different UBA-UBX proteins in the degradation of a single ERAD substrate.

Suboptimal T-cell receptor signaling compromises protein translation, ribosome biogenesis, and proliferation of mouse CD8 T cells.

PubMed

Tan, Thomas C J; Knight, John; Sbarrato, Thomas; Dudek, Kate; Willis, Anne E; Zamoyska, Rose

2017-07-25

Global transcriptomic and proteomic analyses of T cells have been rich sources of unbiased data for understanding T-cell activation. Lack of full concordance of these datasets has illustrated that important facets of T-cell activation are controlled at the level of translation. We undertook translatome analysis of CD8 T-cell activation, combining polysome profiling and microarray analysis. We revealed that altering T-cell receptor stimulation influenced recruitment of mRNAs to heavy polysomes and translation of subsets of genes. A major pathway that was compromised, when TCR signaling was suboptimal, was linked to ribosome biogenesis, a rate-limiting factor in both cell growth and proliferation. Defective TCR signaling affected transcription and processing of ribosomal RNA precursors, as well as the translation of specific ribosomal proteins and translation factors. Mechanistically, IL-2 production was compromised in weakly stimulated T cells, affecting the abundance of Myc protein, a known regulator of ribosome biogenesis. Consequently, weakly activated T cells showed impaired production of ribosomes and a failure to maintain proliferative capacity after stimulation. We demonstrate that primary T cells respond to various environmental cues by regulating ribosome biogenesis and mRNA translation at multiple levels to sustain proliferation and differentiation.

Immune monitoring and TCR sequencing of CD4 T cells in a long term responsive patient with metastasized pancreatic ductal carcinoma treated with individualized, neoepitope-derived multipeptide vaccines: a case report.

PubMed

Sonntag, Katja; Hashimoto, Hisayoshi; Eyrich, Matthias; Menzel, Moritz; Schubach, Max; Döcker, Dennis; Battke, Florian; Courage, Carolina; Lambertz, Helmut; Handgretinger, Rupert; Biskup, Saskia; Schilbach, Karin

2018-02-06

Cancer vaccines can effectively establish clinically relevant tumor immunity. Novel sequencing approaches rapidly identify the mutational fingerprint of tumors, thus allowing to generate personalized tumor vaccines within a few weeks from diagnosis. Here, we report the case of a 62-year-old patient receiving a four-peptide-vaccine targeting the two sole mutations of his pancreatic tumor, identified via exome sequencing. Vaccination started during chemotherapy in second complete remission and continued monthly thereafter. We tracked IFN-γ + T cell responses against vaccine peptides in peripheral blood after 12, 17 and 34 vaccinations by analyzing T-cell receptor (TCR) repertoire diversity and epitope-binding regions of peptide-reactive T-cell lines and clones. By restricting analysis to sorted IFN-γ-producing T cells we could assure epitope-specificity, functionality, and T H 1 polarization. A peptide-specific T-cell response against three of the four vaccine peptides could be detected sequentially. Molecular TCR analysis revealed a broad vaccine-reactive TCR repertoire with clones of discernible specificity. Four identical or convergent TCR sequences could be identified at more than one time-point, indicating timely persistence of vaccine-reactive T cells. One dominant TCR expressing a dual TCRVα chain could be found in three T-cell clones. The observed T-cell responses possibly contributed to clinical outcome: The patient is alive 6 years after initial diagnosis and in complete remission for 4 years now. Therapeutic vaccination with a neoantigen-derived four-peptide vaccine resulted in a diverse and long-lasting immune response against these targets which was associated with prolonged clinical remission. These data warrant confirmation in a larger proof-of concept clinical trial.

Shp1 regulates T cell homeostasis by limiting IL-4 signals

PubMed Central

Johnson, Dylan J.; Pao, Lily I.; Dhanji, Salim; Murakami, Kiichi

2013-01-01

The protein-tyrosine phosphatase Shp1 is expressed ubiquitously in hematopoietic cells and is generally viewed as a negative regulatory molecule. Mutations in Ptpn6, which encodes Shp1, result in widespread inflammation and premature death, known as the motheaten (me) phenotype. Previous studies identified Shp1 as a negative regulator of TCR signaling, but the severe systemic inflammation in me mice may have confounded our understanding of Shp1 function in T cell biology. To define the T cell–intrinsic role of Shp1, we characterized mice with a T cell–specific Shp1 deletion (Shp1fl/fl CD4-cre). Surprisingly, thymocyte selection and peripheral TCR sensitivity were unaltered in the absence of Shp1. Instead, Shp1fl/fl CD4-cre mice had increased frequencies of memory phenotype T cells that expressed elevated levels of CD44. Activation of Shp1-deficient CD4+ T cells also resulted in skewing to the Th2 lineage and increased IL-4 production. After IL-4 stimulation of Shp1-deficient T cells, Stat 6 activation was sustained, leading to enhanced Th2 skewing. Accordingly, we observed elevated serum IgE in the steady state. Blocking or genetic deletion of IL-4 in the absence of Shp1 resulted in a marked reduction of the CD44hi population. Therefore, Shp1 is an essential negative regulator of IL-4 signaling in T lymphocytes. PMID:23797092

PD-1 signalling in CD4+ T cells restrains their clonal expansion to an immunogenic stimulus, but is not critically required for peptide-induced tolerance

PubMed Central

Konkel, Joanne E; Frommer, Friederike; Leech, Melanie D; Yagita, Hideo; Waisman, Ari; Anderton, Stephen M

2010-01-01

The ultimate outcome of T-cell recognition of peptide–major histocompatibility complex (MHC) complexes is determined by the molecular context in which antigen presentation is provided. The paradigm is that, after exposure to peptides presented by steady-state dendritic cells (DCs), inhibitory signals dominate, leading to the deletion and/or functional inactivation of antigen-reactive T cells. This has been utilized in a variety of models providing peptide antigen in soluble form in the absence of adjuvant. A co-inhibitory molecule of considerable current interest is PD-1. Here we show that there is the opportunity for the PD-1/PD-L1 interaction to function in inhibiting the T-cell response during tolerance induction. Using traceable CD4+ T-cell receptor (TCR) transgenic cells, together with a blocking antibody to disrupt PD-1 signalling, we explored the roles of PD-1 in the induction of tolerance versus a productive immune response. Intact PD-1 signalling played a role in limiting the extent of CD4+ T-cell accumulation in response to an immunogenic stimulus. However, PD-1 signalling was not required for either the induction, or the maintenance, of peptide-induced tolerance; a conclusion underlined by successful tolerance induction in TCR transgenic cells genetically deficient for PD-1. These observations contrast with the reported requirement for PD-1 signals in CD8+ T-cell tolerance. PMID:20113370

Achievable Strength-Based Signal Detection in Quantity-Constrained PAM OOK Concentration-Encoded Molecular Communication.

PubMed

Mahfuz, Mohammad Upal

2016-10-01

In this paper, the expressions of achievable strength-based detection probabilities of concentration-encoded molecular communication (CEMC) system have been derived based on finite pulsewidth (FP) pulse-amplitude modulated (PAM) on-off keying (OOK) modulation scheme and strength threshold. An FP-PAM system is characterized by its duty cycle α that indicates the fraction of the entire symbol duration the transmitter remains on and transmits the signal. Results show that the detection performance of an FP-PAM OOK CEMC system significantly depends on the statistical distribution parameters of diffusion-based propagation noise and intersymbol interference (ISI). Analytical detection performance of an FP-PAM OOK CEMC system under ISI scenario has been explained and compared based on receiver operating characteristics (ROC) for impulse (i.e., spike)-modulated (IM) and FP-PAM CEMC schemes. It is shown that the effects of diffusion noise and ISI on ROC can be explained separately based on their communication range-dependent statistics. With full duty cycle, an FP-PAM scheme provides significantly worse performance than an IM scheme. The paper also analyzes the performance of the system when duty cycle, transmission data rate, and quantity of molecules vary.

Investigating strength and frequency effects in recognition memory using type-2 signal detection theory.

PubMed

Higham, Philip A; Perfect, Timothy J; Bruno, Davide

2009-01-01

Criterion- versus distribution-shift accounts of frequency and strength effects in recognition memory were investigated with Type-2 signal detection receiver operating characteristic (ROC) analysis, which provides a measure of metacognitive monitoring. Experiment 1 demonstrated a frequency-based mirror effect, with a higher hit rate and lower false alarm rate, for low frequency words compared with high frequency words. In Experiment 2, the authors manipulated item strength with repetition, which showed an increased hit rate but no effect on the false alarm rate. Whereas Type-1 indices were ambiguous as to whether these effects were based on a criterion- or distribution-shift model, the two models predict opposite effects on Type-2 distractor monitoring under some assumptions. Hence, Type-2 ROC analysis discriminated between potential models of recognition that could not be discriminated using Type-1 indices alone. In Experiment 3, the authors manipulated Type-1 response bias by varying the number of old versus new response categories to confirm the assumptions made in Experiments 1 and 2. The authors conclude that Type-2 analyses are a useful tool for investigating recognition memory when used in conjunction with more traditional Type-1 analyses.

Decreased SAP Expression in T Cells from Patients with Systemic Lupus Erythematosus Contributes to Early Signaling Abnormalities and Reduced IL-2 Production.

PubMed

Karampetsou, Maria P; Comte, Denis; Kis-Toth, Katalin; Terhorst, Cox; Kyttaris, Vasileios C; Tsokos, George C

2016-06-15

T cells from patients with systemic lupus erythematosus (SLE) display a number of abnormalities, including increased early signaling events following engagement of the TCR. Signaling lymphocytic activation molecule family cell surface receptors and the X-chromosome-defined signaling lymphocytic activation molecule-associated protein (SAP) adaptor are important in the development of several immunocyte lineages and modulating the immune response. We present evidence that SAP protein levels are decreased in T cells and in their main subsets isolated from 32 women and three men with SLE, independent of disease activity. In SLE T cells, SAP protein is also subject to increased degradation by caspase-3. Forced expression of SAP in SLE T cells normalized IL-2 production, calcium (Ca(2+)) responses, and tyrosine phosphorylation of a number of proteins. Exposure of normal T cells to SLE serum IgG, known to contain anti-CD3/TCR Abs, resulted in SAP downregulation. We conclude that SLE T cells display reduced levels of the adaptor protein SAP, probably as a result of continuous T cell activation and degradation by caspase-3. Restoration of SAP levels in SLE T cells corrects the overexcitable lupus T cell phenotype. Copyright © 2016 by The American Association of Immunologists, Inc.

Imaging Vesicular Traffic at the Immune Synapse.

PubMed

Bouchet, Jérôme; Del Río-Iñiguez, Iratxe; Alcover, Andrés

2017-01-01

Immunological synapse formation is the result of a profound T cell polarization process that involves the coordinated action of the actin and microtubule cytoskeleton, as well as intracellular vesicle traffic. Endosomal vesicle traffic ensures the targeting of the T cell receptor (TCR) and various signaling molecules to the synapse, being necessary for the generation of signaling complexes downstream of the TCR. Here we describe the microscopy imaging methods that we currently use to unveil how TCR and signaling molecules are associated with endosomal compartments and deliver their cargo to the immunological synapse.

Influenza virus-specific TCR-transduced T cells as a model for adoptive immunotherapy

PubMed Central

Berdien, Belinda; Reinhard, Henrike; Meyer, Sabrina; Spöck, Stefanie; Kröger, Nicolaus; Atanackovic, Djordje; Fehse, Boris

2013-01-01

Adoptive transfer of T lymphocytes equipped with tumor-antigen specific T-cell receptors (TCRs) represents a promising strategy in cancer immunotherapy, but the approach remains technically demanding. Using influenza virus (Flu)-specific T-cell responses as a model system we compared different methods for the generation of T-cell clones and isolation of antigen-specific TCRs. Altogether, we generated 12 CD8+ T-cell clones reacting to the Flu matrix protein (Flu-M) and 6 CD4+ T-cell clones reacting to the Flu nucleoprotein (Flu-NP) from 4 healthy donors. IFN-γ-secretion-based enrichment of antigen-specific cells, optionally combined with tetramer staining, was the most efficient way for generating T-cell clones. In contrast, the commonly used limiting dilution approach was least efficient. TCR genes were isolated from T-cell clones and cloned into both a previously used gammaretroviral LTR-vector, MP91 and the novel lentiviral self-inactivating vector LeGO-MP that contains MP91-derived promotor and regulatory elements. To directly compare their functional efficiencies, we in parallel transduced T-cell lines and primary T cells with the two vectors encoding identical TCRs. Transduction efficiencies were approximately twice higher with the gammaretroviral vector. Secretion of high amounts of IFN-γ, IL-2 and TNF-α by transduced cells after exposure to the respective influenza target epitope proved efficient specificity transfer of the isolated TCRs to primary T-cells for both vectors, at the same time indicating superior functionality of MP91-transduced cells. In conclusion, we have developed optimized strategies to obtain and transfer antigen-specific TCRs as well as designed a novel lentiviral vector for TCR-gene transfer. Our data may help to improve adoptive T-cell therapies. PMID:23428899

Effect Of Simulated Microgravity On Activated T Cell Gene Transcription

NASA Technical Reports Server (NTRS)

Morrow, Maureen A.

2003-01-01

Studies of T lymphocytes under the shear stress environment of clinorotation have demonstrated an inhibition of activation in response to TCR mediated signaling. These results mimic those observed during space flight. This work investigates the molecular signaling events of T lymphocyte activation with clinorotation. Purified human T lymphocytes and the T cell clone Jurkat exhibit an uncoupling of signaling as mediated through the TCR. Activation of the transcription factor AP-1 is inhibited while activation of NFAT occurs. NFAT dephosphorylation and activation is dependent on sustained Ca(++) influx. Alternatively, AP-1, which consists of two transcription factors, jun and fos, is activated by PKC and Ras mediated pathways. TCR signaling is known to be dependent on cytoskeletal rearrangements, in particular, raft aggregation is critical. Raft aggregation, as mediated through GM, crosslinking, overcomes the inhibition of T lymphocyte activation with clinorotation, indicating that the block is occurring upstream of raft aggregation. Clinorotation is shown to have an effect similar to a weak TCR signal.

Enhancement of Beaconless Location-Based Routing with Signal Strength Assistance for Ad-Hoc Networks

NASA Astrophysics Data System (ADS)

Chen, Guowei; Itoh, Kenichi; Sato, Takuro

Routing in Ad-hoc networks is unreliable due to the mobility of the nodes. Location-based routing protocols, unlike other protocols which rely on flooding, excel in network scalability. Furthermore, new location-based routing protocols, like, e. g. BLR [1], IGF [2], & CBF [3] have been proposed, with the feature of not requiring beacons in MAC-layer, which improve more in terms of scalability. Such beaconless routing protocols can work efficiently in dense network areas. However, these protocols' algorithms have no ability to avoid from routing into sparse areas. In this article, historical signal strength has been added as a factor into the BLR algorithm, which avoids routing into sparse area, and consequently improves the global routing efficiency.

Distinct T cell interactions with HLA class II tetramers characterize a spectrum of TCR affinities in the human antigen-specific T cell response.

PubMed

Reichstetter, S; Ettinger, R A; Liu, A W; Gebe, J A; Nepom, G T; Kwok, W W

2000-12-15

The polyclonal nature of T cells expanding in an ongoing immune response results in a range of disparate affinities and activation potential. Recently developed human class II tetramers provide a means to analyze this diversity by direct characterization of the trimolecular TCR-peptide-MHC interaction in live cells. Two HSV-2 VP16(369-379)-specific, DQA1*0102/DQB1*0602 (DQ0602)-restricted T cell clones were compared by means of T cell proliferation assay and HLA-DQ0602 tetramer staining. These two clones were obtained from the same subject, but show different TCR gene usage. Clone 48 was 10-fold more sensitive to VP16(369-379) peptide stimulation than clone 5 as assayed by proliferation assays, correlating with differences in MHC tetramer binding. Clone 48 gave positive staining with the DQ0602/VP16(369-379) tetramer at either 23 or 37 degrees C. Weak staining was also observed at 4 degrees C. Clone 5 showed weaker staining compared with clone 48 at 37 degrees C, and no staining was observed at 23 degrees C or on ice. Receptor internalization was not required for positive staining. Competitive binding indicates that the cell surface TCR of clone 48 has higher affinity for the DQ0602/VP16(369-379) complex than clone 5. The higher binding affinity of clone 48 for the peptide-MHC complex also correlates with a slower dissociation rate compared with clone 5.

Constraints on the off-shell Higgs boson signal strength in the high-mass ZZ and WW final states with the ATLAS detector.

PubMed

Aad, G; Abbott, B; Abdallah, J; Abdinov, O; Aben, R; Abolins, M; AbouZeid, O S; Abramowicz, H; Abreu, H; Abreu, R; Abulaiti, Y; Acharya, B S; Adamczyk, L; Adams, D L; Adelman, J; Adomeit, S; Adye, T; Affolder, A A; Agatonovic-Jovin, T; Aguilar-Saavedra, J A; Agustoni, M; Ahlen, S P; Ahmadov, F; Aielli, G; Akerstedt, H; Åkesson, T P A; Akimoto, G; Akimov, A V; Alberghi, G L; Albert, J; Albrand, S; Alconada Verzini, M J; Aleksa, M; Aleksandrov, I N; Alexa, C; Alexander, G; Alexopoulos, T; Alhroob, M; Alimonti, G; Alio, L; Alison, J; Alkire, S P; Allbrooke, B M M; Allport, P P; Aloisio, A; Alonso, A; Alonso, F; Alpigiani, C; Altheimer, A; Alvarez Gonzalez, B; Piqueras, D Álvarez; Alviggi, M G; Amako, K; Amaral Coutinho, Y; Amelung, C; Amidei, D; Amor Dos Santos, S P; Amorim, A; Amoroso, S; Amram, N; Amundsen, G; Anastopoulos, C; Ancu, L S; Andari, N; Andeen, T; Anders, C F; Anders, G; Anderson, K J; Andreazza, A; Andrei, V; Angelidakis, S; Angelozzi, I; Anger, P; Angerami, A; Anghinolfi, F; 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Yao, W-M; Yasu, Y; Yatsenko, E; Yau Wong, K H; Ye, J; Ye, S; Yeletskikh, I; Yen, A L; Yildirim, E; Yorita, K; Yoshida, R; Yoshihara, K; Young, C; Young, C J S; Youssef, S; Yu, D R; Yu, J; Yu, J M; Yu, J; Yuan, L; Yurkewicz, A; Yusuff, I; Zabinski, B; Zaidan, R; Zaitsev, A M; Zalieckas, J; Zaman, A; Zambito, S; Zanello, L; Zanzi, D; Zeitnitz, C; Zeman, M; Zemla, A; Zengel, K; Zenin, O; Ženiš, T; Zerwas, D; Zhang, D; Zhang, F; Zhang, J; Zhang, L; Zhang, R; Zhang, X; Zhang, Z; Zhao, X; Zhao, Y; Zhao, Z; Zhemchugov, A; Zhong, J; Zhou, B; Zhou, C; Zhou, L; Zhou, L; Zhou, N; Zhu, C G; Zhu, H; Zhu, J; Zhu, Y; Zhuang, X; Zhukov, K; Zibell, A; Zieminska, D; Zimine, N I; Zimmermann, C; Zimmermann, R; Zimmermann, S; Zinonos, Z; Zinser, M; Ziolkowski, M; Živković, L; Zobernig, G; Zoccoli, A; Zur Nedden, M; Zurzolo, G; Zwalinski, L

Measurements of the ZZ and WW final states in the mass range above the [Formula: see text] and [Formula: see text] thresholds provide a unique opportunity to measure the off-shell coupling strength of the Higgs boson. This paper presents constraints on the off-shell Higgs boson event yields normalised to the Standard Model prediction (signal strength) in the [Formula: see text], [Formula: see text] and [Formula: see text] final states. The result is based on pp collision data collected by the ATLAS experiment at the LHC, corresponding to an integrated luminosity of 20.3 fb[Formula: see text] at a collision energy of [Formula: see text] TeV. Using the [Formula: see text] method, the observed 95 [Formula: see text] confidence level (CL) upper limit on the off-shell signal strength is in the range 5.1-8.6, with an expected range of 6.7-11.0. In each case the range is determined by varying the unknown [Formula: see text] and [Formula: see text] background K-factor from higher-order quantum chromodynamics corrections between half and twice the value of the known signal K-factor. Assuming the relevant Higgs boson couplings are independent of the energy scale of the Higgs boson production, a combination with the on-shell measurements yields an observed (expected) 95 [Formula: see text] CL upper limit on [Formula: see text] in the range 4.5-7.5 (6.5-11.2) using the same variations of the background K-factor. Assuming that the unknown [Formula: see text] background K-factor is equal to the signal K-factor, this translates into an observed (expected) 95 [Formula: see text] CL upper limit on the Higgs boson total width of 22.7 (33.0) MeV.

Ezrin tunes T-cell activation by controlling Dlg1 and microtubule positioning at the immunological synapse

PubMed Central

Lasserre, Rémi; Charrin, Stéphanie; Cuche, Céline; Danckaert, Anne; Thoulouze, Maria-Isabel; de Chaumont, Fabrice; Duong, Tarn; Perrault, Nathalie; Varin-Blank, Nadine; Olivo-Marin, Jean-Christophe; Etienne-Manneville, Sandrine; Arpin, Monique; Di Bartolo, Vincenzo; Alcover, Andrés

2010-01-01

T-cell receptor (TCR) signalling is triggered and tuned at immunological synapses by the generation of signalling complexes that associate into dynamic microclusters. Microcluster movement is necessary to tune TCR signalling, but the molecular mechanism involved remains poorly known. We show here that the membrane-microfilament linker ezrin has an important function in microcluster dynamics and in TCR signalling through its ability to set the microtubule network organization at the immunological synapse. Importantly, ezrin and microtubules are important to down-regulate signalling events leading to Erk1/2 activation. In addition, ezrin is required for appropriate NF-AT activation through p38 MAP kinase. Our data strongly support the notion that ezrin regulates immune synapse architecture and T-cell activation through its interaction with the scaffold protein Dlg1. These results uncover a crucial function for ezrin, Dlg1 and microtubules in the organization of the immune synapse and TCR signal down-regulation. Moreover, they underscore the importance of ezrin and Dlg1 in the regulation of NF-AT activation through p38. PMID:20551903

N-ras couples antigen receptor signaling to Eomesodermin and to functional CD8+ T cell memory but not to effector differentiation

PubMed Central

Iborra, Salvador; Ramos, Manuel; Arana, David M.; Lázaro, Silvia; Aguilar, Francisco; Santos, Eugenio; López, Daniel

2013-01-01

Signals from the TCR that specifically contribute to effector versus memory CD8+ T cell differentiation are poorly understood. Using mice and adoptively transferred T lymphocytes lacking the small GTPase N-ras, we found that N-ras–deficient CD8+ T cells differentiate efficiently into antiviral primary effectors but have a severe defect in generating protective memory cells. This defect was rescued, although only partly, by rapamycin-mediated inhibition of mammalian target of rapamycin (mTOR) in vivo. The memory defect correlated with a marked impairment in vitro and in vivo of the antigen-mediated early induction of T-box transcription factor Eomesodermin (Eomes), whereas T-bet was unaffected. Besides N-ras, early Eomes induction in vitro required phosphoinositide 3-kinase (PI3K)–AKT but not extracellular signal-regulated kinase (ERK) activation, and it was largely insensitive to rapamycin. Consistent with N-ras coupling Eomes to T cell memory, retrovirally enforced expression of Eomes in N-ras–deficient CD8+ T cells effectively rescued their memory differentiation. Thus, our study identifies a critical role for N-ras as a TCR-proximal regulator of Eomes for early determination of the CD8+ T cell memory fate. PMID:23776078

The Cish SH2 domain is essential for PLC-γ1 regulation in TCR stimulated CD8+ T cells.

PubMed

Guittard, Geoffrey; Dios-Esponera, Ana; Palmer, Douglas C; Akpan, Itoro; Barr, Valarie A; Manna, Asit; Restifo, Nicholas P; Samelson, Lawrence E

2018-03-28

Cish, participates within a multi-molecular E3 ubiquitin ligase complex, which ubiquitinates target proteins. It has an inhibitory effect on T cell activation mediated by PLC-γ1 regulation, and it functions as a potent checkpoint in CD8 + T cell tumor immunotherapy. To study the structural and functional relationships between Cish and PLC-γ1 during CD8 + T cell activation, we tested mutants of the Cish-SH2 (R107K) and D/BC (L222Q, C226Q) domains. We confirmed that Cish-SH2-specific binding was essential for PLC-γ1 ubiquitination and degradation. This domain was essential for the Cish-mediated inhibition of Ca 2+ release upon TCR stimulation. No effect on inhibition of cytokine release was observed with SH2 or D/BC mutants, although the absence of Cish led to an increased release of IFN-γ and TNF-α. Using imaging we showed that Cish was expressed mostly in the cytoplasm and we did not see any Cish clustering at the plasma membrane upon stimulation. We conclude that the Cish-SH2 domain is essential for PLC-γ1 regulation in TCR-stimulated CD8 + T cells.

Phosphatase CD45 Both Positively and Negatively Regulates T Cell Receptor Phosphorylation in Reconstituted Membrane Protein Clusters*♦

PubMed Central

Furlan, Gabriela; Minowa, Takashi; Hanagata, Nobutaka; Kataoka-Hamai, Chiho; Kaizuka, Yoshihisa

2014-01-01

T cell receptor (TCR) phosphorylation requires the kinase Lck and phosphatase CD45. CD45 activates Lck by dephosphorylating an inhibitory tyrosine of Lck to relieve autoinhibition. However, CD45 also dephosphorylates the TCR, and the spatial exclusion of CD45 from TCR clustering in the plasma membrane appears to attenuate this negative effect of CD45. To further investigate the role of CD45 in signal initiation, we reconstituted membrane TCR clusters in vitro on supported lipid bilayers. Fluorescence microscopy of single clusters showed that incorporation of CD45 enhanced phosphorylation of TCR clusters, but only when Lck co-clustered with TCR. We found that clustered Lck autophosphorylated the inhibitory tyrosine and thus could be activated by CD45, whereas diffusive Lck molecules did not. In the TCR-Lck clusters and at low CD45 density, we speculate that the effect of Lck activation may overcome dephosphorylation of TCR, resulting in a net positive regulation. The CD45 density in physiological TCR clusters is also low because of the exclusion of CD45. Thus, we propose that the spatial organization of TCR/Lck/CD45 in T cell membranes is important not only for modulating the negative role of CD45 but also for creating conditions in which CD45 has a positive role in signal initiation. PMID:25128530

TCR-contacting residues orientation and HLA-DRβ* binding preference determine long-lasting protective immunity against malaria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alba, Martha P.; Suarez, Carlos F.; Universidad del Rosario, Bogotá D. C.

Fully-protective, long-lasting, immunological (FPLLI) memory against Plasmodium falciparum malaria regarding immune protection-inducing protein structures (IMPIPS) vaccinated into monkeys previously challenged and re-challenged 60 days later with a lethal Aotus monkey-adapted P. falciparum strain was found to be associated with preferential high binding capacity to HLA-DRβ1* allelic molecules of the major histocompatibility class II (MHC-II), rather than HLA-DRβ3*, β4*, β5* alleles. Complete PPII{sub L} 3D structure, a longer distance (26.5 Å ± 1.5 Å) between residues perfectly fitting into HLA-DRβ1*PBR pockets 1 and 9, a gauche{sup −} rotamer orientation in p8 TCR-contacting polar residue and a larger volume of polar p2 residues was also found. Thismore » data, in association with previously-described p3 and p7 apolar residues having gauche{sup +} orientation to form a perfect MHC-II-peptide-TCR complex, determines the stereo-electronic and topochemical characteristics associated with FPLLI immunological memory. - Highlights: • Stereo-electronic and topochemical rules associated with FPLLI immunological memory. • Presence of very high long-lasting antibody titres against Plasmodium falciparum Spz. • Protective memory induction associated with a binding capacity to HLA-DRβ1*. • gauche{sup −} rotamer orientation in p8 polar residue is related to is related to immunological memory.« less

Signal Strength-Based Global Navigation Satellite System Performance Assessment in the Space Service Volume

NASA Technical Reports Server (NTRS)

Welch, Bryan W.

2016-01-01

NASA is participating in the International Committee on Global Navigation Satellite Systems (GNSS) (ICG)'s efforts towards demonstrating the benefits to the space user in the Space Service Volume (SSV) when a multi-GNSS solution space approach is utilized. The ICG Working Group: Enhancement of GNSS Performance, New Services and Capabilities has started a three phase analysis initiative as an outcome of recommendations at the ICG-10 meeting, in preparation for the ICG-11 meeting. The second phase of that increasing complexity and fidelity analysis initiative is based on augmenting the Phase 1 pure geometrical approach with signal strength-based limitations to determine if access is valid. The second phase of analysis has been completed, and the results are documented in this paper.

SRC-like adaptor protein regulates B cell development and function.

PubMed

Dragone, Leonard L; Myers, Margaret D; White, Carmen; Sosinowski, Tomasz; Weiss, Arthur

2006-01-01

The avidity of BCRs and TCRs influences signal strength during processes of lymphocyte development. Avidity is determined by both the intrinsic affinity for Ag and surface levels of the Ag receptor. The Src-like adaptor protein (SLAP) is a regulator of TCR levels on thymocytes, and its deficiency alters thymocyte development. We hypothesized that SLAP, which is expressed in B cells, also is important in regulating BCR levels, signal strength, and B cell development. To test this hypothesis, we analyzed the B cell compartment in SLAP-deficient mice. We found increased splenic B cell numbers and decreased surface IgM levels on mature, splenic B cells deficient in SLAP. Immature bone marrow and splenic B cells from BCR-transgenic, SLAP-deficient mice were found to express higher surface levels of IgM. In contrast, mature splenic B cells from BCR-transgenic mice expressed decreased levels of surface BCR associated with decreased calcium flux and activation-induced markers, compared with controls. These data suggest that SLAP regulates BCR levels and signal strength during lymphocyte development.

Electroweak precision observables and Higgs-boson signal strengths in the Standard Model and beyond: present and future

DOE PAGES

de Blas, J.; Ciuchini, M.; Franco, E.; ...

2016-12-27

We present results from a state-of-the-art fit of electroweak precision observables and Higgs-boson signal-strength measurements performed using 7 and 8 TeV data from the Large Hadron Collider. Based on the HEPfit package, our study updates the traditional fit of electroweak precision observables and extends it to include Higgs-boson measurements. As a result we obtain constraints on new physics corrections to both electroweak observables and Higgs-boson couplings. We present the projected accuracy of the fit taking into account the expected sensitivities at future colliders.

Electroweak precision observables and Higgs-boson signal strengths in the Standard Model and beyond: present and future

DOE Office of Scientific and Technical Information (OSTI.GOV)

de Blas, J.; Ciuchini, M.; Franco, E.

We present results from a state-of-the-art fit of electroweak precision observables and Higgs-boson signal-strength measurements performed using 7 and 8 TeV data from the Large Hadron Collider. Based on the HEPfit package, our study updates the traditional fit of electroweak precision observables and extends it to include Higgs-boson measurements. As a result we obtain constraints on new physics corrections to both electroweak observables and Higgs-boson couplings. We present the projected accuracy of the fit taking into account the expected sensitivities at future colliders.

The Adenylate Cyclase Toxins of Bacillus anthracis and Bordetella pertussis Promote Th2 Cell Development by Shaping T Cell Antigen Receptor Signaling

PubMed Central

Rossi Paccani, Silvia; Benagiano, Marisa; Capitani, Nagaja; Zornetta, Irene; Ladant, Daniel; Montecucco, Cesare; D'Elios, Mario M.; Baldari, Cosima T.

2009-01-01

The adjuvanticity of bacterial adenylate cyclase toxins has been ascribed to their capacity, largely mediated by cAMP, to modulate APC activation, resulting in the expression of Th2–driving cytokines. On the other hand, cAMP has been demonstrated to induce a Th2 bias when present during T cell priming, suggesting that bacterial cAMP elevating toxins may directly affect the Th1/Th2 balance. Here we have investigated the effects on human CD4+ T cell differentiation of two adenylate cyclase toxins, Bacillus anthracis edema toxin (ET) and Bordetella pertussis CyaA, which differ in structure, mode of cell entry, and subcellular localization. We show that low concentrations of ET and CyaA, but not of their genetically detoxified adenylate cyclase defective counterparts, potently promote Th2 cell differentiation by inducing expression of the master Th2 transcription factors, c-maf and GATA-3. We also present evidence that the Th2–polarizing concentrations of ET and CyaA selectively inhibit TCR–dependent activation of Akt1, which is required for Th1 cell differentiation, while enhancing the activation of two TCR–signaling mediators, Vav1 and p38, implicated in Th2 cell differentiation. This is at variance from the immunosuppressive toxin concentrations, which interfere with the earliest step in TCR signaling, activation of the tyrosine kinase Lck, resulting in impaired CD3ζ phosphorylation and inhibition of TCR coupling to ZAP-70 and Erk activation. These results demonstrate that, notwithstanding their differences in their intracellular localization, which result in focalized cAMP production, both toxins directly affect the Th1/Th2 balance by interfering with the same steps in TCR signaling, and suggest that their adjuvanticity is likely to result from their combined effects on APC and CD4+ T cells. Furthermore, our results strongly support the key role of cAMP in the adjuvanticity of these toxins. PMID:19266022

Post-exercise cold water immersion attenuates acute anabolic signalling and long-term adaptations in muscle to strength training

PubMed Central

Roberts, Llion A; Raastad, Truls; Markworth, James F; Figueiredo, Vandre C; Egner, Ingrid M; Shield, Anthony; Cameron-Smith, David; Coombes, Jeff S; Peake, Jonathan M

2015-01-01

two treatments on hypertrophy signalling pathways and satellite cell activity in skeletal muscle after acute strength exercise. Cold water immersion attenuated long term gains in muscle mass and strength. It also blunted the activation of key proteins and satellite cells in skeletal muscle up to 2 days after strength exercise. Individuals who use strength training to improve athletic performance, recover from injury or maintain their health should therefore reconsider whether to use cold water immersion as an adjuvant to their training. PMID:26174323

The impact of higher-order aberrations on the strength of directional signals produced by accommodative microfluctuations

PubMed Central

Metlapally, Sangeetha; Tong, Jianliang L.; Tahir, Humza J.; Schor, Clifton M.

2014-01-01

It has been proposed that the accommodation system could perform contrast discrimination between the two dioptric extremes of accommodative microfluctuations to extract directional signals for reflex accommodation. Higher-order aberrations (HOAs) may have a significant influence on the strength of these contrast signals. Our goal was to compute the effect HOAs may have on contrast signals for stimuli within the upper defocus limit by comparing computed microcontrast fluctuations with psychophysical contrast increment thresholds (Bradley & Ohzawa, 1986). Wavefront aberrations were measured while subjects viewed a Maltese spoke stimulus monocularly. Computations were performed for accommodation or disaccommodation stimuli from a 3 Diopter (D) baseline. Microfluctuations were estimated from the standard deviation of the wavefronts over time at baseline. Through-focus Modulation Transfer, optical contrast increments (ΔC), and Weber fractions (ΔC/C) were derived from point spread functions computed from the wavefronts at baseline for 2 and 4 cycles per degree (cpd) components, with and without HOAs. The ΔCs thus computed from the wavefronts were compared with psychophysical contrast increment threshold data. Microfluctuations are potentially useful for extracting directional information for defocus values within 3 D, where contrast increments for the 2 or 4 cpd components exceed psychophysical thresholds. HOAs largely reduce contrast signals produced by microfluctuations, depending on the mean focus error, and their magnitude in individual subjects, and they may shrink the effective stimulus range for reflex accommodation. The upper defocus limit could therefore be constrained by discrimination of microcontrast fluctuations. PMID:25342542

Core Fucosylation on T Cells, Required for Activation of T-Cell Receptor Signaling and Induction of Colitis in Mice, Is Increased in Patients With Inflammatory Bowel Disease.

PubMed

Fujii, Hironobu; Shinzaki, Shinichiro; Iijima, Hideki; Wakamatsu, Kana; Iwamoto, Chizuru; Sobajima, Tomoaki; Kuwahara, Ryusuke; Hiyama, Satoshi; Hayashi, Yoshito; Takamatsu, Shinji; Uozumi, Naofumi; Kamada, Yoshihiro; Tsujii, Masahiko; Taniguchi, Naoyuki; Takehara, Tetsuo; Miyoshi, Eiji

2016-06-01

(-/-) mice did not signal properly after activation and were not transported to lipid rafts. Core fucosylation of the TCR is required for T-cell signaling and production of inflammatory cytokines and induction of colitis in mice. Levels of TCR core fucosylation are increased on T cells from intestinal tissues of patients with IBD; this process might be blocked as a therapeutic strategy. Copyright © 2016 AGA Institute. Published by Elsevier Inc. All rights reserved.

Dominant role of antigen dose in CD4+Foxp3+ regulatory T cell induction and expansion1

PubMed Central

Turner, Michael S.; Kane, Lawrence P.; Morel, Penelope A.

2009-01-01

The definitions of tolerogenic vs. immunogenic dendritic cells (DC) remain controversial. Immature DC have been shown to induce T regulatory cells (Treg) specific for foreign and allo-antigens. However, we have previously reported that mature DC (G4DC) prevented the onset of autoimmune diabetes whereas immature DC (GMDC) were therapeutically ineffective. In this study, islet-specific CD4+ T cells from BDC2.5 TCR Tg mice were stimulated, in the absence of exogenous cytokine, with GMDC or G4DC pulsed with high- or low-affinity antigenic peptides and examined for Treg induction. Both GMDC and G4DC presenting low peptide doses induced weak TCR signaling via the Akt/mTOR pathway, resulting in significant expansion of Foxp3+ Treg. Furthermore, unpulsed G4DC, but not GMDC, also induced Treg. High peptide doses induced strong Akt/mTOR signaling and favored the expansion of Foxp3neg Th cells. The inverse correlation of Foxp3 and Akt/mTOR signaling was also observed in DO11.10 and OT-II TCR-Tg T cells and was recapitulated with anti-CD3/CD28 stimulation in the absence of DC. IL-6 production in these cultures correlated positively with antigen dose and inversely with Treg expansion. Studies with T cells or DC from IL-6−/− mice revealed that IL-6 production by T cells was more important in the inhibition of Treg induction at low antigen doses. These studies indicate that strength of Akt/mTOR signaling, a critical T cell intrinsic determinant for Treg vs Th induction, can be controlled by adjusting the dose of antigenic peptide. Furthermore, this operates in a dominant fashion over DC phenotype and cytokine production. PMID:19801514

Cooperative ethylene receptor signaling

PubMed Central

Liu, Qian; Wen, Chi-Kuang

2012-01-01

The gaseous plant hormone ethylene is perceived by a family of five ethylene receptor members in the dicotyledonous model plant Arabidopsis. Genetic and biochemical studies suggest that the ethylene response is suppressed by ethylene receptor complexes, but the biochemical nature of the receptor signal is unknown. Without appropriate biochemical measures to trace the ethylene receptor signal and quantify the signal strength, the biological significance of the modulation of ethylene responses by multiple ethylene receptors has yet to be fully addressed. Nevertheless, the ethylene receptor signal strength can be reflected by degrees in alteration of various ethylene response phenotypes and in expression levels of ethylene-inducible genes. This mini-review highlights studies that have advanced our understanding of cooperative ethylene receptor signaling. PMID:22827938

Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4+ T Cell Proliferation.

PubMed

Celada, Lindsay J; Rotsinger, Joseph E; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene; Drake, Wonder P

2017-01-01

Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4 + T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4 + T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1 + CD4 + T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P < 0.001). There was a negative correlation with PD-1 expression and proliferative capacity (r = -0.70, P < 0.001). Expression of key mediators of cell cycle progression, including PI3K and AKT, were significantly decreased. Gene and protein expression levels reverted to healthy control levels after PD-1 pathway blockade. Reduction in sarcoidosis CD4 + T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression.

Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4+ T Cell Proliferation

PubMed Central

Celada, Lindsay J.; Rotsinger, Joseph E.; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene

2017-01-01

Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4+ T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4+ T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1+ CD4+ T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P < 0.001). There was a negative correlation with PD-1 expression and proliferative capacity (r = −0.70, P < 0.001). Expression of key mediators of cell cycle progression, including PI3K and AKT, were significantly decreased. Gene and protein expression levels reverted to healthy control levels after PD-1 pathway blockade. Reduction in sarcoidosis CD4+ T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression. PMID:27564547

Cellular Notch responsiveness is defined by phosphoinositide 3-kinase-dependent signals

PubMed Central

Mckenzie, Grahame; Ward, George; Stallwood, Yvette; Briend, Emmanuel; Papadia, Sofia; Lennard, Andrew; Turner, Martin; Champion, Brian; Hardingham, Giles E

2006-01-01

Background Notch plays a wide-ranging role in controlling cell fate, differentiation and development. The PI3K-Akt pathway is a similarly conserved signalling pathway which regulates processes such as differentiation, proliferation and survival. Mice with disrupted Notch and PI3K signalling show phenotypic similarities during haematopoietic cell development, suggesting functional interaction between these pathways. Results We show that cellular responsiveness to Notch signals depends on the activity of the PI3K-Akt pathway in cells as diverse as CHO cells, primary T-cells and hippocampal neurons. Induction of the endogenous PI3K-Akt pathway in CHO cells (by the insulin pathway), in T-cells (via TCR activation) or in neurons (via TrKB activation) potentiates Notch-dependent responses. We propose that the PI3K-Akt pathway exerts its influence on Notch primarily via inhibition of GSK3-beta, a kinase known to phosphorylate and regulate Notch signals. Conclusion The PI3K-Akt pathway acts as a "gain control" for Notch signal responses. Since physiological levels of intracellular Notch are often low, coincidence with PI3K-activation may be crucial for induction of Notch-dependent responses. PMID:16507111

A mechanism for expansion of regulatory T-cell repertoire and its role in self-tolerance.

PubMed

Feng, Yongqiang; van der Veeken, Joris; Shugay, Mikhail; Putintseva, Ekaterina V; Osmanbeyoglu, Hatice U; Dikiy, Stanislav; Hoyos, Beatrice E; Moltedo, Bruno; Hemmers, Saskia; Treuting, Piper; Leslie, Christina S; Chudakov, Dmitriy M; Rudensky, Alexander Y

2015-12-03

T-cell receptor (TCR) signalling has a key role in determining T-cell fate. Precursor cells expressing TCRs within a certain low-affinity range for complexes of self-peptide and major histocompatibility complex (MHC) undergo positive selection and differentiate into naive T cells expressing a highly diverse self-MHC-restricted TCR repertoire. In contrast, precursors displaying TCRs with a high affinity for 'self' are either eliminated through TCR-agonist-induced apoptosis (negative selection) or restrained by regulatory T (Treg) cells, whose differentiation and function are controlled by the X-chromosome-encoded transcription factor Foxp3 (reviewed in ref. 2). Foxp3 is expressed in a fraction of self-reactive T cells that escape negative selection in response to agonist-driven TCR signals combined with interleukin 2 (IL-2) receptor signalling. In addition to Treg cells, TCR-agonist-driven selection results in the generation of several other specialized T-cell lineages such as natural killer T cells and innate mucosal-associated invariant T cells. Although the latter exhibit a restricted TCR repertoire, Treg cells display a highly diverse collection of TCRs. Here we explore in mice whether a specialized mechanism enables agonist-driven selection of Treg cells with a diverse TCR repertoire, and the importance this holds for self-tolerance. We show that the intronic Foxp3 enhancer conserved noncoding sequence 3 (CNS3) acts as an epigenetic switch that confers a poised state to the Foxp3 promoter in precursor cells to make Treg cell lineage commitment responsive to a broad range of TCR stimuli, particularly to suboptimal ones. CNS3-dependent expansion of the TCR repertoire enables Treg cells to control self-reactive T cells effectively, especially when thymic negative selection is genetically impaired. Our findings highlight the complementary roles of these two main mechanisms of self-tolerance.

First-in-class inhibitor of the T cell receptor for the treatment of autoimmune diseases.

PubMed

Borroto, Aldo; Reyes-Garau, Diana; Jiménez, M Angeles; Carrasco, Esther; Moreno, Beatriz; Martínez-Pasamar, Sara; Cortés, José R; Perona, Almudena; Abia, David; Blanco, Soledad; Fuentes, Manuel; Arellano, Irene; Lobo, Juan; Heidarieh, Haleh; Rueda, Javier; Esteve, Pilar; Cibrián, Danay; Martinez-Riaño, Ana; Mendoza, Pilar; Prieto, Cristina; Calleja, Enrique; Oeste, Clara L; Orfao, Alberto; Fresno, Manuel; Sánchez-Madrid, Francisco; Alcamí, Antonio; Bovolenta, Paola; Martín, Pilar; Villoslada, Pablo; Morreale, Antonio; Messeguer, Angel; Alarcon, Balbino

2016-12-21

Modulating T cell activation is critical for treating autoimmune diseases but requires avoiding concomitant opportunistic infections. Antigen binding to the T cell receptor (TCR) triggers the recruitment of the cytosolic adaptor protein Nck to a proline-rich sequence in the cytoplasmic tail of the TCR's CD3ε subunit. Through virtual screening and using combinatorial chemistry, we have generated an orally available, low-molecular weight inhibitor of the TCR-Nck interaction that selectively inhibits TCR-triggered T cell activation with an IC 50 (median inhibitory concentration) ~1 nM. By modulating TCR signaling, the inhibitor prevented the development of psoriasis and asthma and, furthermore, exerted a long-lasting therapeutic effect in a model of autoimmune encephalomyelitis. However, it did not prevent the generation of a protective memory response against a mouse pathogen, suggesting that the compound might not exert its effects through immunosuppression. These results suggest that inhibiting an immediate TCR signal has promise for treating a broad spectrum of human T cell-mediated autoimmune and inflammatory diseases. Copyright © 2016, American Association for the Advancement of Science.

A conserved αβ transmembrane interface forms the core of a compact T-cell receptor-CD3 structure within the membrane.

PubMed

Krshnan, Logesvaran; Park, Soohyung; Im, Wonpil; Call, Melissa J; Call, Matthew E

2016-10-25

The T-cell antigen receptor (TCR) is an assembly of eight type I single-pass membrane proteins that occupies a central position in adaptive immunity. Many TCR-triggering models invoke an alteration in receptor complex structure as the initiating event, but both the precise subunit organization and the pathway by which ligand-induced alterations are transferred to the cytoplasmic signaling domains are unknown. Here, we show that the receptor complex transmembrane (TM) domains form an intimately associated eight-helix bundle organized by a specific interhelical TCR TM interface. The salient features of this core structure are absolutely conserved between αβ and γδ TCR sequences and throughout vertebrate evolution, and mutations at key interface residues caused defects in the formation of stable TCRαβ:CD3δε:CD3γε:ζζ complexes. These findings demonstrate that the eight TCR-CD3 subunits form a compact and precisely organized structure within the membrane and provide a structural basis for further investigation of conformationally regulated models of transbilayer TCR signaling.

Wide-scale quantitative phosphoproteomic analysis reveals that cold treatment of T cells closely mimics soluble antibody stimulation

PubMed Central

Ji, Qinqin; Salomon, Arthur R.

2015-01-01

The activation of T-lymphocytes through antigen-mediated T-cell receptor (TCR) clustering is vital in regulating the adaptive-immune response. Although T cell receptor signaling has been extensively studied, the fundamental mechanisms for signal initiation are not fully understood. Reduced temperature initiated some of the hallmarks of TCR signaling such as increased phosphorylation and activation on ERK and calcium release from the endoplasmic reticulum as well as coalesce T-cell membrane microdomains. The precise mechanism of TCR signaling initiation due to temperature change remains obscure. One critical question is whether signaling initiated by cold treatment of T cells differs from signaling initiated by crosslinking of the T cell receptor. To address this uncertainty, a wide-scale, quantitative mass spectrometry-based phosphoproteomic analysis was performed on T cells stimulated either by temperature shift or through crosslinking of the TCR. Careful statistical comparison between the two stimulations revealed a striking level of identity between the subset of 339 sites that changed significantly with both stimulations. This study demonstrates for the first time, at unprecedented detail, that T cell cold treatment was sufficient to initiate signaling patterns nearly identical to soluble antibody stimulation, shedding new light on the mechanism of activation of these critically important immune cells. PMID:25839225

Received signal strength and local terrain profile data for radio network planning and optimization at GSM frequency bands.

PubMed

Popoola, Segun I; Atayero, Aderemi A; Faruk, Nasir

2018-02-01

The behaviour of radio wave signals in a wireless channel depends on the local terrain profile of the propagation environments. In view of this, Received Signal Strength (RSS) of transmitted signals are measured at different points in space for radio network planning and optimization. However, these important data are often not publicly available for wireless channel characterization and propagation model development. In this data article, RSS data of a commercial base station operating at 900 and 1800 MHz were measured along three different routes of Lagos-Badagry Highway, Nigeria. In addition, local terrain profile data of the study area (terrain elevation, clutter height, altitude, and the distance of the mobile station from the base station) are extracted from Digital Terrain Map (DTM) to account for the unique environmental features. Statistical analyses and probability distributions of the RSS data are presented in tables and graphs. Furthermore, the degree of correlations (and the corresponding significance) between the RSS and the local terrain parameters were computed and analyzed for proper interpretations. The data provided in this article will help radio network engineers to: predict signal path loss; estimate radio coverage; efficiently reuse limited frequencies; avoid interferences; optimize handover; and adjust transmitted power level.

Pulsed Electromagnetic Fields Improve Bone Microstructure and Strength in Ovariectomized Rats through a Wnt/Lrp5/β-Catenin Signaling-Associated Mechanism

PubMed Central

Cai, Jing; Wu, Yan; Xie, Kangning; Wu, Xiaoming; Tang, Chi; Liu, Juan; Guo, Wei; Shen, Guanghao; Luo, Erping

2013-01-01

Growing evidence has demonstrated that pulsed electromagnetic field (PEMF), as an alternative noninvasive method, could promote remarkable in vivo and in vitro osteogenesis. However, the exact mechanism of PEMF on osteopenia/osteoporosis is still poorly understood, which further limits the extensive clinical application of PEMF. In the present study, the efficiency of PEMF on osteoporotic bone microarchitecture and bone quality together with its associated signaling pathway mechanisms was systematically investigated in ovariectomized (OVX) rats. Thirty rats were equally assigned to the Control, OVX and OVX+PEMF groups. The OVX+PEMF group was subjected to daily 8-hour PEMF exposure with 15 Hz, 2.4 mT (peak value). After 10 weeks, the OVX+PEMF group exhibited significantly improved bone mass and bone architecture, evidenced by increased BMD, Tb.N, Tb.Th and BV/TV, and suppressed Tb.Sp and SMI levels in the MicroCT analysis. Three-point bending test suggests that PEMF attenuated the biomechanical strength deterioration of the OVX rat femora, evidenced by increased maximum load and elastic modulus. RT-PCR analysis demonstrated that PEMF exposure significantly promoted the overall gene expressions of Wnt1, LRP5 and β-catenin in the canonical Wnt signaling, but did not exhibit obvious impact on either RANKL or RANK gene expressions. Together, our present findings highlight that PEMF attenuated OVX-induced deterioration of bone microarchitecture and strength in rats by promoting the activation of Wnt/LRP5/β-catenin signaling rather than by inhibiting RANKL-RANK signaling. This study enriches our basic knowledge to the osteogenetic activity of PEMF, and may lead to more efficient and scientific clinical application of PEMF in inhibiting osteopenia/osteoporosis. PMID:24244491

BCR CDR3 length distributions differ between blood and spleen and between old and young patients, and TCR distributions can be used to detect myelodysplastic syndrome

NASA Astrophysics Data System (ADS)

Pickman, Yishai; Dunn-Walters, Deborah; Mehr, Ramit

2013-10-01

Complementarity-determining region 3 (CDR3) is the most hyper-variable region in B cell receptor (BCR) and T cell receptor (TCR) genes, and the most critical structure in antigen recognition and thereby in determining the fates of developing and responding lymphocytes. There are millions of different TCR Vβ chain or BCR heavy chain CDR3 sequences in human blood. Even now, when high-throughput sequencing becomes widely used, CDR3 length distributions (also called spectratypes) are still a much quicker and cheaper method of assessing repertoire diversity. However, distribution complexity and the large amount of information per sample (e.g. 32 distributions of the TCRα chain, and 24 of TCRβ) calls for the use of machine learning tools for full exploration. We have examined the ability of supervised machine learning, which uses computational models to find hidden patterns in predefined biological groups, to analyze CDR3 length distributions from various sources, and distinguish between experimental groups. We found that (a) splenic BCR CDR3 length distributions are characterized by low standard deviations and few local maxima, compared to peripheral blood distributions; (b) healthy elderly people's BCR CDR3 length distributions can be distinguished from those of the young; and (c) a machine learning model based on TCR CDR3 distribution features can detect myelodysplastic syndrome with approximately 93% accuracy. Overall, we demonstrate that using supervised machine learning methods can contribute to our understanding of lymphocyte repertoire diversity.

Weighted least squares techniques for improved received signal strength based localization.

PubMed

Tarrío, Paula; Bernardos, Ana M; Casar, José R

2011-01-01

The practical deployment of wireless positioning systems requires minimizing the calibration procedures while improving the location estimation accuracy. Received Signal Strength localization techniques using propagation channel models are the simplest alternative, but they are usually designed under the assumption that the radio propagation model is to be perfectly characterized a priori. In practice, this assumption does not hold and the localization results are affected by the inaccuracies of the theoretical, roughly calibrated or just imperfect channel models used to compute location. In this paper, we propose the use of weighted multilateration techniques to gain robustness with respect to these inaccuracies, reducing the dependency of having an optimal channel model. In particular, we propose two weighted least squares techniques based on the standard hyperbolic and circular positioning algorithms that specifically consider the accuracies of the different measurements to obtain a better estimation of the position. These techniques are compared to the standard hyperbolic and circular positioning techniques through both numerical simulations and an exhaustive set of real experiments on different types of wireless networks (a wireless sensor network, a WiFi network and a Bluetooth network). The algorithms not only produce better localization results with a very limited overhead in terms of computational cost but also achieve a greater robustness to inaccuracies in channel modeling.

Elevated stress hormone diminishes the strength of female preferences for acoustic signals in the green treefrog.

PubMed

Davis, A Gabriell; Leary, Christopher J

2015-03-01

Mate selection can be stressful; time spent searching for mates can increase predation risk and/or decrease food consumption, resulting in elevated stress hormone levels. Both high predation risk and low food availability are often associated with increased variation in mate choice by females, but it is not clear whether stress hormone levels contribute to such variation in female behavior. We examined how the stress hormone corticosterone (CORT) affects female preferences for acoustic signals in the green treefrog, Hyla cinerea. Specifically, we assessed whether CORT administration affects female preferences for call rate - an acoustic feature that is typically under directional selection via mate choice by females in most anurans and other species that communicate using acoustic signals. Using a dual speaker playback paradigm, we show that females that were administered higher doses of CORT were less likely to choose male advertisement calls broadcast at high rates. Neither CORT dose nor level was related to the latency of female phonotactic responses, suggesting that elevated CORT does not influence the motivation to mate. Results were also not related to circulating sex steroids (i.e., progesterone, androgens or estradiol) that have traditionally been the focus of studies examining the hormonal basis for variation in female mate choice. Our results thus indicate that elevated CORT levels decrease the strength of female preferences for acoustic signals. Copyright © 2015 Elsevier Inc. All rights reserved.

Evaluation of Intracellular Signaling Downstream Chimeric Antigen Receptors

PubMed Central

Karlsson, Hannah; Svensson, Emma; Gigg, Camilla; Jarvius, Malin; Olsson-Strömberg, Ulla; Savoldo, Barbara; Dotti, Gianpietro; Loskog, Angelica

2015-01-01

CD19-targeting CAR T cells have shown potency in clinical trials targeting B cell leukemia. Although mainly second generation (2G) CARs carrying CD28 or 4-1BB have been investigated in patients, preclinical studies suggest that third generation (3G) CARs with both CD28 and 4-1BB have enhanced capacity. However, little is known about the intracellular signaling pathways downstream of CARs. In the present work, we have analyzed the signaling capacity post antigen stimulation in both 2G and 3G CARs. 3G CAR T cells expanded better than 2G CAR T cells upon repeated stimulation with IL-2 and autologous B cells. An antigen-driven accumulation of CAR+ cells was evident post antigen stimulation. The cytotoxicity of both 2G and 3G CAR T cells was maintained by repeated stimulation. The phosphorylation status of intracellular signaling proteins post antigen stimulation showed that 3G CAR T cells had a higher activation status than 2G. Several proteins involved in signaling downstream the TCR were activated, as were proteins involved in the cell cycle, cell adhesion and exocytosis. In conclusion, 3G CAR T cells had a higher degree of intracellular signaling activity than 2G CARs which may explain the increased proliferative capacity seen in 3G CAR T cells. The study also indicates that there may be other signaling pathways to consider when designing or evaluating new generations of CARs. PMID:26700307

Effect of Pulse Shape on Spall Strength

NASA Astrophysics Data System (ADS)

Smirnov, V. I.; Petrov, Yu. V.

2018-03-01

This paper analyzes the effect of the time-dependent shape of a load pulse on the spall strength of materials. Within the framework of a classical one-dimensional scheme, triangular pulses with signal rise and decay portions and with no signal rise portions considered. Calculation results for the threshold characteristics of fracture for rail steel are given. The possibility of optimization of fracture by selecting a loading time with the use of an introduced characteristic of dynamic strength (pulse fracture capacity) is demonstrated. The study is carried out using a structure-time fracture criterion.

Indoor localization using FM radio and DTMB signals

NASA Astrophysics Data System (ADS)

Wu, H.; Wang, Q.; Zhao, Y.; Ma, X.; Yang, M.; Liu, B.; Tang, R.; Xu, X.

2016-07-01

Indoor localization systems based on Wi-Fi signal strength fingerprinting techniques are widely used in office buildings. However, a general problem of these systems pertains to Wi-Fi signal degradation due to the environmental factors. And also, these systems cannot be used in the environments not covered with Wi-Fi signals or the environments with only a single Wi-Fi access point. In this paper, a new indoor location fingerprinting system using both FM radio and Digital Television Terrestrial Multimedia Broadcasting (DTMB) signals is proposed. First, the indoor location fingerprinting using FM radio and DTMB signals is theoretically analyzed to confirm its feasibility. Then, a specially designed combined strength fingerprinting location algorithm is proposed for the location system, which is achieved on the USRP2 platform. Finally, the system is tested in a typical indoor environment. The theoretical analysis and the tests show that the indoor location fingerprinting system using FM radio and DTMB signals has a similar localization accuracy to the Wi-Fi signal strength fingerprinting location system, while it has a wider coverage area, a lower maintenance cost, and more stable signal strength, which makes it a practical indoor positioning method.

Development of an RNA-based kit for easy generation of TCR-engineered lymphocytes to control T-cell assay performance.

PubMed

Bidmon, Nicole; Kind, Sonja; Welters, Marij J P; Joseph-Pietras, Deborah; Laske, Karoline; Maurer, Dominik; Hadrup, Sine Reker; Schreibelt, Gerty; Rae, Richard; Sahin, Ugur; Gouttefangeas, Cécile; Britten, Cedrik M; van der Burg, Sjoerd H

2018-07-01

Cell-based assays to monitor antigen-specific T-cell responses are characterized by their high complexity and should be conducted under controlled conditions to lower multiple possible sources of assay variation. However, the lack of standard reagents makes it difficult to directly compare results generated in one lab over time and across institutions. Therefore TCR-engineered reference samples (TERS) that contain a defined number of antigen-specific T cells and continuously deliver stable results are urgently needed. We successfully established a simple and robust TERS technology that constitutes a useful tool to overcome this issue for commonly used T-cell immuno-assays. To enable users to generate large-scale TERS, on-site using the most commonly used electroporation (EP) devices, an RNA-based kit approach, providing stable TCR mRNA and an optimized manufacturing protocol were established. In preparation for the release of this immuno-control kit, we established optimal EP conditions on six devices and initiated an extended RNA stability study. Furthermore, we coordinated on-site production of TERS with 4 participants. Finally, a proficiency panel was organized to test the unsupervised production of TERS at different laboratories using the kit approach. The results obtained show the feasibility and robustness of the kit approach for versatile in-house production of cellular control samples. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

The Adaptor Protein SAP Directly Associates with CD3ζ Chain and Regulates T Cell Receptor Signaling

PubMed Central

Proust, Richard; Bertoglio, Jacques; Gesbert, Franck

2012-01-01

Mutations altering the gene encoding the SLAM associated protein (SAP) are responsible for the X-linked lymphoproliferative disease or XLP1. Its absence is correlated with a defective NKT cells development, a decrease in B cell functions and a reduced T cells and NK cells cytotoxic activities, thus leading to an immunodeficiency syndrome. SAP is a small 128 amino-acid long protein that is almost exclusively composed of an SH2 domain. It has been shown to interact with the CD150/SLAM family of receptors, and in a non-canonical manner with SH3 containing proteins such as Fyn, βPIX, PKCθ and Nck1. It would thus play the role of a minimal adaptor protein. It has been shown that SAP plays an important function in the activation of T cells through its interaction with the SLAM family of receptors. Therefore SAP defective T cells display a reduced activation of signaling events downstream of the TCR-CD3 complex triggering. In the present work, we evidence that SAP is a direct interactor of the CD3ζ chain. This direct interaction occurs through the first ITAM of CD3ζ, proximal to the membrane. Additionally, we show that, in the context of the TCR-CD3 signaling, an Sh-RNA mediated silencing of SAP is responsible for a decrease of several canonical T cell signaling pathways including Erk, Akt and PLCγ1 and to a reduced induction of IL-2 and IL-4 mRNA. Altogether, we show that SAP plays a central function in the T cell activation processes through a direct association with the CD3 complex. PMID:22912825

LAPTM5 promotes lysosomal degradation of intracellular CD3ζ but not of cell surface CD3ζ.

PubMed

Kawai, Yohei; Ouchida, Rika; Yamasaki, Sho; Dragone, Leonard; Tsubata, Takeshi; Wang, Ji-Yang

2014-07-01

The lysosomal protein LAPTM5 has been shown to negatively regulate cell surface T cell receptor (TCR) expression and T-cell activation by promoting CD3ζ degradation in lysosomes, but the mechanism remains largely unknown. Here we show that LAPTM5 promotes lysosomal translocation of intracellular CD3ζ but not of the cell surface CD3ζ associated with the mature TCR complex. Kinetic analysis of the subcellular localization of the newly synthesized CD3ζ suggests that LAPTM5 targets CD3ζ in the Golgi apparatus and promotes its lysosomal translocation. Consistently, a Golgi-localizing mutant CD3ζ can be transported to and degraded in the lysosome by LAPTM5. A CD3ζ YF mutant in which all six tyrosine residues in the immunoreceptor tyrosine-based activation motif are mutated to phenylalanines is degraded as efficiently as is wild type CD3ζ, further suggesting that TCR signaling-triggered tyrosine phosphorylation of CD3ζ is dispensable for LAPTM5-mediated degradation. Previously, Src-like adapter protein (SLAP) and E3 ubiquitin ligase c-Cbl have been shown to mediate the ubiquitination of CD3ζ in the internalized TCR complex and its subsequent lysosomal degradation. We show that LAPTM5 and SLAP/c-Cbl function in distinct genetic pathways to negatively regulate TCR expression. Collectively, these results suggest that CD3ζ can be degraded by two pathways: SLAP/c-Cbl, which targets internalized cell surface CD3ζ dependent on TCR signaling, and LAPTM5, which targets intracellular CD3ζ independent of TCR signaling.

Secure Cluster Head Sensor Elections Using Signal Strength Estimation and Ordered Transmissions

PubMed Central

Wang, Gicheol; Cho, Gihwan

2009-01-01

In clustered sensor networks, electing CHs (Cluster Heads) in a secure manner is very important because they collect data from sensors and send the aggregated data to the sink. If a compromised node is elected as a CH, it can illegally acquire data from all the members and even send forged data to the sink. Nevertheless, most of the existing CH election schemes have not treated the problem of the secure CH election. Recently, random value based protocols have been proposed to resolve the secure CH election problem. However, these schemes cannot prevent an attacker from suppressing its contribution for the change of CH election result and from selectively forwarding its contribution for the disagreement of CH election result. In this paper, we propose a modified random value scheme to prevent these disturbances. Our scheme dynamically adjusts the forwarding order of contributions and discards a received contribution when its signal strength is lower than the specified level to prevent these malicious actions. The simulation results have shown that our scheme effectively prevents attackers from changing and splitting an agreement of CH election result. Also, they have shown that our scheme is relatively energy-efficient than other schemes. PMID:22408550

Modeling alternative binding registers of a minimal immunogenic peptide on two class II major histocompatibility complex (MHC II) molecules predicts polarized T-cell receptor (TCR) contact positions.

PubMed

Murray, J S; Fois, S D S; Schountz, T; Ford, S R; Tawde, M D; Brown, J C; Siahaan, T J

2002-03-01

Several major histocompatibility complex class II (MHC II) complexes with known minimal immunogenic peptides have now been solved by X-ray crystallography. Specificity pockets within the MHC II binding groove provide distinct peptide contacts that influence peptide conformation and define the binding register within different allelic MHC II molecules. Altering peptide ligands with respect to the residues that contact the T-cell receptor (TCR) can drastically change the nature of the ensuing immune response. Here, we provide an example of how MHC II (I-A) molecules may indirectly effect TCR contacts with a peptide and drive functionally distinct immune responses. We modeled the same immunogenic 12-amino acid peptide into the binding grooves of two allelic MHC II molecules linked to distinct cytokine responses against the peptide. Surprisingly, the favored conformation of the peptide in each molecule was distinct with respect to the exposure of the N- or C-terminus of the peptide above the MHC II binding groove. T-cell clones derived from each allelic MHC II genotype were found to be allele-restricted with respect to the recognition of these N- vs. C-terminal residues on the bound peptide. Taken together, these data suggest that MHC II alleles may influence T-cell functions by restricting TCR access to specific residues of the I-A-bound peptide. Thus, these data are of significance to diseases that display genetic linkage to specific MHC II alleles, e.g. type 1 diabetes and rheumatoid arthritis.

Vitamin C and E supplementation alters protein signalling after a strength training session, but not muscle growth during 10 weeks of training

PubMed Central

Paulsen, G; Hamarsland, H; Cumming, K T; Johansen, R E; Hulmi, J J; Børsheim, E; Wiig, H; Garthe, I; Raastad, T

2014-01-01

This study investigated the effects of vitamin C and E supplementation on acute responses and adaptations to strength training. Thirty-two recreationally strength-trained men and women were randomly allocated to receive a vitamin C and E supplement (1000 mg day−1 and 235 mg day−1, respectively), or a placebo, for 10 weeks. During this period the participants’ training involved heavy-load resistance exercise four times per week. Muscle biopsies from m. vastus lateralis were collected, and 1 repetition maximum (1RM) and maximal isometric voluntary contraction force, body composition (dual-energy X-ray absorptiometry), and muscle cross-sectional area (magnetic resonance imaging) were measured before and after the intervention. Furthermore, the cellular responses to a single exercise session were assessed midway in the training period by measurements of muscle protein fractional synthetic rate and phosphorylation of several hypertrophic signalling proteins. Muscle biopsies were obtained from m. vastus lateralis twice before, and 100 and 150 min after, the exercise session (4 × 8RM, leg press and knee-extension). The supplementation did not affect the increase in muscle mass or the acute change in protein synthesis, but it hampered certain strength increases (biceps curl). Moreover, increased phosphorylation of p38 mitogen-activated protein kinase, Extracellular signal-regulated protein kinases 1 and 2 and p70S6 kinase after the exercise session was blunted by vitamin C and E supplementation. The total ubiquitination levels after the exercise session, however, were lower with vitamin C and E than placebo. We concluded that vitamin C and E supplementation interfered with the acute cellular response to heavy-load resistance exercise and demonstrated tentative long-term negative effects on adaptation to strength training. PMID:25384788

Vitamin C and E supplementation alters protein signalling after a strength training session, but not muscle growth during 10 weeks of training.

PubMed

Paulsen, G; Hamarsland, H; Cumming, K T; Johansen, R E; Hulmi, J J; Børsheim, E; Wiig, H; Garthe, I; Raastad, T

2014-12-15

This study investigated the effects of vitamin C and E supplementation on acute responses and adaptations to strength training. Thirty-two recreationally strength-trained men and women were randomly allocated to receive a vitamin C and E supplement (1000 mg day(-1) and 235 mg day(-1), respectively), or a placebo, for 10 weeks. During this period the participants' training involved heavy-load resistance exercise four times per week. Muscle biopsies from m. vastus lateralis were collected, and 1 repetition maximum (1RM) and maximal isometric voluntary contraction force, body composition (dual-energy X-ray absorptiometry), and muscle cross-sectional area (magnetic resonance imaging) were measured before and after the intervention. Furthermore, the cellular responses to a single exercise session were assessed midway in the training period by measurements of muscle protein fractional synthetic rate and phosphorylation of several hypertrophic signalling proteins. Muscle biopsies were obtained from m. vastus lateralis twice before, and 100 and 150 min after, the exercise session (4 × 8RM, leg press and knee-extension). The supplementation did not affect the increase in muscle mass or the acute change in protein synthesis, but it hampered certain strength increases (biceps curl). Moreover, increased phosphorylation of p38 mitogen-activated protein kinase, Extracellular signal-regulated protein kinases 1 and 2 and p70S6 kinase after the exercise session was blunted by vitamin C and E supplementation. The total ubiquitination levels after the exercise session, however, were lower with vitamin C and E than placebo. We concluded that vitamin C and E supplementation interfered with the acute cellular response to heavy-load resistance exercise and demonstrated tentative long-term negative effects on adaptation to strength training. © 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.

Intimate association of Thy-1 and the T-cell antigen receptor with the CD45 tyrosine phosphatase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Volarevic, S.; Burns, C.M.; Sussman, J.J.

1990-09-01

Immunoprecipitation of Thy-1 from Triton X-100 detergent lysates of surface-iodinated and chemically cross-linked T cells precipitated at least first major and discrete bands. Four of these bands were identified as Thy-1, CD45 (a trasmembrane tyrosine phosphatase), a major histocompatibility complex-encoded class I molecule, and {beta}{sub 2}-microglobulin. Similar analyses revealed that CD45 was coprecipitated from lysates of cross-linker-treated cells by antibodies to the T-cell antigen receptor (TCR). The same pattern of coprecipitated bands was observed when digitonin was used to lyse untreated cells. Immunoprecipitation of Thy-1 or the TCR from lysates of cross-linked T cells precipitated CD45 tyrosine phosphatase activity. Calculationsmore » based upon the amounts of coprecipitated enzymatic activity or TCR {zeta} chain indicate that a substantial fraction of Thy-1 and TCR complexes can be cross-linked to CD45. These data support a model in which the dependence of Thy-1 signaling on TCR coexpression is due to their common interaction with a tyrosine phosphatase and provide a possible structural basis for the influence of CD45 on TCR-mediated signaling.« less

T Cell Calcium Signaling Regulation by the Co-Receptor CD5

PubMed Central

Freitas, Claudia M. Tellez

2018-01-01

Calcium influx is critical for T cell effector function and fate. T cells are activated when T cell receptors (TCRs) engage peptides presented by antigen-presenting cells (APC), causing an increase of intracellular calcium (Ca2+) concentration. Co-receptors stabilize interactions between the TCR and its ligand, the peptide-major histocompatibility complex (pMHC), and enhance Ca2+ signaling and T cell activation. Conversely, some co-receptors can dampen Ca2+ signaling and inhibit T cell activation. Immune checkpoint therapies block inhibitory co-receptors, such as cytotoxic T-lymphocyte associated antigen 4 (CTLA-4) and programmed death 1 (PD-1), to increase T cell Ca2+ signaling and promote T cell survival. Similar to CTLA-4 and PD-1, the co-receptor CD5 has been known to act as a negative regulator of T cell activation and to alter Ca2+ signaling and T cell function. Though much is known about the role of CD5 in B cells, recent research has expanded our understanding of CD5 function in T cells. Here we review these recent findings and discuss how our improved understanding of CD5 Ca2+ signaling regulation could be useful for basic and clinical research. PMID:29701673

THEMIS, a new T cell specific protein important for late thymocyte development

PubMed Central

Lesourne, Renaud; Uehara, Shoji; Lee, Jan; Song, Ki-Duk; Li, LiQi; Pinkhasov, Julia; Zhang, Yongqing; Weng, Nan-Ping; Wildt, Kathryn F.; Wang, Lie; Bosselut, Remy; Love, Paul E.

2010-01-01

During positive selection, thymocytes transition through a stage during which T cell receptor (TCR) signaling controls CD4 versus CD8 lineage choice and subsequent maturation. Here, we describe a new T cell specific protein, THEMIS, that performs a distinct function during this stage. In Themis-/- mice, thymocyte selection was impaired and the number of transitional CD4+CD8int thymocytes as well as CD4 and CD8 single positive thymocytes was decreased. Remarkably, although no overt TCR-proximal signaling deficiencies were detected, Themis-/-CD4+CD8int thymocytes exhibited developmental defects consistent with attenuated signaling that were reversible by increased TCR stimulation. These results identify THEMIS as a critical component of the T cell developmental program and suggest that THEMIS functions to sustain and/or integrate signals required for proper lineage commitment and maturation. PMID:19597498

Weighted Least Squares Techniques for Improved Received Signal Strength Based Localization

PubMed Central

Tarrío, Paula; Bernardos, Ana M.; Casar, José R.

2011-01-01

The practical deployment of wireless positioning systems requires minimizing the calibration procedures while improving the location estimation accuracy. Received Signal Strength localization techniques using propagation channel models are the simplest alternative, but they are usually designed under the assumption that the radio propagation model is to be perfectly characterized a priori. In practice, this assumption does not hold and the localization results are affected by the inaccuracies of the theoretical, roughly calibrated or just imperfect channel models used to compute location. In this paper, we propose the use of weighted multilateration techniques to gain robustness with respect to these inaccuracies, reducing the dependency of having an optimal channel model. In particular, we propose two weighted least squares techniques based on the standard hyperbolic and circular positioning algorithms that specifically consider the accuracies of the different measurements to obtain a better estimation of the position. These techniques are compared to the standard hyperbolic and circular positioning techniques through both numerical simulations and an exhaustive set of real experiments on different types of wireless networks (a wireless sensor network, a WiFi network and a Bluetooth network). The algorithms not only produce better localization results with a very limited overhead in terms of computational cost but also achieve a greater robustness to inaccuracies in channel modeling. PMID:22164092

T Cell Development in Mice Lacking All T Cell Receptor ζ Family Members (ζ, η, and FcεRIγ)

PubMed Central

Shores, Elizabeth W.; Ono, Masao; Kawabe, Tsutomo; Sommers, Connie L.; Tran, Tom; Lui, Kin; Udey, Mark C.; Ravetch, Jeffrey; Love, Paul E.

1998-01-01

The ζ family includes ζ, η, and FcεRIγ (Fcγ). Dimers of the ζ family proteins function as signal transducing subunits of the T cell antigen receptor (TCR), the pre-TCR, and a subset of Fc receptors. In mice lacking ζ/η chains, T cell development is impaired, yet low numbers of CD4+ and CD8+ T cells develop. This finding suggests either that pre-TCR and TCR complexes lacking a ζ family dimer can promote T cell maturation, or that in the absence of ζ/η, Fcγ serves as a subunit in TCR complexes. To elucidate the role of ζ family dimers in T cell development, we generated mice lacking expression of all of these proteins and compared their phenotype to mice lacking only ζ/η or Fcγ. The data reveal that surface complexes that are expressed in the absence of ζ family dimers are capable of transducing signals required for α/β–T cell development. Strikingly, T cells generated in both ζ/η−/− and ζ/η−/−–Fcγ−/− mice exhibit a memory phenotype and elaborate interferon γ. Finally, examination of different T cell populations reveals that ζ/η and Fcγ have distinct expression patterns that correlate with their thymus dependency. A possible function for the differential expression of ζ family proteins may be to impart distinctive signaling properties to TCR complexes expressed on specific T cell populations. PMID:9529325

Androgen signaling in myocytes contributes to the maintenance of muscle mass and fiber type regulation but not to muscle strength or fatigue.

PubMed

Ophoff, Jill; Van Proeyen, Karen; Callewaert, Filip; De Gendt, Karel; De Bock, Katrien; Vanden Bosch, An; Verhoeven, Guido; Hespel, Peter; Vanderschueren, Dirk

2009-08-01

Muscle frailty is considered a major cause of disability in the elderly and chronically ill. However, the exact role of androgen receptor (AR) signaling in muscle remains unclear. Therefore, a postmitotic myocyte-specific AR knockout (mARKO) mouse model was created and investigated together with a mouse model with ubiquitous AR deletion. Muscles from mARKO mice displayed a marked reduction in AR protein (60-88%). Interestingly, body weights and lean body mass were lower in mARKO vs. control mice (-8%). The weight of the highly androgen-sensitive musculus levator ani was significantly reduced (-46%), whereas the weights of other peripheral skeletal muscles were not or only slightly reduced. mARKO mice had lower intra-abdominal fat but did not demonstrate a cortical or trabecular bone phenotype, indicating that selective ablation of the AR in myocytes affected male body composition but not skeletal homeostasis. Furthermore, muscle contractile performance in mARKO mice did not differ from their controls. Myocyte-specific AR ablation resulted in a conversion of fast toward slow fibers, without affecting muscle strength or fatigue. Similar results were obtained in ubiquitous AR deletion, showing lower body weight, whereas some but not all muscle weights were reduced. The percent slow fibers was increased, but no changes in muscle strength or fatigue could be detected. Together, our findings show that myocyte AR signaling contributes to the maintenance of muscle mass and fiber type regulation but not to muscle strength or fatigue. The levator ani weight remains the most sensitive and specific marker of AR-mediated anabolic action on muscle.

In vivo1H NMR spectroscopy of the human brain at 9.4 T: Initial results

NASA Astrophysics Data System (ADS)

Deelchand, Dinesh Kumar; Moortele, Pierre-François Van de; Adriany, Gregor; Iltis, Isabelle; Andersen, Peter; Strupp, John P.; Thomas Vaughan, J.; Uğurbil, Kâmil; Henry, Pierre-Gilles

2010-09-01

In vivo proton NMR spectroscopy allows non-invasive detection and quantification of a wide range of biochemical compounds in the brain. Higher field strength is generally considered advantageous for spectroscopy due to increased signal-to-noise and increased spectral dispersion. So far 1H NMR spectra have been reported in the human brain up to 7 T. In this study we show that excellent quality short echo time STEAM and LASER 1H NMR spectra can be measured in the human brain at 9.4 T. The information content of the human brain spectra appears very similar to that measured in the past decade in rodent brains at the same field strength, in spite of broader linewidth in human brain. Compared to lower fields, the T1 relaxation times of metabolites were slightly longer while T2 relaxation values of metabolites were shorter (<100 ms) at 9.4 T. The linewidth of the total creatine (tCr) resonance at 3.03 ppm increased linearly with magnetic field (1.35 Hz/T from 1.5 T to 9.4 T), with a minimum achievable tCr linewidth of around 12.5 Hz at 9.4 T. At very high field, B0 microsusceptibility effects are the main contributor to the minimum achievable linewidth.

Constitutive Lck Activity Drives Sensitivity Differences between CD8+ Memory T Cell Subsets.

PubMed

Moogk, Duane; Zhong, Shi; Yu, Zhiya; Liadi, Ivan; Rittase, William; Fang, Victoria; Dougherty, Janna; Perez-Garcia, Arianne; Osman, Iman; Zhu, Cheng; Varadarajan, Navin; Restifo, Nicholas P; Frey, Alan B; Krogsgaard, Michelle

2016-07-15

CD8(+) T cells develop increased sensitivity following Ag experience, and differences in sensitivity exist between T cell memory subsets. How differential TCR signaling between memory subsets contributes to sensitivity differences is unclear. We show in mouse effector memory T cells (TEM) that >50% of lymphocyte-specific protein tyrosine kinase (Lck) exists in a constitutively active conformation, compared with <20% in central memory T cells (TCM). Immediately proximal to Lck signaling, we observed enhanced Zap-70 phosphorylation in TEM following TCR ligation compared with TCM Furthermore, we observed superior cytotoxic effector function in TEM compared with TCM, and we provide evidence that this results from a lower probability of TCM reaching threshold signaling owing to the decreased magnitude of TCR-proximal signaling. We provide evidence that the differences in Lck constitutive activity between CD8(+) TCM and TEM are due to differential regulation by SH2 domain-containing phosphatase-1 (Shp-1) and C-terminal Src kinase, and we use modeling of early TCR signaling to reveal the significance of these differences. We show that inhibition of Shp-1 results in increased constitutive Lck activity in TCM to levels similar to TEM, as well as increased cytotoxic effector function in TCM Collectively, this work demonstrates a role for constitutive Lck activity in controlling Ag sensitivity, and it suggests that differential activities of TCR-proximal signaling components may contribute to establishing the divergent effector properties of TCM and TEM. This work also identifies Shp-1 as a potential target to improve the cytotoxic effector functions of TCM for adoptive cell therapy applications. Copyright © 2016 by The American Association of Immunologists, Inc.

A Model of an Integrated Immune System Pathway in Homo sapiens and Its Interaction with Superantigen Producing Expression Regulatory Pathway in Staphylococcus aureus: Comparing Behavior of Pathogen Perturbed and Unperturbed Pathway

PubMed Central

Tomar, Namrata; De, Rajat K.

2013-01-01

Response of an immune system to a pathogen attack depends on the balance between the host immune defense and the virulence of the pathogen. Investigation of molecular interactions between the proteins of a host and a pathogen helps in identifying the pathogenic proteins. It is necessary to understand the dynamics of a normally behaved host system to evaluate the capacity of its immune system upon pathogen attack. In this study, we have compared the behavior of an unperturbed and pathogen perturbed host system. Moreover, we have developed a formalism under Flux Balance Analysis (FBA) for the optimization of conflicting objective functions. We have constructed an integrated pathway system, which includes Staphylococcal Superantigen (SAg) expression regulatory pathway and TCR signaling pathway of Homo sapiens. We have implemented the method on this pathway system and observed the behavior of host signaling molecules upon pathogen attack. The entire study has been divided into six different cases, based on the perturbed/unperturbed conditions. In other words, we have investigated unperturbed and pathogen perturbed human TCR signaling pathway, with different combinations of optimization of concentrations of regulatory and signaling molecules. One of these cases has aimed at finding out whether minimization of the toxin production in a pathogen leads to the change in the concentration levels of the proteins coded by TCR signaling pathway genes in the infected host. Based on the computed results, we have hypothesized that the balance between TCR signaling inhibitory and stimulatory molecules can keep TCR signaling system into resting/stimulating state, depending upon the perturbation. The proposed integrated host-pathogen interaction pathway model has accurately reflected the experimental evidences, which we have used for validation purpose. The significance of this kind of investigation lies in revealing the susceptible interaction points that can take back the

Conformational analysis of the ΜΒΡ83-99 (Phe91) and ΜΒΡ83-99 (Tyr91) peptide analogues and study of their interactions with the HLA-DR2 and human TCR receptors by using Molecular Dynamics

NASA Astrophysics Data System (ADS)

Potamitis, C.; Matsoukas, M.-T.; Tselios, T.; Mavromoustakos, T.; Golič Grdadolnik, S.

2011-09-01

The two new synthetic analogues of the MBP83-99 epitope substituted at Lys91 (primary TCR contact) with Phe [MBP83-99 (Phe91)] or Tyr [MBP83-99 (Tyr91)], have been structurally elucidated using 1D and 2D high resolution NMR studies. The conformational analysis of the two altered peptide ligands (APLs) has been performed and showed that they adopt a linear and extended conformation which is in agreement with the structural requirements of the peptides that interact with the HLA-DR2 and TCR receptors. In addition, Molecular Dynamics (MD) simulations of the two analogues in complex with HLA-DR2 (DRA, DRB1*1501) and TCR were performed. Similarities and differences of the binding motif of the two analogues were observed which provide a possible explanation of their biological activity. Their differences in the binding mode in comparison with the MBP83-99 epitope may also explain their antagonistic versus agonistic activity. The obtained results clearly indicate that substitutions in crucial amino acids (TCR contacts) in combination with the specific conformational characteristics of the MBP83-99 immunodominant epitope lead to an alteration of their biological activity. These results make the rational drug design intriguing since the biological activity is very sensitive to the substitution and conformation of the mutated MBP epitopes.

A Mechanical Switch Couples T Cell Receptor Triggering to the Cytoplasmic Juxtamembrane Regions of CD3ζζ.

PubMed

Lee, Mark S; Glassman, Caleb R; Deshpande, Neha R; Badgandi, Hemant B; Parrish, Heather L; Uttamapinant, Chayasith; Stawski, Philipp S; Ting, Alice Y; Kuhns, Michael S

2015-08-18

The eight-subunit T cell receptor (TCR)-CD3 complex is the primary determinant for T cell fate decisions. Yet how it relays ligand-specific information across the cell membrane for conversion to chemical signals remains unresolved. We hypothesized that TCR engagement triggers a change in the spatial relationship between the associated CD3ζζ subunits at the junction where they emerge from the membrane into the cytoplasm. Using three in situ proximity assays based on ID-PRIME, FRET, and EPOR activity, we determined that the cytosolic juxtamembrane regions of the CD3ζζ subunits are spread apart upon assembly into the TCR-CD3 complex. TCR engagement then triggered their apposition. This mechanical switch resides upstream of the CD3ζζ intracellular motifs that initiate chemical signaling, as well as the polybasic stretches that regulate signal potentiation. These findings provide a framework from which to examine triggering events for activating immune receptors and other complex molecular machines. Copyright © 2015 Elsevier Inc. All rights reserved.

A conserved αβ transmembrane interface forms the core of a compact T-cell receptor–CD3 structure within the membrane

PubMed Central

Krshnan, Logesvaran; Park, Soohyung; Im, Wonpil; Call, Melissa J.; Call, Matthew E.

2016-01-01

The T-cell antigen receptor (TCR) is an assembly of eight type I single-pass membrane proteins that occupies a central position in adaptive immunity. Many TCR-triggering models invoke an alteration in receptor complex structure as the initiating event, but both the precise subunit organization and the pathway by which ligand-induced alterations are transferred to the cytoplasmic signaling domains are unknown. Here, we show that the receptor complex transmembrane (TM) domains form an intimately associated eight-helix bundle organized by a specific interhelical TCR TM interface. The salient features of this core structure are absolutely conserved between αβ and γδ TCR sequences and throughout vertebrate evolution, and mutations at key interface residues caused defects in the formation of stable TCRαβ:CD3δε:CD3γε:ζζ complexes. These findings demonstrate that the eight TCR–CD3 subunits form a compact and precisely organized structure within the membrane and provide a structural basis for further investigation of conformationally regulated models of transbilayer TCR signaling. PMID:27791034

Enhanced Wnt signaling improves bone mass and strength, but not brittleness, in the Col1a1(+/mov13) mouse model of type I Osteogenesis Imperfecta.

PubMed

Jacobsen, Christina M; Schwartz, Marissa A; Roberts, Heather J; Lim, Kyung-Eun; Spevak, Lyudmila; Boskey, Adele L; Zurakowski, David; Robling, Alexander G; Warman, Matthew L

2016-09-01

Osteogenesis Imperfecta (OI) comprises a group of genetic skeletal fragility disorders. The mildest form of OI, Osteogenesis Imperfecta type I, is frequently caused by haploinsufficiency mutations in COL1A1, the gene encoding the α1(I) chain of type 1 collagen. Children with OI type I have a 95-fold higher fracture rate compared to unaffected children. Therapies for OI type I in the pediatric population are limited to anti-catabolic agents. In adults with osteoporosis, anabolic therapies that enhance Wnt signaling in bone improve bone mass, and ongoing clinical trials are determining if these therapies also reduce fracture risk. We performed a proof-of-principle experiment in mice to determine whether enhancing Wnt signaling in bone could benefit children with OI type I. We crossed a mouse model of OI type I (Col1a1(+/Mov13)) with a high bone mass (HBM) mouse (Lrp5(+/p.A214V)) that has increased bone strength from enhanced Wnt signaling. Offspring that inherited the OI and HBM alleles had higher bone mass and strength than mice that inherited the OI allele alone. However, OI+HBM and OI mice still had bones with lower ductility compared to wild-type mice. We conclude that enhancing Wnt signaling does not make OI bone normal, but does improve bone properties that could reduce fracture risk. Therefore, agents that enhance Wnt signaling are likely to benefit children and adults with OI type 1. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of varying Notch1 signal strength on embryogenesis and vasculogenesis in compound mutant heterozygotes

PubMed Central

2010-01-01

Background Identifying developmental processes regulated by Notch1 can be addressed in part by characterizing mice with graded levels of Notch1 signaling strength. Here we examine development in embryos expressing various combinations of Notch1 mutant alleles. Mice homozygous for the hypomorphic Notch112f allele, which removes the single O-fucose glycan in epidermal growth factor-like repeat 12 (EGF12) of the Notch1 ligand binding domain (lbd), exhibit reduced growth after weaning and defective T cell development. Mice homozygous for the inactive Notch1lbd allele express Notch1 missing an ~20 kDa internal segment including the canonical Notch1 ligand binding domain, and die at embryonic day ~E9.5. The embryonic and vascular phenotypes of compound heterozygous Notch112f/lbd embryos were compared with Notch1+/12f, Notch112f/12f, and Notch1lbd/lbd embryos. Embryonic stem (ES) cells derived from these embryos were also examined in Notch signaling assays. While Notch1 signaling was stronger in Notch112f/lbd compound heterozygotes compared to Notch1lbd/lbd embryos and ES cells, Notch1 signaling was even stronger in embryos carrying Notch112f and a null Notch1 allele. Results Mouse embryos expressing the hypomorphic Notch112f allele, in combination with the inactive Notch1lbd allele which lacks the Notch1 ligand binding domain, died at ~E11.5-12.5. Notch112f/lbd ES cells signaled less well than Notch112f/12f ES cells but more strongly than Notch1lbd/lbd ES cells. However, vascular defects in Notch112f/lbd yolk sac were severe and similar to Notch1lbd/lbd yolk sac. By contrast, vascular disorganization was milder in Notch112f/lbd compared to Notch1lbd/lbd embryos. The expression of Notch1 target genes was low in Notch112f/lbd yolk sac and embryo head, whereas Vegf and Vegfr2 transcripts were increased. The severity of the compound heterozygous Notch112f/lbd yolk sac phenotype suggested that the allelic products may functionally interact. By contrast, compound heterozygotes

Chimeric antigen receptor T cells form nonclassical and potent immune synapses driving rapid cytotoxicity.

PubMed

Davenport, A J; Cross, R S; Watson, K A; Liao, Y; Shi, W; Prince, H M; Beavis, P A; Trapani, J A; Kershaw, M H; Ritchie, D S; Darcy, P K; Neeson, P J; Jenkins, M R

2018-02-27

Chimeric antigen receptor T (CAR-T) cells are effective serial killers with a faster off-rate from dying tumor cells than CAR-T cells binding target cells through their T cell receptor (TCR). Here we explored the functional consequences of CAR-mediated signaling using a dual-specific CAR-T cell, where the same cell was triggered via TCR (tcrCTL) or CAR (carCTL). The carCTL immune synapse lacked distinct LFA-1 adhesion rings and was less reliant on LFA to form stable conjugates with target cells. carCTL receptors associated with the synapse were found to be disrupted and formed a convoluted multifocal pattern of Lck microclusters. Both proximal and distal receptor signaling pathways were induced more rapidly and subsequently decreased more rapidly in carCTL than in tcrCTL. The functional consequence of this rapid signaling in carCTL cells included faster lytic granule recruitment to the immune synapse, correlating with faster detachment of the CTL from the target cell. This study provides a mechanism for how CAR-T cells can debulk large tumor burden quickly and may contribute to further refinement of CAR design for enhancing the quality of signaling and programming of the T cell. Copyright © 2018 the Author(s). Published by PNAS.

Chimeric antigen receptor T cells form nonclassical and potent immune synapses driving rapid cytotoxicity

PubMed Central

Davenport, A. J.; Cross, R. S.; Watson, K. A.; Liao, Y.; Shi, W.; Prince, H. M.; Beavis, P. A.; Trapani, J. A.; Kershaw, M. H.; Ritchie, D. S.; Darcy, P. K.; Jenkins, M. R.

2018-01-01

Chimeric antigen receptor T (CAR-T) cells are effective serial killers with a faster off-rate from dying tumor cells than CAR-T cells binding target cells through their T cell receptor (TCR). Here we explored the functional consequences of CAR-mediated signaling using a dual-specific CAR-T cell, where the same cell was triggered via TCR (tcrCTL) or CAR (carCTL). The carCTL immune synapse lacked distinct LFA-1 adhesion rings and was less reliant on LFA to form stable conjugates with target cells. carCTL receptors associated with the synapse were found to be disrupted and formed a convoluted multifocal pattern of Lck microclusters. Both proximal and distal receptor signaling pathways were induced more rapidly and subsequently decreased more rapidly in carCTL than in tcrCTL. The functional consequence of this rapid signaling in carCTL cells included faster lytic granule recruitment to the immune synapse, correlating with faster detachment of the CTL from the target cell. This study provides a mechanism for how CAR-T cells can debulk large tumor burden quickly and may contribute to further refinement of CAR design for enhancing the quality of signaling and programming of the T cell. PMID:29440406

Method and Apparatus to Access Optimum Strength During Processing of Precipitation Strengthened Alloys

NASA Technical Reports Server (NTRS)

Cantrell, John H. (Inventor); Yost, William T. (Inventor)

2001-01-01

A method and apparatus are provided which enable the nondestructive testing of strength of a heat treated alloy. An alloy is insonified with an ultrasonic signal. The resulting convoluted signal is detected and the acoustic nonlinearity parameter is determined. The acoustic nonlinearity parameter shows a peak corresponding to a peak in material strength.

Development of a Novel CD4+ TCR Transgenic Line That Reveals a Dominant Role for CD8+ Dendritic Cells and CD40 Signaling in the Generation of Helper and CTL Responses to Blood-Stage Malaria.

PubMed

Fernandez-Ruiz, Daniel; Lau, Lei Shong; Ghazanfari, Nazanin; Jones, Claerwen M; Ng, Wei Yi; Davey, Gayle M; Berthold, Dorothee; Holz, Lauren; Kato, Yu; Enders, Matthias H; Bayarsaikhan, Ganchimeg; Hendriks, Sanne H; Lansink, Lianne I M; Engel, Jessica A; Soon, Megan S F; James, Kylie R; Cozijnsen, Anton; Mollard, Vanessa; Uboldi, Alessandro D; Tonkin, Christopher J; de Koning-Ward, Tania F; Gilson, Paul R; Kaisho, Tsuneyasu; Haque, Ashraful; Crabb, Brendan S; Carbone, Francis R; McFadden, Geoffrey I; Heath, William R

2017-12-15

We describe an MHC class II (I-A b )-restricted TCR transgenic mouse line that produces CD4 + T cells specific for Plasmodium species. This line, termed PbT-II, was derived from a CD4 + T cell hybridoma generated to blood-stage Plasmodium berghei ANKA (PbA). PbT-II cells responded to all Plasmodium species and stages tested so far, including rodent (PbA, P. berghei NK65, Plasmodium chabaudi AS, and Plasmodium yoelii 17XNL) and human ( Plasmodium falciparum ) blood-stage parasites as well as irradiated PbA sporozoites. PbT-II cells can provide help for generation of Ab to P. chabaudi infection and can control this otherwise lethal infection in CD40L-deficient mice. PbT-II cells can also provide help for development of CD8 + T cell-mediated experimental cerebral malaria (ECM) during PbA infection. Using PbT-II CD4 + T cells and the previously described PbT-I CD8 + T cells, we determined the dendritic cell (DC) subsets responsible for immunity to PbA blood-stage infection. CD8 + DC (a subset of XCR1 + DC) were the major APC responsible for activation of both T cell subsets, although other DC also contributed to CD4 + T cell responses. Depletion of CD8 + DC at the beginning of infection prevented ECM development and impaired both Th1 and follicular Th cell responses; in contrast, late depletion did not affect ECM. This study describes a novel and versatile tool for examining CD4 + T cell immunity during malaria and provides evidence that CD4 + T cell help, acting via CD40L signaling, can promote immunity or pathology to blood-stage malaria largely through Ag presentation by CD8 + DC. Copyright © 2017 by The American Association of Immunologists, Inc.

Crystal structure of a complete ternary complex of T-cell receptor, peptide-MHC, and CD4

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yin, Yiyuan; Wang, Xin Xiang; Mariuzza, Roy A

2012-07-11

Adaptive immunity depends on specific recognition by a T-cell receptor (TCR) of an antigenic peptide bound to a major histocompatibility complex (pMHC) molecule on an antigen-presenting cell (APC). In addition, T-cell activation generally requires binding of this same pMHC to a CD4 or CD8 coreceptor. Here, we report the structure of a complete TCR-pMHC-CD4 ternary complex involving a human autoimmune TCR, a myelin-derived self-peptide bound to HLA-DR4, and CD4. The complex resembles a pointed arch in which TCR and CD4 are each tilted ~65° relative to the T-cell membrane. By precluding direct contacts between TCR and CD4, the structure explainsmore » how TCR and CD4 on the T cell can simultaneously, yet independently, engage the same pMHC on the APC. The structure, in conjunction with previous mutagenesis data, places TCR-associated CD3εγ and CD3εδ subunits, which transmit activation signals to the T cell, inside the TCR-pMHC-CD4 arch, facing CD4. By establishing anchor points for TCR and CD4 on the T-cell membrane, the complex provides a basis for understanding how the CD4 coreceptor focuses TCR on MHC to guide TCR docking on pMHC during thymic T-cell selection.« less

MHC drives TCR repertoire shaping, but not maturation, in recent thymic emigrants.

PubMed

Houston, Evan G; Fink, Pamela J

2009-12-01

After developing in the thymus, recent thymic emigrants (RTEs) enter the lymphoid periphery and undergo a maturation process as they transition into the mature naive (MN) T cell compartment. This maturation presumably shapes RTEs into a pool of T cells best fit to function robustly in the periphery without causing autoimmunity; however, the mechanism and consequences of this maturation process remain unknown. Using a transgenic mouse system that specifically labels RTEs, we tested the influence of MHC molecules, key drivers of intrathymic T cell selection and naive peripheral T cell homeostasis, in shaping the RTE pool in the lymphoid periphery. We found that the TCRs expressed by RTEs are skewed to longer CDR3 regions compared with those of MN T cells, suggesting that MHC does streamline the TCR repertoire of T cells as they transition from the RTE to the MN T cell stage. This conclusion is borne out in studies in which the representation of individual TCRs was followed as a function of time since thymic egress. Surprisingly, we found that MHC is dispensable for the phenotypic and functional maturation of RTEs.

TET proteins regulate the lineage specification and TCR-mediated expansion of iNKT cells.

PubMed

Tsagaratou, Ageliki; González-Avalos, Edahí; Rautio, Sini; Scott-Browne, James P; Togher, Susan; Pastor, William A; Rothenberg, Ellen V; Chavez, Lukas; Lähdesmäki, Harri; Rao, Anjana

2017-01-01

TET proteins oxidize 5-methylcytosine in DNA to 5-hydroxymethylcytosine and other oxidation products. We found that simultaneous deletion of Tet2 and Tet3 in mouse CD4 + CD8 + double-positive thymocytes resulted in dysregulated development and proliferation of invariant natural killer T cells (iNKT cells). Tet2-Tet3 double-knockout (DKO) iNKT cells displayed pronounced skewing toward the NKT17 lineage, with increased DNA methylation and impaired expression of genes encoding the key lineage-specifying factors T-bet and ThPOK. Transfer of purified Tet2-Tet3 DKO iNKT cells into immunocompetent recipient mice resulted in an uncontrolled expansion that was dependent on the nonclassical major histocompatibility complex (MHC) protein CD1d, which presents lipid antigens to iNKT cells. Our data indicate that TET proteins regulate iNKT cell fate by ensuring their proper development and maturation and by suppressing aberrant proliferation mediated by the T cell antigen receptor (TCR).

Hyper-Expression of PD-1 Is Associated with the Levels of Exhausted and Dysfunctional Phenotypes of Circulating CD161++TCR iVα7.2+ Mucosal-Associated Invariant T Cells in Chronic Hepatitis B Virus Infection.

PubMed

Yong, Yean K; Saeidi, Alireza; Tan, Hong Y; Rosmawati, Mohamed; Enström, Philip F; Batran, Rami Al; Vasuki, V; Chattopadhyay, Indranil; Murugesan, Amudhan; Vignesh, Ramachandran; Kamarulzaman, Adeeba; Rajarajeswaran, Jayakumar; Ansari, Abdul W; Vadivelu, Jamuna; Ussher, James E; Velu, Vijayakumar; Larsson, Marie; Shankar, Esaki M

2018-01-01

Mucosal-associated invariant T (MAIT) cells, defined as CD161 ++ TCR iVα7.2 + T cells, play an important role in the innate defense against bacterial infections, and their functionality is impaired in chronic viral infections. Here, we investigated the frequency and functional role of MAIT cells in chronic hepatitis B virus (HBV) infection. The peripheral CD3 + CD161 ++ TCR iVα7.2 + MAIT cells in chronic HBV-infected patients and healthy controls were phenotypically characterized based on CD57, PD-1, TIM-3, and CTLA-4, as well as HLA-DR and CD38 expression. The frequency of MAIT cells was significantly decreased among chronic HBV-infected individuals as compared to controls. Expression of CD57, PD-1, CTLA-4, as well as HLA-DR and CD38 on MAIT cells was significantly elevated in chronic HBV-infected individuals relative to controls. The percentage of T cell receptor (TCR) iVα7.2 + CD161 + MAIT cells did not correlate with HBV viral load but inversely with HLA-DR on CD4 + T cells and MAIT cells and with CD57 on CD8 + T cells suggesting that decrease of MAIT cells may not be attributed to direct infection by HBV but driven by HBV-induced chronic immune activation. The percentage and expression levels of PD-1 as well as CTLA-4 on MAIT cells inversely correlated with plasma HBV-DNA levels, which may suggest either a role for MAIT cells in the control of HBV infection or the effect of HBV replication in the liver on MAIT cell phenotype. We report that decrease of TCR iVα7.2 + MAIT cells in the peripheral blood and their functions were seemingly impaired in chronic HBV-infected patients likely because of the increased expression of PD-1.

Influence of aging on isometric muscle strength, fat-free mass and electromyographic signal power of the upper and lower limbs in women

PubMed Central

Amaral, Josária F.; Alvim, Felipe C.; Castro, Eliane A.; Doimo, Leonice A.; Silva, Marcus V.; Novo, José M.

2014-01-01

Background Aging is a multifactorial process that leads to changes in the quantity and quality of skeletal muscle and contributes to decreased levels of muscle strength. Objective This study sought to investigate whether the isometric muscle strength, fat-free mass (FFM) and power of the electromyographic (EMG) signal of the upper and lower limbs of women are similarly affected by aging. Method The sample consisted of 63 women, who were subdivided into three groups (young (YO) n=33, 24.7±3.5 years; middle age (MA) n=15, 58.6±4.2 years; and older adults (OA). n=15, 72.0±4.2 years). Isometric strength was recorded simultaneously with the capture of the electrical activity of the flexor muscles of the fingers and the vastus lateralis during handgrip and knee extension tests, respectively. FFM was assessed using dual-energy X-ray absorptiometry. Results The handgrip strength measurements were similar among groups (p=0.523), whereas the FFM of the upper limbs was lower in group OA compared to group YO (p=0.108). The RMSn values of the hand flexors were similar among groups (p=0.754). However, the strength of the knee extensors, the FFM of the lower limbs and the RMSn values of the vastus lateralis were lower in groups MA (p=0.014, p=0.006 and p=0.013, respectively) and OA (p=0.000, p=0.000 and p<0.000, respectively) compared to group YO. Conclusions The results of this study demonstrate that changes in isometric muscle strength in MLG and electromyographic activity of the lower limbs are more pronounced with the aging process of the upper limb. PMID:24676705

Signal-to-noise ratio, T2 , and T2* for hyperpolarized helium-3 MRI of the human lung at three magnetic field strengths.

PubMed

Komlosi, Peter; Altes, Talissa A; Qing, Kun; Mooney, Karen E; Miller, G Wilson; Mata, Jaime F; de Lange, Eduard E; Tobias, William A; Cates, Gordon D; Mugler, John P

2017-10-01

To evaluate T 2 , T2*, and signal-to-noise ratio (SNR) for hyperpolarized helium-3 ( 3 He) MRI of the human lung at three magnetic field strengths ranging from 0.43T to 1.5T. Sixteen healthy volunteers were imaged using a commercial whole body scanner at 0.43T, 0.79T, and 1.5T. Whole-lung T 2 values were calculated from a Carr-Purcell-Meiboom-Gill spin-echo-train acquisition. T2* maps and SNR were determined from dual-echo and single-echo gradient-echo images, respectively. Mean whole-lung SNR values were normalized by ventilated lung volume and administered 3 He dose. As expected, T 2 and T2* values demonstrated a significant inverse relationship to field strength. Hyperpolarized 3 He images acquired at all three field strengths had comparable SNR values and thus appeared visually very similar. Nonetheless, the relatively small SNR differences among field strengths were statistically significant. Hyperpolarized 3 He images of the human lung with similar image quality were obtained at three field strengths ranging from 0.43T and 1.5T. The decrease in susceptibility effects at lower fields that are reflected in longer T 2 and T2* values may be advantageous for optimizing pulse sequences inherently sensitive to such effects. The three-fold increase in T2* at lower field strength would allow lower receiver bandwidths, providing a concomitant decrease in noise and relative increase in SNR. Magn Reson Med 78:1458-1463, 2017. © 2016 International Society for Magnetic Resonance in Medicine. © 2016 International Society for Magnetic Resonance in Medicine.

BASH, a novel signaling molecule preferentially expressed in B cells of the bursa of Fabricius.

PubMed

Goitsuka, R; Fujimura, Y; Mamada, H; Umeda, A; Morimura, T; Uetsuka, K; Doi, K; Tsuji, S; Kitamura, D

1998-12-01

The bursa of Fabricius is a gut-associated lymphoid organ that is essential for the generation of a diversified B cell repertoire in the chicken. We describe here a novel gene preferentially expressed in bursal B cells. The gene encodes an 85-kDa protein, designated BASH (B cell adaptor containing SH2 domain), that contains N-terminal acidic domains with SH2 domain-binding phosphotyrosine-based motifs, a proline-rich domain, and a C-terminal SH2 domain. BASH shows a substantial sequence similarity to SLP-76, an adaptor protein functioning in TCR-signal transduction. BASH becomes tyrosine-phosphorylated with the B cell Ag receptor (BCR) cross-link or by coexpression with Syk and Lyn and associates with signaling molecules including Syk and a putative chicken Shc homologue. Overexpression of BASH results in suppression of the NF-AT activation induced by BCR-cross-linking. These findings suggest that BASH is involved in BCR-mediated signal transduction and could play a critical role in B cell development in the bursa.

Critical biological parameters modulate affinity as a determinant of function in T-cell receptor gene-modified T-cells.

PubMed

Spear, Timothy T; Wang, Yuan; Foley, Kendra C; Murray, David C; Scurti, Gina M; Simms, Patricia E; Garrett-Mayer, Elizabeth; Hellman, Lance M; Baker, Brian M; Nishimura, Michael I

2017-11-01

T-cell receptor (TCR)-pMHC affinity has been generally accepted to be the most important factor dictating antigen recognition in gene-modified T-cells. As such, there is great interest in optimizing TCR-based immunotherapies by enhancing TCR affinity to augment the therapeutic benefit of TCR gene-modified T-cells in cancer patients. However, recent clinical trials using affinity-enhanced TCRs in adoptive cell transfer (ACT) have observed unintended and serious adverse events, including death, attributed to unpredicted off-tumor or off-target cross-reactivity. It is critical to re-evaluate the importance of other biophysical, structural, or cellular factors that drive the reactivity of TCR gene-modified T-cells. Using a model for altered antigen recognition, we determined how TCR-pMHC affinity influenced the reactivity of hepatitis C virus (HCV) TCR gene-modified T-cells against a panel of naturally occurring HCV peptides and HCV-expressing tumor targets. The impact of other factors, such as TCR-pMHC stabilization and signaling contributions by the CD8 co-receptor, as well as antigen and TCR density were also evaluated. We found that changes in TCR-pMHC affinity did not always predict or dictate IFNγ release or degranulation by TCR gene-modified T-cells, suggesting that less emphasis might need to be placed on TCR-pMHC affinity as a means of predicting or augmenting the therapeutic potential of TCR gene-modified T-cells used in ACT. A more complete understanding of antigen recognition by gene-modified T-cells and a more rational approach to improve the design and implementation of novel TCR-based immunotherapies is necessary to enhance efficacy and maximize safety in patients.

Different immunological mechanisms govern protection from experimental stroke in young and older mice with recombinant TCR ligand therapy

PubMed Central

Dotson, Abby L.; Zhu, Wenbin; Libal, Nicole; Alkayed, Nabil J.; Offner, Halina

2014-01-01

Stroke is a leading cause of death and disability in the United States. The lack of clinical success in stroke therapies can be attributed, in part, to inadequate basic research on aging rodents. The current study demonstrates that recombinant TCR ligand therapy uses different immunological mechanisms to protect young and older mice from experimental stroke. In young mice, RTL1000 therapy inhibited splenocyte efflux while reducing frequency of T cells and macrophages in the spleen. Older mice treated with RTL1000 exhibited a significant reduction in inflammatory cells in the brain and inhibition of splenic atrophy. Our data suggest age specific differences in immune response to stroke that allow unique targeting of stroke immunotherapies. PMID:25309326

Conditional deletion reveals a cell-autonomous requirement of SLP-76 for thymocyte selection.

PubMed

Maltzman, Jonathan S; Kovoor, Lisa; Clements, James L; Koretzky, Gary A

2005-10-03

The SH2 domain containing leukocyte phosphoprotein of 76 kD (SLP-76) is critical for pre-TCR-mediated maturation to the CD4+CD8+ double positive (DP) stage in the thymus. The absolute block in SLP-76null mice at the CD4-CD8-CD44-CD25+ (double-negative 3, DN3) stage has hindered our understanding of the role of this adaptor in alphabeta TCR-mediated signal transduction in primary thymocytes and peripheral T lymphocytes. To evaluate the requirements for SLP-76 in these events, we used a cre-loxP approach to generate mice that conditionally delete SLP-76 after the DN3 checkpoint. These mice develop DP thymocytes that express the alphabeta TCR on the surface, but lack SLP-76 at the genomic DNA and protein levels. The DP compartment has reduced cellularity in young mice and fails to undergo positive selection to CD4+ or CD8+ single positive (SP) cells in vivo or activation-induced cell death in vitro. A small number of CD4+SP thymocytes are generated, but these cells fail to flux calcium in response to an alphabeta TCR-generated signal. Peripheral T cells are reduced in number, lack SLP-76 protein, and have an abnormal surface phenotype. These studies show for the first time that SLP-76 is required for signal transduction through the mature alphabeta TCR in primary cells of the T lineage.

CCL21 mediates CD4+ T-cell costimulation via a DOCK2/Rac-dependent pathway.

PubMed

Gollmer, Kathrin; Asperti-Boursin, François; Tanaka, Yoshihiko; Okkenhaug, Klaus; Vanhaesebroeck, Bart; Peterson, Jeffrey R; Fukui, Yoshinori; Donnadieu, Emmanuel; Stein, Jens V

2009-07-16

CD4(+) T cells use the chemokine receptor CCR7 to home to and migrate within lymphoid tissue, where T-cell activation takes place. Using primary T-cell receptor (TCR)-transgenic (tg) CD4(+) T cells, we explored the effect of CCR7 ligands, in particular CCL21, on T-cell activation. We found that the presence of CCL21 during early time points strongly increased in vitro T-cell proliferation after TCR stimulation, correlating with increased expression of early activation markers. CCL21 costimulation resulted in increased Ras- and Rac-GTP formation and enhanced phosphorylation of Akt, MEK, and ERK but not p38 or JNK. Kinase-dead PI3Kdelta(D910A/D910A) or PI3Kgamma-deficient TCR-tg CD4(+) T cells showed similar responsiveness to CCL21 costimulation as control CD4(+) T cells. Conversely, deficiency in the Rac guanine exchange factor DOCK2 significantly impaired CCL21-mediated costimulation in TCR-tg CD4(+) T cells, concomitant with impaired Rac- but not Ras-GTP formation. Using lymph node slices for live monitoring of T-cell behavior and activation, we found that G protein-coupled receptor signaling was required for early CD69 expression but not for Ca(2+) signaling. Our data suggest that the presence of CCL21 during early TCR signaling lowers the activation threshold through Ras- and Rac-dependent pathways leading to increased ERK phosphorylation.

Sympathetic neural signaling via the β2-adrenergic receptor suppresses T-cell receptor-mediated human and mouse CD8(+) T-cell effector function.

PubMed

Estrada, Leonardo D; Ağaç, Didem; Farrar, J David

2016-08-01

Postganglionic sympathetic neurons innervate secondary lymphoid organs and secrete norepinephrine (NE) as the primary neurotransmitter. NE binds and signals through five distinct members of the adrenergic receptor family. In this study, we show elevated expression of the β2-adrenergic receptor (ADRB2) on primary human CD8(+) effector memory T cells. Treatment of both human and murine CD8(+) T cells with NE decreased IFN-γ and TNF-α secretion and suppressed their cytolytic capacity in response to T-cell receptor (TCR) activation. The effects of NE were specifically reversed by β2-specific antagonists. Adrb2(-/-) CD8(+) T cells were completely resistant to the effects of NE. Further, the ADRB2-specific pharmacological ligand, albuterol, significantly suppressed effector functions in both human and mouse CD8(+) T cells. While both TCR activation and stimulation with IL-12 + IL-18 were able to induce inflammatory cytokine secretion, NE failed to suppress IFN-γ secretion in response to IL-12 + IL18. Finally, the long-acting ADRB2-specific agonist, salmeterol, markedly reduced the cytokine secretion capacity of CD8(+) T cells in response to infection with vesicular stomatitis virus. This study reveals a novel intrinsic role for ADRB2 signaling in CD8(+) T-cell function and underscores the novel role this pathway plays in adaptive T-cell responses to infection. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quantitative Biology of Exercise-Induced Signal Transduction Pathways.

PubMed

Liu, Timon Cheng-Yi; Liu, Gang; Hu, Shao-Juan; Zhu, Ling; Yang, Xiang-Bo; Zhang, Quan-Guang

2017-01-01

Exercise is essential in regulating energy metabolism. Exercise activates cellular, molecular, and biochemical pathways with regulatory roles in training response adaptation. Among them, endurance/strength training of an individual has been shown to activate its respective signal transduction pathways in skeletal muscle. This was further studied from the viewpoint of quantitative difference (QD). For the mean values, [Formula: see text], of two sets of data, their QD is defined as [Formula: see text] ([Formula: see text]). The function-specific homeostasis (FSH) of a function of a biosystem is a negative-feedback response of the biosystem to maintain the function-specific conditions inside the biosystem so that the function is perfectly performed. A function in/far from its FSH is called a normal/dysfunctional function. A cellular normal function can resist the activation of other signal transduction pathways so that there are normal function-specific signal transduction pathways which full activation maintains the normal function. An acute endurance/strength training may be dysfunctional, but its regular training may be normal. The normal endurance/strength training of an individual may resist the activation of other signal transduction pathways in skeletal muscle so that there may be normal endurance/strength training-specific signal transduction pathways (NEPs/NSPs) in skeletal muscle. The endurance/strength training may activate NSPs/NEPs, but the QD from the control is smaller than 0.80. The simultaneous activation of both NSPs and NEPs may enhance their respective activation, and the QD from the control is larger than 0.80. The low level laser irradiation pretreatment of rats may promote the activation of NSPs in endurance training skeletal muscle. There may be NEPs/NSPs in skeletal muscle trained by normal endurance/strength training.

47 CFR 90.359 - Field strength limits for EA-licensed LMS systems.

Code of Federal Regulations, 2011 CFR

2011-10-01

... 47 Telecommunication 5 2011-10-01 2011-10-01 false Field strength limits for EA-licensed LMS... § 90.359 Field strength limits for EA-licensed LMS systems. EA-licensed multilateration systems shall limit the field strength of signals transmitted from their base stations to 47 dBuV/m at their EA...

Alteration of the gene expression profile of T-cell receptor αβ-modified T-cells with diffuse large B-cell lymphoma specificity.

PubMed

Zha, Xianfeng; Yin, Qingsong; Tan, Huo; Wang, Chunyan; Chen, Shaohua; Yang, Lijian; Li, Bo; Wu, Xiuli; Li, Yangqiu

2013-05-01

Antigen-specific, T-cell receptor (TCR)-modified cytotoxic T lymphocytes (CTLs) that target tumors are an attractive strategy for specific adoptive immunotherapy. Little is known about whether there are any alterations in the gene expression profile after TCR gene transduction in T cells. We constructed TCR gene-redirected CTLs with specificity for diffuse large B-cell lymphoma (DLBCL)-associated antigens to elucidate the gene expression profiles of TCR gene-redirected T-cells, and we further analyzed the gene expression profile pattern of these redirected T-cells by Affymetrix microarrays. The resulting data were analyzed using Bioconductor software, a two-fold cut-off expression change was applied together with anti-correlation of the profile ratios to render the microarray analysis set. The fold change of all genes was calculated by comparing the three TCR gene-modified T-cells and a negative control counterpart. The gene pathways were analyzed using Bioconductor and Kyoto Encyclopedia of Genes and Genomes. Identical genes whose fold change was greater than or equal to 2.0 in all three TCR gene-redirected T-cell groups in comparison with the negative control were identified as the differentially expressed genes. The differentially expressed genes were comprised of 33 up-regulated genes and 1 down-regulated gene including JUNB, FOS, TNF, INF-γ, DUSP2, IL-1B, CXCL1, CXCL2, CXCL9, CCL2, CCL4, and CCL8. These genes are mainly involved in the TCR signaling, mitogen-activated protein kinase signaling, and cytokine-cytokine receptor interaction pathways. In conclusion, we characterized the gene expression profile of DLBCL-specific TCR gene-redirected T-cells. The changes corresponded to an up-regulation in the differentiation and proliferation of the T-cells. These data may help to explain some of the characteristics of the redirected T-cells.

Conditional deletion of SLP-76 in mature T cells abrogates peripheral immune responses.

PubMed

Wu, Gregory F; Corbo, Evann; Schmidt, Michelle; Smith-Garvin, Jennifer E; Riese, Matthew J; Jordan, Martha S; Laufer, Terri M; Brown, Eric J; Maltzman, Jonathan S

2011-07-01

The adaptor protein Src homology 2 domain-containing leukocyte-specific protein of 76 kDa (SLP-76) is central to the organization of intracellular signaling downstream of the T-cell receptor (TCR). Evaluation of its role in mature, primary T cells has been hampered by developmental defects that occur in the absence of WT SLP-76 protein in thymocytes. Here, we show that following tamoxifen-regulated conditional deletion of SLP-76, mature, antigen-inexperienced T cells maintain normal TCR surface expression but fail to transduce TCR-generated signals. Conditionally deficient T cells fail to proliferate in response to antigenic stimulation or a lymphopenic environment. Mice with induced deletion of SLP-76 are resistant to induction of the CD4+ T-cell-mediated autoimmune disease experimental autoimmune encephalomyelitis. Altogether, our findings demonstrate the critical role of SLP-76-mediated signaling in initiating T-cell-directed immune responses both in vitro and in vivo and highlight the ability to analyze signaling processes in mature T cells in the absence of developmental defects. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Conditional deletion of SLP-76 in mature T cells abrogates peripheral immune responses1

PubMed Central

Wu, Gregory F.; Corbo, Evann; Schmidt, Michelle; Smith-Garvin, Jennifer E.; Riese, Matthew J.; Jordan, Martha S.; Laufer, Terri M.; Brown, Eric J.; Maltzman, Jonathan S.

2011-01-01

SUMMARY The adaptor protein Src homology 2 domain-containing leukocyte-specific protein of 76 kDa (SLP-76) is central to the organization of intracellular signaling downstream of the T cell receptor (TCR). Evaluation of its role in mature, primary T cells has been hampered by developmental defects that occur in the absence of wild-type SLP-76 protein in thymocytes. Following tamoxifen-regulated conditional deletion of SLP-76, mature, antigen-inexperienced T cells maintain normal TCR surface expression but fail to transduce TCR generated signals. Conditionally deficient T cells fail to proliferate in response to antigenic stimulation or a lymphopenic environment. Mice with induced deletion of SLP-76 are resistant to induction of the CD4+ T cell mediated autoimmune disease experimental autoimmune encephalomyelitis. Our findings demonstrate the critical role of SLP-76-mediated signaling in initiating T cell-directed immune responses both in vitro and in vivo and highlight the ability to analyze signaling processes in mature T cells in the absence of developmental defects. PMID:21469089

Simulate different environments TDLAS On the analysis of the test signal strength

NASA Astrophysics Data System (ADS)

Li, Xin; Zhou, Tao; Jia, Xiaodong

2014-12-01

TDLAS system is the use of the wavelength tuning characteristics of the laser diode, for detecting the absorption spectrum of the gas absorption line. Detecting the gas space, temperature, pressure and flow rate and concentration. The use of laboratory techniques TDLAS gas detection, experimental simulation engine combustion water vapor and smoke. using an optical lens system receives the signal acquisition and signal interference test analysis. Analog water vapor and smoke in two different environments in the sample pool interference. In both experiments environmental interference gas absorption in the optical signal acquisition, signal amplitude variation analysis, and records related to the signal data. In order to study site conditions in the engine combustion process for signal acquisition provides an ideal experimental data .

The Cytoplasmic Tail of the T Cell Receptor CD3 ε Subunit Contains a Phospholipid-Binding Motif that Regulates T Cell Functions1

PubMed Central

DeFord-Watts, Laura M.; Tassin, Tara C.; Becker, Amy M.; Medeiros, Jennifer J.; Albanesi, Joseph P.; Love, Paul E.; Wülfing, Christoph; van Oers, Nicolai S. C.

2010-01-01

The CD3 ε subunit of the TCR complex contains two defined signaling domains, a proline-rich sequence and an ITAM. We identified a third signaling sequence in CD3 ε, termed the basic-rich stretch (BRS). Herein, we show that the positively charged residues of the BRS enable this region of CD3 ε to complex a subset of acidic phospholipids, including PI(3)P, PI(4)P, PI(5)P, PI(3,4,5)P3, and PI(4,5)P2. Transgenic mice containing mutations of the BRS exhibited varying developmental defects, ranging from reduced thymic cellularity to a complete block in T cell development. Peripheral T cells from BRS-modified mice also exhibited several defects, including decreased TCR surface expression, reduced TCR-mediated signaling responses to agonist peptide-loaded APCs, and delayed CD3 ε localization to the immunological synapse. Overall, these findings demonstrate a functional role for the CD3 ε lipid-binding domain in T cell biology. PMID:19542373

D120 and K152 within the PH Domain of T Cell Adapter SKAP55 Regulate Plasma Membrane Targeting of SKAP55 and LFA-1 Affinity Modulation in Human T Lymphocytes

PubMed Central

Witte, Amelie; Meineke, Bernhard; Sticht, Jana; Philipsen, Lars; Kuropka, Benno; Müller, Andreas J.; Freund, Christian

2017-01-01

ABSTRACT The β2-integrin lymphocyte function-associated antigen 1 (LFA-1) is needed for the T cell receptor (TCR)-induced activation of LFA-1 to promote T cell adhesion and interaction with antigen-presenting cells (APCs). LFA-1-mediated cell-cell interactions are critical for proper T cell differentiation and proliferation. The Src kinase-associated phosphoprotein of 55 kDa (SKAP55) is a key regulator of TCR-mediated LFA-1 signaling (inside-out/outside-in signaling). To gain an understanding of how SKAP55 controls TCR-mediated LFA-1 activation, we assessed the functional role of its pleckstrin homology (PH) domain. We identified two critical amino acid residues within the PH domain of SKAP55, aspartic acid 120 (D120) and lysine 152 (K152). D120 facilitates the retention of SKAP55 in the cytoplasm of nonstimulated T cells, while K152 promotes SKAP55 membrane recruitment via actin binding upon TCR triggering. Importantly, the K152-dependent interaction of the PH domain with actin promotes the binding of talin to LFA-1, thus facilitating LFA-1 activation. These data suggest that K152 and D120 within the PH domain of SKAP55 regulate plasma membrane targeting and TCR-mediated activation of LFA-1. PMID:28052935

Enhanced astroglial Ca2+ signaling increases excitatory synaptic strength in the epileptic brain.

PubMed

Álvarez-Ferradas, Carla; Morales, Juan Carlos; Wellmann, Mario; Nualart, Francisco; Roncagliolo, Manuel; Fuenzalida, Marco; Bonansco, Christian

2015-09-01

The fine-tuning of synaptic transmission by astrocyte signaling is crucial to CNS physiology. However, how exactly astroglial excitability and gliotransmission are affected in several neuropathologies, including epilepsy, remains unclear. Here, using a chronic model of temporal lobe epilepsy (TLE) in rats, we found that astrocytes from astrogliotic hippocampal slices displayed an augmented incidence of TTX-insensitive spontaneous slow Ca(2+) transients (STs), suggesting a hyperexcitable pattern of astroglial activity. As a consequence, elevated glutamate-mediated gliotransmission, observed as increased slow inward current (SICs) frequency, up-regulates the probability of neurotransmitter release in CA3-CA1 synapses. Selective blockade of spontaneous astroglial Ca(2+) elevations as well as the inhibition of purinergic P2Y1 or mGluR5 receptors relieves the abnormal enhancement of synaptic strength. Moreover, mGluR5 blockade eliminates any synaptic effects induced by P2Y1R inhibition alone, suggesting that the Pr modulation via mGluR occurs downstream of P2Y1R-mediated Ca(2+)-dependent glutamate release from astrocyte. Our findings show that elevated Ca(2+)-dependent glutamate gliotransmission from hyperexcitable astrocytes up-regulates excitatory neurotransmission in epileptic hippocampus, suggesting that gliotransmission should be considered as a novel functional key in a broad spectrum of neuropathological conditions. © 2015 Wiley Periodicals, Inc.

Chimeric Antigen Receptor- and TCR-Modified T Cells Enter Main Street and Wall Street.

PubMed

Barrett, David M; Grupp, Stephan A; June, Carl H

2015-08-01

The field of adoptive cell transfer (ACT) is currently comprised of chimeric Ag receptor (CAR)- and TCR-engineered T cells and has emerged from principles of basic immunology to paradigm-shifting clinical immunotherapy. ACT of T cells engineered to express artificial receptors that target cells of choice is an exciting new approach for cancer, and it holds equal promise for chronic infection and autoimmunity. Using principles of synthetic biology, advances in immunology, and genetic engineering have made it possible to generate human T cells that display desired specificities and enhanced functionalities. Clinical trials in patients with advanced B cell leukemias and lymphomas treated with CD19-specific CAR T cells have induced durable remissions in adults and children. The prospects for the widespread availability of engineered T cells have changed dramatically given the recent entry of the pharmaceutical industry to this arena. In this overview, we discuss some of the challenges and opportunities that face the field of ACT. Copyright © 2015 by The American Association of Immunologists, Inc.

Histogram Matching Extends Acceptable Signal Strength Range on Optical Coherence Tomography Images

PubMed Central

Chen, Chieh-Li; Ishikawa, Hiroshi; Wollstein, Gadi; Bilonick, Richard A.; Sigal, Ian A.; Kagemann, Larry; Schuman, Joel S.

2015-01-01

Purpose. We minimized the influence of image quality variability, as measured by signal strength (SS), on optical coherence tomography (OCT) thickness measurements using the histogram matching (HM) method. Methods. We scanned 12 eyes from 12 healthy subjects with the Cirrus HD-OCT device to obtain a series of OCT images with a wide range of SS (maximal range, 1–10) at the same visit. For each eye, the histogram of an image with the highest SS (best image quality) was set as the reference. We applied HM to the images with lower SS by shaping the input histogram into the reference histogram. Retinal nerve fiber layer (RNFL) thickness was automatically measured before and after HM processing (defined as original and HM measurements), and compared to the device output (device measurements). Nonlinear mixed effects models were used to analyze the relationship between RNFL thickness and SS. In addition, the lowest tolerable SSs, which gave the RNFL thickness within the variability margin of manufacturer recommended SS range (6–10), were determined for device, original, and HM measurements. Results. The HM measurements showed less variability across a wide range of image quality than the original and device measurements (slope = 1.17 vs. 4.89 and 1.72 μm/SS, respectively). The lowest tolerable SS was successfully reduced to 4.5 after HM processing. Conclusions. The HM method successfully extended the acceptable SS range on OCT images. This would qualify more OCT images with low SS for clinical assessment, broadening the OCT application to a wider range of subjects. PMID:26066749

Estimation of muscle strength during motion recognition using multichannel surface EMG signals.

PubMed

Nagata, Kentaro; Nakano, Takemi; Magatani, Kazushige; Yamada, Masafumi

2008-01-01

The use of kinesiological electromyography is established as an evaluation tool for various kinds of applied research, and surface electromyogram (SEMG) has been widely used as a control source for human interfaces such as in a myoelectric prosthetic hand (we call them 'SEMG interfaces'). It is desirable to be able to control the SEMG interfaces with the same feeling as body movement. The existing SEMG interface mainly focuses on how to achieve accurate recognition of the intended movement. However, detecting muscular strength and reduced number of electrodes are also an important factor in controlling them. Therefore, our objective in this study is the development of and the estimation method for muscular strength that maintains the accuracy of hand motion recognition to reflect the result of measured power in a controlled object. Although the muscular strength can be evaluated by various methods, in this study a grasp force index was applied to evaluate the muscular strength. In order to achieve our objective, we directed our attention to measuring all valuable information for SEMG. This work proposes an application method of two simple linear models, and the selection method of an optimal electrode configuration to use them effectively. Our system required four SEMG measurement electrodes in which locations differed for every subject depending on the individual's characteristics, and those were selected from a 96ch multi electrode using the Monte Carlo method. From the experimental results, the performance in six normal subjects indicated that the recognition rate of four motions were perfect and the grasp force estimated result fit well with the actual measurement result.

Right- and left-brain hemisphere. Rhythm in reaction time to light signals is task-load-dependent: age, gender, and handgrip strength rhythm comparisons.

PubMed

Reinberg, Alain; Bicakova-Rocher, Alena; Mechkouri, Mohamed; Ashkenazi, Israel

2002-11-01

In healthy mature subjects simple reaction time (SRT) to a single light signal (an easy task) is associated with a prominent rhythm with tau = 24 h of dominant (DH) as well as nondominant (NDH) hand performance, while three-choice reaction time (CRT), a complex task, is associated with tau = 24 h of the DH but tau < 24 h of the NDH. The aims of the study were to assess the influence of age and gender on the difference in tau of the NDH and DH, as it relates to the corresponding cortical hemisphere of the brain, in comparison to the rhythm in handgrip strength. Healthy subjects, 9 (5 M and 4 F) adolescents 10-16 yr of age and 15 (8 M and 7 F) adults 18-67 yr of age, active between 08:00 +/- 1 h and 23:00 +/- 1:30 h and free of alcohol, tobacco, and drug consumption volunteered. Data were gathered longitudinally at home and work 4-7 times daily for 11-20 d. At each test time the following variables were assessed: grip strength of both hands (Dynamometer: Colin-Gentile, Paris, France); single reaction time to a yellow signal (SRT); and CRT to randomized yellow, red, or green signal series with varying instruction from test to test (Psycholog-24: Biophyderm, France). Rhythms in the performance in SRT, CRT, and handgrip strength of both DH and NDH were explored. The sleep-wake rhythm was assessed by sleep-logs, and in a subset of 14 subjects it was also assessed by wrist actigraphy (Mini-Motionlogger: AMI, Ardsley NY). Exploration of the prominent period tau of time series was achieved by a special power spectra analysis for unequally spaced data. Cosinor analysis was used to quantify the rhythm amplitude A and rhythm-adjusted mean M of the power spectral analysis determined trial tau. A 24h sleep-wake rhythm was detected in almost all cases. In adults, a prominent tau of 24 h characterized the performance of the easy task by both the DH and NDH. In adults a prominent tau of 24 h was also detected in the complex CRT task performed by the DH, but for the NDH the tau was

Tonic ubiquitylation controls T-cell receptor:CD3 complex expression during T-cell development.

PubMed

Wang, Haopeng; Holst, Jeff; Woo, Seng-Ryong; Guy, Cliff; Bettini, Matt; Wang, Yao; Shafer, Aaron; Naramura, Mayumi; Mingueneau, Michaël; Dragone, Leonard L; Hayes, Sandra M; Malissen, Bernard; Band, Hamid; Vignali, Dario A A

2010-04-07

Expression of the T-cell receptor (TCR):CD3 complex is tightly regulated during T-cell development. The mechanism and physiological role of this regulation are unclear. Here, we show that the TCR:CD3 complex is constitutively ubiquitylated in immature double positive (DP) thymocytes, but not mature single positive (SP) thymocytes or splenic T cells. This steady state, tonic CD3 monoubiquitylation is mediated by the CD3varepsilon proline-rich sequence, Lck, c-Cbl, and SLAP, which collectively trigger the dynamin-dependent downmodulation, lysosomal sequestration and degradation of surface TCR:CD3 complexes. Blocking this tonic ubiquitylation by mutating all the lysines in the CD3 cytoplasmic tails significantly upregulates TCR levels on DP thymocytes. Mimicking monoubiquitylation by expression of a CD3zeta-monoubiquitin (monoUb) fusion molecule significantly reduces TCR levels on immature thymocytes. Moreover, modulating CD3 ubiquitylation alters immunological synapse (IS) formation and Erk phosphorylation, thereby shifting the signalling threshold for positive and negative selection, and regulatory T-cell development. Thus, tonic TCR:CD3 ubiquitylation results in precise regulation of TCR expression on immature T cells, which is required to maintain the fidelity of T-cell development.

Tonic ubiquitylation controls T-cell receptor:CD3 complex expression during T-cell development

PubMed Central

Wang, Haopeng; Holst, Jeff; Woo, Seng-Ryong; Guy, Cliff; Bettini, Matt; Wang, Yao; Shafer, Aaron; Naramura, Mayumi; Mingueneau, Michaël; Dragone, Leonard L; Hayes, Sandra M; Malissen, Bernard; Band, Hamid; Vignali, Dario A A

2010-01-01

Expression of the T-cell receptor (TCR):CD3 complex is tightly regulated during T-cell development. The mechanism and physiological role of this regulation are unclear. Here, we show that the TCR:CD3 complex is constitutively ubiquitylated in immature double positive (DP) thymocytes, but not mature single positive (SP) thymocytes or splenic T cells. This steady state, tonic CD3 monoubiquitylation is mediated by the CD3ɛ proline-rich sequence, Lck, c-Cbl, and SLAP, which collectively trigger the dynamin-dependent downmodulation, lysosomal sequestration and degradation of surface TCR:CD3 complexes. Blocking this tonic ubiquitylation by mutating all the lysines in the CD3 cytoplasmic tails significantly upregulates TCR levels on DP thymocytes. Mimicking monoubiquitylation by expression of a CD3ζ-monoubiquitin (monoUb) fusion molecule significantly reduces TCR levels on immature thymocytes. Moreover, modulating CD3 ubiquitylation alters immunological synapse (IS) formation and Erk phosphorylation, thereby shifting the signalling threshold for positive and negative selection, and regulatory T-cell development. Thus, tonic TCR:CD3 ubiquitylation results in precise regulation of TCR expression on immature T cells, which is required to maintain the fidelity of T-cell development. PMID:20150895

Thin film materials and devices for resistive temperature sensing applications

NASA Astrophysics Data System (ADS)

Basantani, Hitesh A.

Thin films of vanadium oxide (VOx) and hydrogenated amorphous silicon (a-Si:H) are the two dominant material systems used in resistive infrared radiation detectors (microbolometers) for sensing long wave infrared (LWIR) wavelengths in the 8--14 microm range. Typical thin films of VO x (x < 2) currently used in the bolometer industry have a magnitude of temperature coefficient of resistance (TCR) between 2%/K -- 3%/K. In contrast, thin films of hydrogenated germanium (SiGe:H) have |TCR| between 3%/K to 4%/K. Devices made from either of these materials have resulted in similar device performance with NETD ≈ 25 mK. The performance of the microbolometers is limited by the electronic noise, especially 1/f noise. Therefore, regardless of the choice of bolometer sensing material and read out circuitry, manufacturers are constantly striving to reduce 1/f noise while simultaneously increasing TCR to give better signal to noise ratios in their bolometers and ultimately, better image quality with more thermal information to the end user. In this work, thin films of VOx and hydrogenated germanium (Ge:H), having TCR values > 4 %/K are investigated as potential candidates for higher sensitivity next generation of microbolometers. Thin films of VO x were deposited by Biased Target Ion Beam Deposition (BTIBD) (˜85 nm thick). Electrical characterization of lateral resistor structures showed resistivity ranging from 104 O--cm to 2.1 x 104 O--cm, TCR varying from --4%/K to --5%/K, normalized Hooge parameter (alphaH/n) of 5 x 10 -21 to 5 x 10-18 cm3. Thin films of Ge:H were deposited by plasma enhanced chemical vapor deposition (PECVD) by incorporating an increasing amount of crystal fraction in the growing thin films. Thin films of Ge:H having a mixed phase, amorphous + nanocrystalline, having a |TCR| > 6 %/K were deposited with resistivity < 2,300 O--cm and a normalized Hooge's parameter 'alphaH/n' < 2 x 10-20 cm3. Higher TCR materials are desired, however, such materials have

Graphene nanoribbon field effect transistor for nanometer-size on-chip temperature sensor

NASA Astrophysics Data System (ADS)

Banadaki, Yaser M.; Srivastava, Ashok; Sharifi, Safura

2016-04-01

Graphene has been extensively investigated as a promising material for various types of high performance sensors due to its large surface-to-volume ratio, remarkably high carrier mobility, high carrier density, high thermal conductivity, extremely high mechanical strength and high signal-to-noise ratio. The power density and the corresponding die temperature can be tremendously high in scaled emerging technology designs, urging the on-chip sensing and controlling of the generated heat in nanometer dimensions. In this paper, we have explored the feasibility of a thin oxide graphene nanoribbon (GNR) as nanometer-size temperature sensor for detecting local on-chip temperature at scaled bias voltages of emerging technology. We have introduced an analytical model for GNR FET for 22nm technology node, which incorporates both thermionic emission of high-energy carriers and band-to-band-tunneling (BTBT) of carriers from drain to channel regions together with different scattering mechanisms due to intrinsic acoustic phonons and optical phonons and line-edge roughness in narrow GNRs. The temperature coefficient of resistivity (TCR) of GNR FET-based temperature sensor shows approximately an order of magnitude higher TCR than large-area graphene FET temperature sensor by accurately choosing of GNR width and bias condition for a temperature set point. At gate bias VGS = 0.55 V, TCR maximizes at room temperature to 2.1×10-2 /K, which is also independent of GNR width, allowing the design of width-free GNR FET for room temperature sensing applications.

Received Signal Strength Recovery in Green WLAN Indoor Positioning System Using Singular Value Thresholding

PubMed Central

Ma, Lin; Xu, Yubin

2015-01-01

Green WLAN is a promising technique for accessing future indoor Internet services. It is designed not only for high-speed data communication purposes but also for energy efficiency. The basic strategy of green WLAN is that all the access points are not always powered on, but rather work on-demand. Though powering off idle access points does not affect data communication, a serious asymmetric matching problem will arise in a WLAN indoor positioning system due to the fact the received signal strength (RSS) readings from the available access points are different in their offline and online phases. This asymmetry problem will no doubt invalidate the fingerprint algorithm used to estimate the mobile device location. Therefore, in this paper we propose a green WLAN indoor positioning system, which can recover RSS readings and achieve good localization performance based on singular value thresholding (SVT) theory. By solving the nuclear norm minimization problem, SVT recovers not only the radio map, but also online RSS readings from a sparse matrix by sensing only a fraction of the RSS readings. We have implemented the method in our lab and evaluated its performances. The experimental results indicate the proposed system could recover the RSS readings and achieve good localization performance. PMID:25587977

Impaired thymic selection in mice expressing altered levels of the SLP-76 adaptor protein.

PubMed

Ramsey, Kimberley; Luckashenak, Nancy; Koretzky, Gary A; Clements, James L

2008-02-01

Intracellular signaling initiated by ligation of the TCR influences cell fate at multiple points during the lifespan of a T cell. This is especially evident during thymic selection, where the nature of TCR-dependent signaling helps to establish a MHC-restricted, self-tolerant T cell repertoire. The Src homology 2 domain-containing leukocyte-specific phosphoprotein of 76 kDa (SLP-76) adaptor protein is a required intermediate in multiple signaling pathways triggered by TCR engagement, several of which have been implicated in dictating the outcome of thymic selection (e.g., intracellular calcium flux and activation of ERK family MAPKs). To determine if thymocyte maturation and selection at later stages of development are sensitive to perturbations in SLP-76 levels, we analyzed these crucial events using several transgenic (Tg) lines of mice expressing altered levels of SLP-76 in the thymus. In Tg mice expressing low levels of SLP-76 in preselection thymocytes, the CD4:CD8 ratio in the thymus and spleen was skewed in a manner consistent with impaired selection and/or maturation of CD4+ thymocytes. Low SLP-76 expression also correlated with reduced CD5 expression on immature thymocytes, consistent with reduced TCR signaling potential. In contrast, reconstitution of SLP-76 at higher levels resulted in normal thymic CD5 expression and CD4:CD8 ratios in the thymus and periphery. It is curious that thymic deletion of TCR-Tg (HY) thymocytes was markedly impaired in both lines of Tg-reconstituted SLP-76-/- mice. Studies using chimeric mice indicate that the defect in deletion of HY+ thymocytes is intrinsic to the developing thymocyte, suggesting that maintenance of sufficient SLP-76 expression from the endogenous locus is a key element in the selection process.

Collective signaling behavior in a networked-oscillator model

NASA Astrophysics Data System (ADS)

Liu, Z.-H.; Hui, P. M.

2007-09-01

We propose and study the collective behavior of a model of networked signaling objects that incorporates several ingredients of real-life systems. These ingredients include spatial inhomogeneity with grouping of signaling objects, signal attenuation with distance, and delayed and impulsive coupling between non-identical signaling objects. Depending on the coupling strength and/or time-delay effect, the model exhibits completely, partially, and locally collective signaling behavior. In particular, a correlated signaling (CS) behavior is observed in which there exist time durations when nearly a constant fraction of oscillators in the system are in the signaling state. These time durations are much longer than the duration of a spike when a single oscillator signals, and they are separated by regular intervals in which nearly all oscillators are silent. Such CS behavior is similar to that observed in biological systems such as fireflies, cicadas, crickets, and frogs. The robustness of the CS behavior against noise is also studied. It is found that properly adjusting the coupling strength and noise level could enhance the correlated behavior.

R2NA: Received Signal Strength (RSS) Ratio-Based Node Authentication for Body Area Network

PubMed Central

Wu, Yang; Wang, Kai; Sun, Yongmei; Ji, Yuefeng

2013-01-01

The body area network (BAN) is an emerging branch of wireless sensor networks for personalized applications. The services in BAN usually have a high requirement on security, especially for the medical diagnosis. One of the fundamental directions to ensure security in BAN is how to provide node authentication. Traditional research using cryptography relies on prior secrets shared among nodes, which leads to high resource cost. In addition, most existing non-cryptographic solutions exploit out-of-band (OOB) channels, but they need the help of additional hardware support or significant modifications to the system software. To avoid the above problems, this paper presents a proximity-based node authentication scheme, which only uses wireless modules equipped on sensors. With only one sensor and one control unit (CU) in BAN, we could detect a unique physical layer characteristic, namely, the difference between the received signal strength (RSS) measured on different devices in BAN. Through the above-mentioned particular difference, we can tell whether the sender is close enough to be legitimate. We validate our scheme through both theoretical analysis and experiments, which are conducted on the real Shimmer nodes. The results demonstrate that our proposed scheme has a good security performance.

Signalling through NK1.1 triggers NK cells to die but induces NK T cells to produce interleukin-4.

PubMed

Asea, A; Stein-Streilein, J

1998-02-01

In vivo inoculation of specific antibody is an accepted protocol for elimination of specific cell populations. Except for anti-CD3 and anti-CD4, it is not known if the depleted cells are eliminated by signalling through the target molecule or through a more non-specific mechanism. C57BL/6 mice were inoculated with anti-natural killer (NK1.1) monoclonal antibody (mAb). Thereafter spleen cells were harvested, stained for both surface and intracellular markers, and analysed by flow cytometry. As early as 2 hr post inoculation, NK cells were signalled to become apoptotic while signalling through the NK1.1 molecule activated NK1.1+ T-cell receptor (TCR)+ (NK T) cells to increase in number, and produce interleukin-4 (IL-4). Anti NK1.1 mAb was less efficient at signalling apoptosis in NK cells when NK T-cell deficient [beta 2-microglobulin beta 2m-deficient] mice were used compared with wild type mice. Efficient apoptotic signalling was restored when beta 2m-deficient mice were reconstituted with NK T cells. NK-specific antibody best signals the apoptotic process in susceptible NK cells when resistant NK T cells are present, activated, and secrete IL-4.

Noncore RAG1 regions promote Vβ rearrangements and αβ T cell development by overcoming inherent inefficiency of Vβ recombination signal sequences.

PubMed

Horowitz, Julie E; Bassing, Craig H

2014-02-15

The RAG proteins are comprised of core endonuclease domains and noncore regions that modulate endonuclease activity. Mutation or deletion of noncore RAG regions in humans causes immunodeficiency and altered TCR repertoire, and mice expressing core but not full-length Rag1 (Rag1(C/C)) or Rag2 (Rag2(C/C)) exhibit lymphopenia, reflecting impaired V(D)J recombination and lymphocyte development. Rag1(C/C) mice display reduced D-to-J and V-to-DJ rearrangements of TCRβ and IgH loci, whereas Rag2(C/C) mice show decreased V-to-DJ rearrangements and altered Vβ/VH repertoire. Because Vβs/VHs only recombine to DJ complexes, the Rag1(C/C) phenotype could reflect roles for noncore RAG1 regions in promoting recombination during only the D-to-J step or during both steps. In this study, we demonstrate that a preassembled TCRβ gene, but not a preassembled DβJβ complex or the prosurvival BCL2 protein, completely rescues αβ T cell development in Rag1(C/C) mice. We find that Rag1(C/C) mice exhibit altered Vβ utilization in Vβ-to-DJβ rearrangements, increased usage of 3'Jα gene segments in Vα-to-Jα rearrangements, and abnormal changes in Vβ repertoire during αβ TCR selection. Inefficient Vβ/VH recombination signal sequences (RSSs) have been hypothesized to cause impaired V-to-DJ recombination on the background of a defective recombinase as in core-Rag mice. We show that replacement of the Vβ14 RSS with a more efficient RSS increases Vβ14 recombination and rescues αβ T cell development in Rag1(C/C) mice. Our data indicate that noncore RAG1 regions establish a diverse TCR repertoire by overcoming Vβ RSS inefficiency to promote Vβ recombination and αβ T cell development, and by modulating TCRβ and TCRα gene segment utilization.

A Proteomic View at T Cell Costimulation

PubMed Central

Hombach, Andreas A.; Recktenwald, Christian V.; Dressler, Sven P.; Abken, Hinrich; Seliger, Barbara

2012-01-01

The “two-signal paradigm” in T cell activation predicts that the cooperation of “signal 1,” provided by the T cell receptor (TCR) through engagement of major histocompatility complex (MHC)-presented peptide, with “signal 2″ provided by costimulatory molecules, the prototype of which is CD28, is required to induce T cell effector functions. While the individual signalling pathways are well understood, little is known about global changes in the proteome pattern during TCR/CD28-mediated activation. Therefore, comparative 2-DE-based proteome analyses of CD3+ CD69- resting T cells versus cells incubated with (i) the agonistic anti-CD3 antibody OKT3 mimicking signal 1 in absence or presence of IL-2 and/or with (ii) the agonistic antibody 15E8 triggering CD28-mediated signaling were performed. Differentially regulated spots were defined leading to the identification of proteins involved in the regulation of the metabolism, shaping and maintenance of the cytoskeleton and signal transduction. Representative members of the differentially expressed protein families, such as calmodulin (CALM), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), L-lactate dehydrogenase (LDH), Rho GDP-dissociation inhibitor 2 (GDIR2), and platelet basic protein (CXCL7), were independently verified by flow cytometry. Data provide a detailed map of individual protein alterations at the global proteome level in response to TCR/CD28-mediated T cell activation. PMID:22539942

Self-Recognition Sensitizes Mouse and Human Regulatory T Cells to Low-Dose CD28 Superagonist Stimulation.

PubMed

Langenhorst, Daniela; Tabares, Paula; Gulde, Tobias; Becklund, Bryan R; Berr, Susanne; Surh, Charles D; Beyersdorf, Niklas; Hünig, Thomas

2017-01-01

In rodents, low doses of CD28-specific superagonistic monoclonal antibodies (CD28 superagonists, CD28SA) selectively activate regulatory T cells (Treg). This observation has recently been extended to humans, suggesting an option for the treatment of autoimmune and inflammatory diseases. However, a mechanistic explanation for this phenomenon is still lacking. Given that CD28SA amplify T cell receptor (TCR) signals, we tested the hypothesis that the weak tonic TCR signals received by conventional CD4 + T cells (Tconv) in the absence of cognate antigen require more CD28 signaling input for full activation than the stronger TCR signals received by self-reactive Treg. We report that in vitro , the response of mouse Treg and Tconv to CD28SA strongly depends on MHC class II expression by antigen-presenting cells. To separate the effect of tonic TCR signals from self-peptide recognition, we compared the response of wild-type Treg and Tconv to low and high CD28SA doses upon transfer into wild-type or H-2M knockout mice, which lack a self-peptide repertoire. We found that the superior response of Treg to low CD28SA doses was lost in the absence of self-peptide presentation. We also tested if potentially pathogenic autoreactive Tconv would benefit from self-recognition-induced sensitivity to CD28SA stimulation by transferring TCR transgenic OVA-specific Tconv into OVA-expressing mice and found that low-dose CD28SA application inhibited, rather than supported, their expansion, presumably due to the massive concomitant activation of Treg. Finally, we report that also in the in vitro response of human peripheral blood mononuclear cells to CD28SA, HLA II blockade interferes with the expansion of Treg by low-dose CD28SA stimulation. These results provide a rational basis for the further development of low-dose CD28SA therapy for the improvement of Treg activity.

Oral-resident natural Th17 cells and γδ T cells control opportunistic Candida albicans infections.

PubMed

Conti, Heather R; Peterson, Alanna C; Brane, Lucas; Huppler, Anna R; Hernández-Santos, Nydiaris; Whibley, Natasha; Garg, Abhishek V; Simpson-Abelson, Michelle R; Gibson, Gregory A; Mamo, Anna J; Osborne, Lisa C; Bishu, Shrinivas; Ghilardi, Nico; Siebenlist, Ulrich; Watkins, Simon C; Artis, David; McGeachy, Mandy J; Gaffen, Sarah L

2014-09-22

Oropharyngeal candidiasis (OPC) is an opportunistic fungal infection caused by Candida albicans. OPC is frequent in HIV/AIDS, implicating adaptive immunity. Mice are naive to Candida, yet IL-17 is induced within 24 h of infection, and susceptibility is strongly dependent on IL-17R signaling. We sought to identify the source of IL-17 during the early innate response to candidiasis. We show that innate responses to Candida require an intact TCR, as SCID, IL-7Rα(-/-), and Rag1(-/-) mice were susceptible to OPC, and blockade of TCR signaling by cyclosporine induced susceptibility. Using fate-tracking IL-17 reporter mice, we found that IL-17 is produced within 1-2 d by tongue-resident populations of γδ T cells and CD3(+)CD4(+)CD44(hi)TCRβ(+)CCR6(+) natural Th17 (nTh17) cells, but not by TCR-deficient innate lymphoid cells (ILCs) or NK cells. These cells function redundantly, as TCR-β(-/-) and TCR-δ(-/-) mice were both resistant to OPC. Whereas γδ T cells were previously shown to produce IL-17 during dermal candidiasis and are known to mediate host defense at mucosal surfaces, nTh17 cells are poorly understood. The oral nTh17 population expanded rapidly after OPC, exhibited high TCR-β clonal diversity, and was absent in Rag1(-/-), IL-7Rα(-/-), and germ-free mice. These findings indicate that nTh17 and γδ T cells, but not ILCs, are key mucosal sentinels that control oral pathogens. © 2014 Conti et al.

An Improved Compressive Sensing and Received Signal Strength-Based Target Localization Algorithm with Unknown Target Population for Wireless Local Area Networks.

PubMed

Yan, Jun; Yu, Kegen; Chen, Ruizhi; Chen, Liang

2017-05-30

In this paper a two-phase compressive sensing (CS) and received signal strength (RSS)-based target localization approach is proposed to improve position accuracy by dealing with the unknown target population and the effect of grid dimensions on position error. In the coarse localization phase, by formulating target localization as a sparse signal recovery problem, grids with recovery vector components greater than a threshold are chosen as the candidate target grids. In the fine localization phase, by partitioning each candidate grid, the target position in a grid is iteratively refined by using the minimum residual error rule and the least-squares technique. When all the candidate target grids are iteratively partitioned and the measurement matrix is updated, the recovery vector is re-estimated. Threshold-based detection is employed again to determine the target grids and hence the target population. As a consequence, both the target population and the position estimation accuracy can be significantly improved. Simulation results demonstrate that the proposed approach achieves the best accuracy among all the algorithms compared.

Specific recycling receptors are targeted to the immune synapse by the intraflagellar transport system

PubMed Central

Finetti, Francesca; Patrussi, Laura; Masi, Giulia; Onnis, Anna; Galgano, Donatella; Lucherini, Orso Maria; Pazour, Gregory J.; Baldari, Cosima T.

2014-01-01

ABSTRACT T cell activation requires sustained signaling at the immune synapse, a specialized interface with the antigen-presenting cell (APC) that assembles following T cell antigen receptor (TCR) engagement by major histocompatibility complex (MHC)-bound peptide. Central to sustained signaling is the continuous recruitment of TCRs to the immune synapse. These TCRs are partly mobilized from an endosomal pool by polarized recycling. We have identified IFT20, a component of the intraflagellar transport (IFT) system that controls ciliogenesis, as a central regulator of TCR recycling to the immune synapse. Here, we have investigated the interplay of IFT20 with the Rab GTPase network that controls recycling. We found that IFT20 forms a complex with Rab5 and the TCR on early endosomes. IFT20 knockdown (IFT20KD) resulted in a block in the recycling pathway, leading to a build-up of recycling TCRs in Rab5+ endosomes. Recycling of the transferrin receptor (TfR), but not of CXCR4, was disrupted by IFT20 deficiency. The IFT components IFT52 and IFT57 were found to act together with IFT20 to regulate TCR and TfR recycling. The results provide novel insights into the mechanisms that control TCR recycling and immune synapse assembly, and underscore the trafficking-related function of the IFT system beyond ciliogenesis. PMID:24554435

T-Cell Receptor- and CD28-induced Vav1 activity is required for the accumulation of primed T cells into antigenic tissue

PubMed Central

David, Rachel; Ma, Liang; Ivetic, Aleksandar; Takesono, Aya; Ridley, Anne J.; Chai, Jian-Guo; Tybulewicz, Victor; Marelli-Berg, Federica M.

2016-01-01

Localization of primed T cells to antigenic tissue is essential for the development of effective immunity. Together with tissue-selective homing molecules, T-cell receptor (TCR)- and CD28-mediated signals have been shown to promote transendothelial migration of specific T cells into non-lymphoid antigen-rich tissue tissue. However, the cellular and molecular requirements for T-cell accumulation to target tissue following their recruitment are largely undefined. The guanine nucleotide exchange factor (GEF) Vav1 has an integral role in coupling TCR and CD28 to signalling pathways that regulate T cell activation and migration. Here, we have investigated the contribution of TCR- and CD28-induced Vav1 activity to the trafficking and localization of primed HY-specific CD4+ T cells to antigenic sites. Severe migratory defects displayed by Vav1-/- T cells in vitro were fully compensated by a combination of shear flow and chemokines, leading to normal recruitment of Vav1-/- T cells in vivo. In contrast, Vav1-/- T-cell retention into antigen-rich tissue was severely impaired, reflecting their inability to engage in sustained TCR- and CD28-mediated interactions with tissue-resident antigen-presenting cells (APCs). This novel function of APC-induced, TCR- and CD28-mediated Vav1 activity in the regulation of effector T-cell immunity highlights its potential as a therapeutic target in T-cell-mediated tissue damage. PMID:19060239

TCR hypervariable regions expressed by T cells that respond to effective tumor vaccines.

PubMed

Jordan, Kimberly R; Buhrman, Jonathan D; Sprague, Jonathan; Moore, Brandon L; Gao, Dexiang; Kappler, John W; Slansky, Jill E

2012-10-01

A major goal of immunotherapy for cancer is the activation of T cell responses against tumor-associated antigens (TAAs). One important strategy for improving antitumor immunity is vaccination with peptide variants of TAAs. Understanding the mechanisms underlying the expansion of T cells that respond to the native tumor antigen is an important step in developing effective peptide-variant vaccines. Using an immunogenic mouse colon cancer model, we compare the binding properties and the TCR genes expressed by T cells elicited by peptide variants that elicit variable antitumor immunity directly ex vivo. The steady-state affinity of the natural tumor antigen for the T cells responding to effective peptide vaccines was higher relative to ineffective peptides, consistent with their improved function. Ex vivo analysis showed that T cells responding to the effective peptides expressed a CDR3β motif, which was also shared by T cells responding to the natural antigen and not those responding to the less effective peptide vaccines. Importantly, these data demonstrate that peptide vaccines can expand T cells that naturally respond to tumor antigens, resulting in more effective antitumor immunity. Future immunotherapies may require similar stringent analysis of the responding T cells to select optimal peptides as vaccine candidates.

T Cell Receptor Engagement Triggers Its CD3ε and CD3ζ Subunits to Adopt a Compact, Locked Conformation

PubMed Central

Risueño, Ruth M.; Schamel, Wolfgang W. A.; Alarcón, Balbino

2008-01-01

How the T cell antigen receptor (TCR) discriminates between molecularly related peptide/Major Histocompatibility Complex (pMHC) ligands and converts this information into different possible signaling outcomes is still not understood. One current model proposes that strong pMHC ligands, but not weak ones, induce a conformational change in the TCR. Evidence supporting this comes from a pull-down assay that detects ligand-induced binding of the TCR to the N-terminal SH3 domain of the adapter protein Nck, and also from studies with a neoepitope-specific antibody. Both methods rely on the exposure of a polyproline sequence in the CD3ε subunit of the TCR, and neither indicates whether the conformational change is transmitted to other CD3 subunits. Using a protease-sensitivity assay, we now show that the cytoplasmic tails of CD3ε and CD3ζ subunits become fully protected from degradation upon TCR triggering. These results suggest that the TCR conformational change is transmitted to the tails of CD3ε and CD3ζ, and perhaps all CD3 subunits. Furthermore, the resistance to protease digestion suggests that CD3 cytoplasmic tails adopt a compact structure in the triggered TCR. These results are consistent with a model in which transduction of the conformational change induced upon TCR triggering promotes condensation and shielding of the CD3 cytoplasmic tails. PMID:18320063

Effect of pulse sequence parameter selection on signal strength in positive-contrast MRI markers for MRI-based prostate postimplant assessment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lim, Tze Yee

Purpose: For postimplant dosimetric assessment, computed tomography (CT) is commonly used to identify prostate brachytherapy seeds, at the expense of accurate anatomical contouring. Magnetic resonance imaging (MRI) is superior to CT for anatomical delineation, but identification of the negative-contrast seeds is challenging. Positive-contrast MRI markers were proposed to replace spacers to assist seed localization on MRI images. Visualization of these markers under varying scan parameters was investigated. Methods: To simulate a clinical scenario, a prostate phantom was implanted with 66 markers and 86 seeds, and imaged on a 3.0T MRI scanner using a 3D fast radiofrequency-spoiled gradient recalled echo acquisitionmore » with various combinations of scan parameters. Scan parameters, including flip angle, number of excitations, bandwidth, field-of-view, slice thickness, and encoding steps were systematically varied to study their effects on signal, noise, scan time, image resolution, and artifacts. Results: The effects of pulse sequence parameter selection on the marker signal strength and image noise were characterized. The authors also examined the tradeoff between signal-to-noise ratio, scan time, and image artifacts, such as the wraparound artifact, susceptibility artifact, chemical shift artifact, and partial volume averaging artifact. Given reasonable scan time and managable artifacts, the authors recommended scan parameter combinations that can provide robust visualization of the MRI markers. Conclusions: The recommended MRI pulse sequence protocol allows for consistent visualization of the markers to assist seed localization, potentially enabling MRI-only prostate postimplant dosimetry.« less

Weakly self-reactive T-cell clones can homeostatically expand when present at low numbers.

PubMed

Vrisekoop, Nienke; Artusa, Patricio; Monteiro, Joao P; Mandl, Judith N

2017-01-01

T-cell division is central to maintaining a stable T-cell pool in adults. It also enables T-cell expansion in neonates, and after depletion by chemotherapy, bone marrow transplantation, or infection. The same signals required for T-cell survival in lymphoreplete settings, IL-7 and T-cell receptor (TCR) interactions with self-peptide MHC (pMHC), induce division when T-cell numbers are low. The strength of reactivity for self-pMHC has been shown to correlate with the capacity of T cells to undergo lymphopenia-induced proliferation (LIP), in that weakly self-reactive T cells are unable to divide, implying that T-cell reconstitution would significantly skew the TCR repertoire toward TCRs with greater self-reactivity and thus compromise T-cell diversity. Here, we show that while CD4 + T cells with low self-pMHC reactivity experience more intense competition, they are able to divide when present at low enough cell numbers. Thus, at physiological precursor frequencies CD4 + T cells with low self-pMHC reactivity are able to contribute to the reconstitution of the T-cell pool. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

T-cell receptor revision: friend or foe?

PubMed Central

Hale, J Scott; Fink, Pamela J

2010-01-01

T-cell receptor (TCR) revision is a process of tolerance induction by which peripheral T cells lose surface expression of an autoreactive TCR, reinduce expression of the recombinase machinery, rearrange genes encoding extrathymically generated TCRs for antigen, and express these new receptors on the cell surface. We discuss the evidence for this controversial tolerance mechanism below. Despite the apparent heresy of post-thymic gene rearrangement, we argue here that TCR revision follows the rules obeyed by maturing thymocytes undergoing gene recombination. Expression of the recombinase is carefully controlled both spatially and temporally, and may be initiated by loss of signals through surface TCRs. The resulting TCR repertoire is characterized by its diversity, self major histocompatibility complex restriction, self tolerance, and ability to mount productive immune responses specific for foreign antigens. Hence, TCR revision is a carefully regulated process of tolerance induction that can contribute to the protection of the individual against invading pathogens while preserving the integrity of self tissue. PMID:20201984

T-cell receptor revision: friend or foe?

PubMed

Hale, J Scott; Fink, Pamela J

2010-04-01

T-cell receptor (TCR) revision is a process of tolerance induction by which peripheral T cells lose surface expression of an autoreactive TCR, reinduce expression of the recombinase machinery, rearrange genes encoding extrathymically generated TCRs for antigen, and express these new receptors on the cell surface. We discuss the evidence for this controversial tolerance mechanism below. Despite the apparent heresy of post-thymic gene rearrangement, we argue here that TCR revision follows the rules obeyed by maturing thymocytes undergoing gene recombination. Expression of the recombinase is carefully controlled both spatially and temporally, and may be initiated by loss of signals through surface TCRs. The resulting TCR repertoire is characterized by its diversity, self major histocompatibility complex restriction, self tolerance, and ability to mount productive immune responses specific for foreign antigens. Hence, TCR revision is a carefully regulated process of tolerance induction that can contribute to the protection of the individual against invading pathogens while preserving the integrity of self tissue.

Magnetic Field-Induced T Cell Receptor Clustering by Nanoparticles Enhances T Cell Activation and Stimulates Antitumor Activity

PubMed Central

2015-01-01

Iron–dextran nanoparticles functionalized with T cell activating proteins have been used to study T cell receptor (TCR) signaling. However, nanoparticle triggering of membrane receptors is poorly understood and may be sensitive to physiologically regulated changes in TCR clustering that occur after T cell activation. Nano-aAPC bound 2-fold more TCR on activated T cells, which have clustered TCR, than on naive T cells, resulting in a lower threshold for activation. To enhance T cell activation, a magnetic field was used to drive aggregation of paramagnetic nano-aAPC, resulting in a doubling of TCR cluster size and increased T cell expansion in vitro and after adoptive transfer in vivo. T cells activated by nano-aAPC in a magnetic field inhibited growth of B16 melanoma, showing that this novel approach, using magnetic field-enhanced nano-aAPC stimulation, can generate large numbers of activated antigen-specific T cells and has clinically relevant applications for adoptive immunotherapy. PMID:24564881

Differences In Early T-Cell Signaling In Cultures Grown In a Rotating Clinostat vs. Static Controls

NASA Technical Reports Server (NTRS)

Alexamder. M.; Nelman-Gonzales, M.; Penkala, J.; Sams, C.

1999-01-01

Altered gravity has previously been demonstrated to be a stress that can influence components of the immune system. Specifically, T-cell activation has been shown to be affected by changes in gravity, exhibiting a decrease in proliferative response to in vitro stimulation in microgravity. Subsequent ground based studies utilizing a rotating clinostat to model some of the effects of microgravity have been consistent with earlier flight based experiments. These ground and flight experiments have examined T-cell activation by measuring various responses including production of cytokines, DNA synthesis and the production of various cell surface activation markers. These indicators of T-cell activation were measured anywhere from 4 to 72 hours after stimulation. Prior to the work described here, the initial signaling events in T-cell activation had not been directly examined. The goal of this project was to determine how the process of early signal transduction was affected by growth in a rotating clinostat. Here we directly show a defect in signaling from TCR to MAPK in purified peripheral T-cells activated in the clinostat by OKT3/antiCD28 coated microbeads as compared to static controls.

Regulation of vesicular traffic at the T cell immune synapse: lessons from the primary cilium.

PubMed

Finetti, Francesca; Onnis, Anna; Baldari, Cosima T

2015-03-01

The signals that orchestrate the process of T cell activation are coordinated at the specialized interface that forms upon contact with an antigen presenting cell displaying a specific MHC-associated peptide ligand, known as the immune synapse. The central role of vesicular traffic in the assembly of the immune synapse has emerged only in recent years with the finding that sustained T-cell receptor (TCR) signaling involves delivery of TCR/CD3 complexes from an intracellular pool associated with recycling endosomes. A number of receptors as well as membrane-associated signaling mediators have since been demonstrated to exploit this process to localize to the immune synapse. Here, we will review our current understanding of the mechanisms responsible for TCR recycling, with a focus on the intraflagellar transport system, a multimolecular complex that is responsible for the assembly and function of the primary cilium which we have recently implicated in polarized endosome recycling to the immune synapse. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Context matters: sexual signaling loss in digital organisms

PubMed Central

Weigel, Emily G; Testa, Nicholas D; Peer, Alex; Garnett, Sara C

2015-01-01

Sexual signals are important in attracting and choosing mates; however, these signals and their associated preferences are often costly and frequently lost. Despite the prevalence of signaling system loss in many taxa, the factors leading to signal loss remain poorly understood. Here, we test the hypothesis that complexity in signal loss scenarios is due to the context-dependent nature of the many factors affecting signal loss itself. Using the Avida digital life platform, we evolved 50 replicates of ∼250 lineages, each with a unique combination of parameters, including whether signaling is obligate or facultative; genetic linkage between signaling and receiving genes; population size; and strength of preference for signals. Each of these factors ostensibly plays a crucial role in signal loss, but was found to do so only under specific conditions. Under obligate signaling, genetic linkage, but not population size, influenced signal loss; under facultative signaling, genetic linkage does not have significant influence. Somewhat surprisingly, only a total loss of preference in the obligate signaling populations led to total signal loss, indicating that even a modest amount of preference is enough to maintain signaling systems. Strength of preference proved to be the strongest single force preventing signal loss, as it consistently overcame the potential effects of drift within our study. Our findings suggest that signaling loss is often dependent on not just preference for signals, population size, and genetic linkage, but also whether signals are required to initiate mating. These data provide an understanding of the factors (and their interactions) that may facilitate the maintenance of sexual signals. PMID:26380700

Tunable signal processing in synthetic MAP kinase cascades.

PubMed

O'Shaughnessy, Ellen C; Palani, Santhosh; Collins, James J; Sarkar, Casim A

2011-01-07

The flexibility of MAPK cascade responses enables regulation of a vast array of cell fate decisions, but elucidating the mechanisms underlying this plasticity is difficult in endogenous signaling networks. We constructed insulated mammalian MAPK cascades in yeast to explore how intrinsic and extrinsic perturbations affect the flexibility of these synthetic signaling modules. Contrary to biphasic dependence on scaffold concentration, we observe monotonic decreases in signal strength as scaffold concentration increases. We find that augmenting the concentration of sequential kinases can enhance ultrasensitivity and lower the activation threshold. Further, integrating negative regulation and concentration variation can decouple ultrasensitivity and threshold from the strength of the response. Computational analyses show that cascading can generate ultrasensitivity and that natural cascades with different kinase concentrations are innately biased toward their distinct activation profiles. This work demonstrates that tunable signal processing is inherent to minimal MAPK modules and elucidates principles for rational design of synthetic signaling systems. Copyright © 2011 Elsevier Inc. All rights reserved.

NFκB–Pim-1–Eomesodermin axis is critical for maintaining CD8 T-cell memory quality

PubMed Central

Knudson, Karin M.; Saxena, Vikas; Altman, Amnon; Daniels, Mark A.; Teixeiro, Emma

2017-01-01

T-cell memory is critical for long-term immunity. However, the factors involved in maintaining the persistence, function, and phenotype of the memory pool are undefined. Eomesodermin (Eomes) is required for the establishment of the memory pool. Here, we show that in T cells transitioning to memory, the expression of high levels of Eomes is not constitutive but rather requires a continuum of cell-intrinsic NFκB signaling. Failure to maintain NFκB signals after the peak of the response led to impaired Eomes expression and a defect in the maintenance of CD8 T-cell memory. Strikingly, we found that antigen receptor [T-cell receptor (TCR)] signaling regulates this process through expression of the NFκB-dependent kinase proviral integration site for Moloney murine leukemia virus-1 (PIM-1), which in turn regulates NFκB and Eomes. T cells defective in TCR-dependent NFκB signaling were impaired in late expression of Pim-1, Eomes, and CD8 memory. These defects were rescued when TCR-dependent NFκB signaling was restored. We also found that NFκB–Pim-1 signals were required at memory to maintain memory CD8 T-cell longevity, effector function, and Eomes expression. Hence, an NFκB–Pim-1–Eomes axis regulates Eomes levels to maintain memory fitness. PMID:28193872

Spatial and Temporal Control of T Cell Activation Using a Photoactivatable Agonist.

PubMed

Sanchez, Elisa; Huse, Morgan

2018-04-25

T lymphocytes engage in rapid, polarized signaling, occurring within minutes following TCR activation. This induces formation of the immunological synapse, a stereotyped cell-cell junction that regulates T cell activation and directionally targets effector responses. To study these processes effectively, an imaging approach that is tailored to capturing fast, polarized responses is necessary. This protocol describes such a system, which is based on a photoactivatable peptide-major histocompatibility complex (pMHC) that is non-stimulatory until it is exposed to ultraviolet light. Targeted decaging of this reagent during videomicroscopy experiments enables precise spatiotemporal control of TCR activation and high-resolution monitoring of subsequent cellular responses by total internal reflection (TIRF) imaging. This approach is also compatible with genetic and pharmacological perturbation strategies. This allows for the assembly of well-defined molecular pathways that link TCR signaling to the formation of the polarized cytoskeletal structures that underlie the immunological synapse.

Inhibition of Canonical NF-κB Signaling by a Small Molecule Targeting NEMO-Ubiquitin Interaction

PubMed Central

Vincendeau, Michelle; Hadian, Kamyar; Messias, Ana C.; Brenke, Jara K.; Halander, Jenny; Griesbach, Richard; Greczmiel, Ute; Bertossi, Arianna; Stehle, Ralf; Nagel, Daniel; Demski, Katrin; Velvarska, Hana; Niessing, Dierk; Geerlof, Arie; Sattler, Michael; Krappmann, Daniel

2016-01-01

The IκB kinase (IKK) complex acts as the gatekeeper of canonical NF-κB signaling, thereby regulating immunity, inflammation and cancer. It consists of the catalytic subunits IKKα and IKKβ and the regulatory subunit NEMO/IKKγ. Here, we show that the ubiquitin binding domain (UBAN) in NEMO is essential for IKK/NF-κB activation in response to TNFα, but not IL-1β stimulation. By screening a natural compound library we identified an anthraquinone derivative that acts as an inhibitor of NEMO-ubiquitin binding (iNUB). Using biochemical and NMR experiments we demonstrate that iNUB binds to NEMOUBAN and competes for interaction with methionine-1-linked linear ubiquitin chains. iNUB inhibited NF-κB activation upon UBAN-dependent TNFα and TCR/CD28, but not UBAN-independent IL-1β stimulation. Moreover, iNUB was selectively killing lymphoma cells that are addicted to chronic B-cell receptor triggered IKK/NF-κB activation. Thus, iNUB disrupts the NEMO-ubiquitin protein-protein interaction interface and thereby inhibits physiological and pathological NF-κB signaling. PMID:26740240

Actin polymerization‐dependent activation of Cas‐L promotes immunological synapse stability

PubMed Central

Santos, Luís C; Blair, David A; Kumari, Sudha; Cammer, Michael; Iskratsch, Thomas; Herbin, Olivier; Alexandropoulos, Konstantina

2016-01-01

The immunological synapse formed between a T‐cell and an antigen‐presenting cell is important for cell–cell communication during T‐cell‐mediated immune responses. Immunological synapse formation begins with stimulation of the T‐cell receptor (TCR). TCR microclusters are assembled and transported to the center of the immunological synapse in an actin polymerization‐dependent process. However, the physical link between TCR and actin remains elusive. Here we show that lymphocyte‐specific Crk‐associated substrate (Cas‐L), a member of a force sensing protein family, is required for transport of TCR microclusters and for establishing synapse stability. We found that Cas‐L is phosphorylated at TCR microclusters in an actin polymerization‐dependent fashion. Furthermore, Cas‐L participates in a positive feedback loop leading to amplification of Ca2+ signaling, inside–out integrin activation, and actomyosin contraction. We propose a new role for Cas‐L in T‐cell activation as a mechanical transducer linking TCR microclusters to the underlying actin network and coordinating multiple actin‐dependent structures in the immunological synapse. Our studies highlight the importance of mechanotransduction processes in T‐cell‐mediated immune responses. PMID:27359298

Auditory Warnings, Signal-Referent Relations, and Natural Indicators: Re-Thinking Theory and Application

ERIC Educational Resources Information Center

Petocz, Agnes; Keller, Peter E.; Stevens, Catherine J.

2008-01-01

In auditory warning design the idea of the strength of the association between sound and referent has been pivotal. Research has proceeded via constructing classification systems of signal-referent associations and then testing predictions about ease of learning of different levels of signal-referent relation strength across and within different…

Oral-resident natural Th17 cells and γδ T cells control opportunistic Candida albicans infections

PubMed Central

Conti, Heather R.; Peterson, Alanna C.; Brane, Lucas; Huppler, Anna R.; Hernández-Santos, Nydiaris; Whibley, Natasha; Garg, Abhishek V.; Simpson-Abelson, Michelle R.; Gibson, Gregory A.; Mamo, Anna J.; Osborne, Lisa C.; Bishu, Shrinivas; Ghilardi, Nico; Siebenlist, Ulrich; Watkins, Simon C.; Artis, David; McGeachy, Mandy J.

2014-01-01

Oropharyngeal candidiasis (OPC) is an opportunistic fungal infection caused by Candida albicans. OPC is frequent in HIV/AIDS, implicating adaptive immunity. Mice are naive to Candida, yet IL-17 is induced within 24 h of infection, and susceptibility is strongly dependent on IL-17R signaling. We sought to identify the source of IL-17 during the early innate response to candidiasis. We show that innate responses to Candida require an intact TCR, as SCID, IL-7Rα−/−, and Rag1−/− mice were susceptible to OPC, and blockade of TCR signaling by cyclosporine induced susceptibility. Using fate-tracking IL-17 reporter mice, we found that IL-17 is produced within 1–2 d by tongue-resident populations of γδ T cells and CD3+CD4+CD44hiTCRβ+CCR6+ natural Th17 (nTh17) cells, but not by TCR-deficient innate lymphoid cells (ILCs) or NK cells. These cells function redundantly, as TCR-β−/− and TCR-δ−/− mice were both resistant to OPC. Whereas γδ T cells were previously shown to produce IL-17 during dermal candidiasis and are known to mediate host defense at mucosal surfaces, nTh17 cells are poorly understood. The oral nTh17 population expanded rapidly after OPC, exhibited high TCR-β clonal diversity, and was absent in Rag1−/−, IL-7Rα−/−, and germ-free mice. These findings indicate that nTh17 and γδ T cells, but not ILCs, are key mucosal sentinels that control oral pathogens. PMID:25200028

Co-potentiation of antigen recognition: A mechanism to boost weak T cell responses and provide immunotherapy in vivo

PubMed Central

Hoffmann, Michele M.; Molina-Mendiola, Carlos; Nelson, Alfreda D.; Parks, Christopher A.; Reyes, Edwin E.; Hansen, Michael J.; Rajagopalan, Govindarajan; Pease, Larry R.; Schrum, Adam G.; Gil, Diana

2015-01-01

Adaptive immunity is mediated by antigen receptors that can induce weak or strong immune responses depending on the nature of the antigen that is bound. In T lymphocytes, antigen recognition triggers signal transduction by clustering T cell receptor (TCR)/CD3 multiprotein complexes. In addition, it hypothesized that biophysical changes induced in TCR/CD3 that accompany receptor engagement may contribute to signal intensity. Nonclustering monovalent TCR/CD3 engagement is functionally inert despite the fact that it may induce changes in conformational arrangement or in the flexibility of receptor subunits. We report that the intrinsically inert monovalent engagement of TCR/CD3 can specifically enhance physiologic T cell responses to weak antigens in vitro and in vivo without stimulating antigen-unengaged T cells and without interrupting T cell responses to strong antigens, an effect that we term as “co-potentiation.” We identified Mono-7D6-Fab, which biophysically altered TCR/CD3 when bound and functionally enhanced immune reactivity to several weak antigens in vitro, including a gp100-derived peptide associated with melanoma. In vivo, Mono-7D6-Fab induced T cell antigen–dependent therapeutic responses against melanoma lung metastases, an effect that synergized with other anti-melanoma immunotherapies to significantly improve outcome and survival. We conclude that Mono-7D6-Fab directly co-potentiated TCR/CD3 engagement by weak antigens and that such concept can be translated into an immunotherapeutic design. The co-potentiation principle may be applicable to other receptors that could be regulated by otherwise inert compounds whose latent potency is only invoked in concert with specific physiologic ligands. PMID:26601285

Metabolic incentives for dishonest signals of strength in the fiddler crab Uca vomeris.

PubMed

Bywater, Candice L; White, Craig R; Wilson, Robbie S

2014-08-15

To reduce the potential costs of combat, animals may rely upon signals to resolve territorial disputes. Signals also provide a means for individuals to appear better than they actually are, deceiving opponents and gaining access to resources that would otherwise be unattainable. However, other than resource gains, incentives for dishonest signalling remain unexplored. In this study, we tested the idea that unreliable signallers pay lower metabolic costs for their signals, and that energetic savings could represent an incentive for cheating. We focused on two-toned fiddler crabs (Uca vomeris), a species that frequently uses its enlarged claws as signals of dominance to opponents. Previously, we found that regenerated U. vomeris claws are often large but weak (i.e. unreliable). Here, we found that the original claws of male U. vomeris consumed 43% more oxygen than weaker, regenerated claws, suggesting that muscle quantity drives variation in metabolic costs. Therefore, it seems that metabolic savings could provide a powerful incentive for dishonesty within fiddler crabs. © 2014. Published by The Company of Biologists Ltd.

Astrocytes regulate heterogeneity of presynaptic strengths in hippocampal networks

PubMed Central

Letellier, Mathieu; Park, Yun Kyung; Chater, Thomas E.; Chipman, Peter H.; Gautam, Sunita Ghimire; Oshima-Takago, Tomoko; Goda, Yukiko

2016-01-01

Dendrites are neuronal structures specialized for receiving and processing information through their many synaptic inputs. How input strengths are modified across dendrites in ways that are crucial for synaptic integration and plasticity remains unclear. We examined in single hippocampal neurons the mechanism of heterosynaptic interactions and the heterogeneity of synaptic strengths of pyramidal cell inputs. Heterosynaptic presynaptic plasticity that counterbalances input strengths requires N-methyl-d-aspartate receptors (NMDARs) and astrocytes. Importantly, this mechanism is shared with the mechanism for maintaining highly heterogeneous basal presynaptic strengths, which requires astrocyte Ca2+ signaling involving NMDAR activation, astrocyte membrane depolarization, and L-type Ca2+ channels. Intracellular infusion of NMDARs or Ca2+-channel blockers into astrocytes, conditionally ablating the GluN1 NMDAR subunit, or optogenetically hyperpolarizing astrocytes with archaerhodopsin promotes homogenization of convergent presynaptic inputs. Our findings support the presence of an astrocyte-dependent cellular mechanism that enhances the heterogeneity of presynaptic strengths of convergent connections, which may help boost the computational power of dendrites. PMID:27118849

Subsurface event detection and classification using Wireless Signal Networks.

PubMed

Yoon, Suk-Un; Ghazanfari, Ehsan; Cheng, Liang; Pamukcu, Sibel; Suleiman, Muhannad T

2012-11-05

Subsurface environment sensing and monitoring applications such as detection of water intrusion or a landslide, which could significantly change the physical properties of the host soil, can be accomplished using a novel concept, Wireless Signal Networks (WSiNs). The wireless signal networks take advantage of the variations of radio signal strength on the distributed underground sensor nodes of WSiNs to monitor and characterize the sensed area. To characterize subsurface environments for event detection and classification, this paper provides a detailed list and experimental data of soil properties on how radio propagation is affected by soil properties in subsurface communication environments. Experiments demonstrated that calibrated wireless signal strength variations can be used as indicators to sense changes in the subsurface environment. The concept of WSiNs for the subsurface event detection is evaluated with applications such as detection of water intrusion, relative density change, and relative motion using actual underground sensor nodes. To classify geo-events using the measured signal strength as a main indicator of geo-events, we propose a window-based minimum distance classifier based on Bayesian decision theory. The window-based classifier for wireless signal networks has two steps: event detection and event classification. With the event detection, the window-based classifier classifies geo-events on the event occurring regions that are called a classification window. The proposed window-based classification method is evaluated with a water leakage experiment in which the data has been measured in laboratory experiments. In these experiments, the proposed detection and classification method based on wireless signal network can detect and classify subsurface events.

Multisite Phosphorylation Modulates the T Cell Receptor ζ-Chain Potency but not the Switchlike Response.

PubMed

Mukhopadhyay, Himadri; de Wet, Ben; Clemens, Lara; Maini, Philip K; Allard, Jun; van der Merwe, P Anton; Dushek, Omer

2016-04-26

Multisite phosphorylation is ubiquitous in cellular signaling and is thought to provide signaling proteins with additional regulatory mechanisms. Indeed, mathematical models have revealed a large number of mechanisms by which multisite phosphorylation can produce switchlike responses. The T cell antigen receptor (TCR) is a multisubunit receptor on the surface of T cells that is a prototypical multisite substrate as it contains 20 sites that are distributed on 10 conserved immunoreceptor tyrosine-based activation motifs (ITAMs). The TCR ζ-chain is a homodimer subunit that contains six ITAMs (12 sites) and exhibits a number of properties that are predicted to be sufficient for a switchlike response. We have used cellular reconstitution to systematically study multisite phosphorylation of the TCR ζ-chain. We find that multisite phosphorylation proceeds by a nonsequential random mechanism, and find no evidence that multiple ITAMs modulate a switchlike response but do find that they alter receptor potency and maximum phosphorylation. Modulation of receptor potency can be explained by a reduction in molecular entropy of the disordered ζ-chain upon phosphorylation. We further find that the tyrosine kinase ZAP-70 increases receptor potency but does not modulate the switchlike response. In contrast to other multisite proteins, where phosphorylations act in strong concert to modulate protein function, we suggest that the multiple ITAMs on the TCR function mainly to amplify subsequent signaling. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

A Phosphosite within the SH2 Domain of Lck Regulates Its Activation by CD45.

PubMed

Courtney, Adam H; Amacher, Jeanine F; Kadlecek, Theresa A; Mollenauer, Marianne N; Au-Yeung, Byron B; Kuriyan, John; Weiss, Arthur

2017-08-03

The Src Family kinase Lck sets a critical threshold for T cell activation because it phosphorylates the TCR complex and the Zap70 kinase. How a T cell controls the abundance of active Lck molecules remains poorly understood. We have identified an unappreciated role for a phosphosite, Y192, within the Lck SH2 domain that profoundly affects the amount of active Lck in cells. Notably, mutation of Y192 blocks critical TCR-proximal signaling events and impairs thymocyte development in retrogenic mice. We determined that these defects are caused by hyperphosphorylation of the inhibitory C-terminal tail of Lck. Our findings reveal that modification of Y192 inhibits the ability of CD45 to associate with Lck in cells and dephosphorylate the C-terminal tail of Lck, which prevents its adoption of an active open conformation. These results suggest a negative feedback loop that responds to signaling events that tune active Lck amounts and TCR sensitivity. Copyright © 2017 Elsevier Inc. All rights reserved.

ERK-dependent T cell receptor threshold calibration in rheumatoid arthritis.

PubMed

Singh, Karnail; Deshpande, Pratima; Pryshchep, Sergey; Colmegna, Inés; Liarski, Vladimir; Weyand, Cornelia M; Goronzy, Jörg J

2009-12-15

Immune responses to citrullinated neoantigens and clinical efficacy of costimulation blockade indicate a general defect in maintaining T cell tolerance in rheumatoid arthritis (RA). To examine whether TCR threshold calibration contributes to disease pathogenesis, signaling in RA T cells was quantified. RA patients had a selective increase in ERK phosphorylation compared with demographically matched controls due to a mechanism distal of Ras activation. Increased ERK responses included naive and memory CD4 and CD8 T cells and did not correlate with disease activity. The augmented ERK activity delayed SHP-1 recruitment to the TCR synapse and sustained TCR-induced Zap70 and NF-kappaB signaling, facilitating responses to suboptimal stimulation. Increased responsiveness of the ERK pathway was also a characteristic finding in the SKG mouse model of RA where it preceded clinical symptoms. Treatment with subtherapeutic doses of a MEK-1/2 inhibitor delayed arthritis onset and reduced severity, suggesting that increased ERK phosphorylation predisposes for autoimmunity and can be targeted to prevent disease.

Natural variants of cytotoxic epitopes are T-cell receptor antagonists for antiviral cytotoxic T cells

NASA Astrophysics Data System (ADS)

Bertoletti, Antonio; Sette, Alessandro; Chisari, Francis V.; Penna, Amalia; Levrero, Massimo; Carli, Marco De; Fiaccadori, Franco; Ferrari, Carlo

1994-06-01

IT has been suggested that mutations within immunodominant cytotoxic T-lymphocyte (CTL) epitopes may be exploited by viruses to evade protective immune responses critical for clearance1-4. Viral escape could originate from passive mechanisms, such as mutations within crucial CTL epitopes, either affecting major histocompatibility complex binding or T-cell antigen receptor (TCR) recognition. Additionally, it has recently been shown that substitutions of TCR contact sites can yield analogue peptides that can still interact with the T-cell receptor but be unable to deliver a full stimulatory signal, thus inducing anergy5 or acting as an antagonist for the TCR6-8. We report here that hepatitis B virus isolates derived from two chronically infected patients display variant epitopes that act as natural TCR antagonists with the capacity to inhibit the CTL response to the wild-type epitope. During natural infection, TCR antagonist mutations of CTL epitopes could contribute to the development of viral persistence, especially if the antiviral CTL response is monospecific or the epitope is strongly immunodominant.

Signaling aggression.

PubMed

van Staaden, Moira J; Searcy, William A; Hanlon, Roger T

2011-01-01

From psychological and sociological standpoints, aggression is regarded as intentional behavior aimed at inflicting pain and manifested by hostility and attacking behaviors. In contrast, biologists define aggression as behavior associated with attack or escalation toward attack, omitting any stipulation about intentions and goals. Certain animal signals are strongly associated with escalation toward attack and have the same function as physical attack in intimidating opponents and winning contests, and ethologists therefore consider them an integral part of aggressive behavior. Aggressive signals have been molded by evolution to make them ever more effective in mediating interactions between the contestants. Early theoretical analyses of aggressive signaling suggested that signals could never be honest about fighting ability or aggressive intentions because weak individuals would exaggerate such signals whenever they were effective in influencing the behavior of opponents. More recent game theory models, however, demonstrate that given the right costs and constraints, aggressive signals are both reliable about strength and intentions and effective in influencing contest outcomes. Here, we review the role of signaling in lieu of physical violence, considering threat displays from an ethological perspective as an adaptive outcome of evolutionary selection pressures. Fighting prowess is conveyed by performance signals whose production is constrained by physical ability and thus limited to just some individuals, whereas aggressive intent is encoded in strategic signals that all signalers are able to produce. We illustrate recent advances in the study of aggressive signaling with case studies of charismatic taxa that employ a range of sensory modalities, viz. visual and chemical signaling in cephalopod behavior, and indicators of aggressive intent in the territorial calls of songbirds. Copyright © 2011 Elsevier Inc. All rights reserved.

Strength-based criterion shifts in recognition memory.

PubMed

Singer, Murray

2009-10-01

In manipulations of stimulus strength between lists, a more lenient signal detection criterion is more frequently applied to a weak than to a strong stimulus class. However, with randomly intermixed weak and strong test probes, such a criterion shift often does not result. A procedure that has yielded delay-based within-list criterion shifts was applied to strength manipulations in recognition memory for categorized word lists. When participants made semantic ratings about each stimulus word, strength-based criterion shifts emerged regardless of whether words from pairs of categories were studied in separate blocks (Experiment 1) or in intermixed blocks (Experiment 2). In Experiment 3, the criterion shift persisted under the semantic-rating study task, but not under rote memorization. These findings suggest that continually adjusting the recognition decision criterion is cognitively feasible. They provide a technique for manipulating the criterion shift, and they identify competing theoretical accounts of these effects.

Evaluation of rotator cuff muscle strength in healthy individuals

PubMed Central

Cortez, Paulo José Oliveira; Tomazini, José Elias

2015-01-01

OBJECTIVE: To compare the strength generated by the rotator muscles of the shoulder joint between the right upper limb and left upper limb among healthy individuals. METHODS: To evaluate the muscle strength of upper limbs from isometric contractions in the horizontal direction (rotation) an isometric dynamometer was used, equipped with transducers, signal conditioning, a data acquisition board, and finally, a computer. Study participants were 22 male military subjects, aged between 18 and 19 years old, body mass between 57.7 and 93.0 kg (71.8 ± 9.45 kg) and height between 1.67 and 1.90 m (1.75 ± 0.06 m), healthy and without clinical diseases or any type of orthopedic injury in the muscle skeletal system. RESULTS: The internal rotation in the right upper limb (RUL) was higher than the average strength of internal rotation in the left upper limb (LUL) (p = 0.723). The external rotation strength in RUL was lower than the average strength of external rotation in the LUL (p=0.788). No statistical difference was observed by comparing the strength values of all isometric strength tests. CONCLUSION: For the sample and methodology used to assess muscle strength, there was no statistical difference between the strength generated by the muscles of the rotator cuff of the right and left upper limbs. Experimental Study. PMID:26207091

Equations for Estimating the Strength of TV Signals Scattered by Wind Turbines

NASA Technical Reports Server (NTRS)

Spera, David A.; Sengupta, Dipak L.

1994-01-01

During the late 1970's and early 1980's, concerns about the potential interference of wind turbine generators with electromagnetic communication signals led to a series of research studies, both in the laboratory and in the field, conducted by the staff of the University of Michigan Radiation Laboratory. These studies were sponsored by organizations such as the U.S. Department of Energy, the Solar Energy Research Institute, and private developers of wind power stations. Research objectives were to identify the mechanisms by which wind turbines might adversely affect communication signals, estimate the severity of these effects for different types of signals (e.g. television, radio, microwave, and navigation), and formulate mathematical models with which to predict the sizes of potential interference zones around wind turbines and wind power plants. This work formed the basis for preliminary standards on assessing electromagnetic interference (EMI) by wind turbines. With the current renewal of interest in wind energy projects, it is appropriate that the many experimental and analytical aspects of this pioneering work be reviewed and correlated. The purpose of this study is to combine test data and theory from previously published and unpublished research reports into a unified and consistent set of equations which are useful for estimating potential levels of television interference from wind turbines. To be comprehensive, these equations will include both horizontal-axis and vertical-axis wind turbines (HAWT's and VAWT's), blade configuration parameters (e.g. number, size, material, twist, and coning), signal frequency and power, and directional characteristics of the receiving antenna. The approach that is followed in this report is as follows. First, some basic equations that describe electromagnetic signals with interference are presented without detailed derivations, since the latter are available in the references. Minor changes in terminology are made for

WAVE2 Regulates High-Affinity Integrin Binding by Recruiting Vinculin and Talin to the Immunological Synapse▿

PubMed Central

Nolz, Jeffrey C.; Medeiros, Ricardo B.; Mitchell, Jason S.; Zhu, Peimin; Freedman, Bruce D.; Shimizu, Yoji; Billadeau, Daniel D.

2007-01-01

T-cell-receptor (TCR)-mediated integrin activation is required for T-cell-antigen-presenting cell conjugation and adhesion to extracellular matrix components. While it has been demonstrated that the actin cytoskeleton and its regulators play an essential role in this process, no mechanism has been established which directly links TCR-induced actin polymerization to the activation of integrins. Here, we demonstrate that TCR stimulation results in WAVE2-ARP2/3-dependent F-actin nucleation and the formation of a complex containing WAVE2, ARP2/3, vinculin, and talin. The verprolin-connecting-acidic (VCA) domain of WAVE2 mediates the formation of the ARP2/3-vinculin-talin signaling complex and talin recruitment to the immunological synapse (IS). Interestingly, although vinculin is not required for F-actin or integrin accumulation at the IS, it is required for the recruitment of talin. In addition, RNA interference of either WAVE2 or vinculin inhibits activation-dependent induction of high-affinity integrin binding to VCAM-1. Overall, these findings demonstrate a mechanism in which signals from the TCR produce WAVE2-ARP2/3-mediated de novo actin polymerization, leading to integrin clustering and high-affinity binding through the recruitment of vinculin and talin. PMID:17591693

WAVE2 regulates high-affinity integrin binding by recruiting vinculin and talin to the immunological synapse.

PubMed

Nolz, Jeffrey C; Medeiros, Ricardo B; Mitchell, Jason S; Zhu, Peimin; Freedman, Bruce D; Shimizu, Yoji; Billadeau, Daniel D

2007-09-01

T-cell-receptor (TCR)-mediated integrin activation is required for T-cell-antigen-presenting cell conjugation and adhesion to extracellular matrix components. While it has been demonstrated that the actin cytoskeleton and its regulators play an essential role in this process, no mechanism has been established which directly links TCR-induced actin polymerization to the activation of integrins. Here, we demonstrate that TCR stimulation results in WAVE2-ARP2/3-dependent F-actin nucleation and the formation of a complex containing WAVE2, ARP2/3, vinculin, and talin. The verprolin-connecting-acidic (VCA) domain of WAVE2 mediates the formation of the ARP2/3-vinculin-talin signaling complex and talin recruitment to the immunological synapse (IS). Interestingly, although vinculin is not required for F-actin or integrin accumulation at the IS, it is required for the recruitment of talin. In addition, RNA interference of either WAVE2 or vinculin inhibits activation-dependent induction of high-affinity integrin binding to VCAM-1. Overall, these findings demonstrate a mechanism in which signals from the TCR produce WAVE2-ARP2/3-mediated de novo actin polymerization, leading to integrin clustering and high-affinity binding through the recruitment of vinculin and talin.

Force Dynamics During T Cell Activation

NASA Astrophysics Data System (ADS)

Garcia, David A.; Upadhyaya, Arpita

T cell activation is an essential step in the adaptive immune response. The binding of the T cell receptor (TCR) with antigen triggers signaling cascades and cell spreading. Physical forces exerted on the TCR by the cytoskeleton have been shown to induce signaling events. While cellular forces are known to depend on the mechanical properties of the cytoskeleton, the biophysical mechanisms underlying force induced activation of TCR-antigen interactions unknown. Here, we use traction force microscopy to measure the force dynamics of activated Jurkat T cells. The movements of beads embedded in an elastic gel serve as a non-invasive reporter of cytoskeletal and molecular motor dynamics. We examined the statistical structure of the force profiles throughout the cell during signaling activation. We found two spatially distinct active regimes of force generation characterized by different time scales. Typically, the interior of the cells was found to be more active than the periphery. Inhibition of myosin motor activity altered the correlation time of the bead displacements indicating additional sources of stochastic force generation. Our results indicate a complex interaction between myosin activity and actin polymerization dynamics in producing cellular forces in immune cells.

Functional MRI of Auditory Responses in the Zebra Finch Forebrain Reveals a Hierarchical Organisation Based on Signal Strength but Not Selectivity

PubMed Central

Boumans, Tiny; Gobes, Sharon M. H.; Poirier, Colline; Theunissen, Frederic E.; Vandersmissen, Liesbeth; Pintjens, Wouter; Verhoye, Marleen; Bolhuis, Johan J.; Van der Linden, Annemie

2008-01-01

Background Male songbirds learn their songs from an adult tutor when they are young. A network of brain nuclei known as the ‘song system’ is the likely neural substrate for sensorimotor learning and production of song, but the neural networks involved in processing the auditory feedback signals necessary for song learning and maintenance remain unknown. Determining which regions show preferential responsiveness to the bird's own song (BOS) is of great importance because neurons sensitive to self-generated vocalisations could mediate this auditory feedback process. Neurons in the song nuclei and in a secondary auditory area, the caudal medial mesopallium (CMM), show selective responses to the BOS. The aim of the present study is to investigate the emergence of BOS selectivity within the network of primary auditory sub-regions in the avian pallium. Methods and Findings Using blood oxygen level-dependent (BOLD) fMRI, we investigated neural responsiveness to natural and manipulated self-generated vocalisations and compared the selectivity for BOS and conspecific song in different sub-regions of the thalamo-recipient area Field L. Zebra finch males were exposed to conspecific song, BOS and to synthetic variations on BOS that differed in spectro-temporal and/or modulation phase structure. We found significant differences in the strength of BOLD responses between regions L2a, L2b and CMM, but no inter-stimuli differences within regions. In particular, we have shown that the overall signal strength to song and synthetic variations thereof was different within two sub-regions of Field L2: zone L2a was significantly more activated compared to the adjacent sub-region L2b. Conclusions Based on our results we suggest that unlike nuclei in the song system, sub-regions in the primary auditory pallium do not show selectivity for the BOS, but appear to show different levels of activity with exposure to any sound according to their place in the auditory processing stream. PMID

Phosphorylation of the Grb2- and phosphatidylinositol 3-kinase p85-binding p36/38 by Syk in Lck-negative T cells.

PubMed

von Willebrand, M; Williams, S; Tailor, P; Mustelin, T

1998-06-01

Activation of the mitogen-activated protein kinase (MAPK) pathway by the T-cell antigen receptor (TCR) in T cells involves a positive role for phosphatidylinositol 3-kinase (PI3K) activity. We recently reported that over-expression of the Syk protein tyrosine kinase in the Lck-negative JCaM1 cells enabled the TCR to induce a normal activation of the Erk2 MAPK and enhanced transcription of a reporter gene driven by the nuclear factor of activated T cells and AP-1. Because this system allows us to analyse the targets for Syk in receptor-mediated signalling, we examined the role of PI3K in signalling events between the TCR-regulated Syk and the downstream activation of Erk2. We report that inhibition of PI3K by wortmannin or an inhibitory p85 construct, p85deltaiSH2, reduced the TCR-induced Syk-dependent activation of Erk2, as well as the appearance of phospho-Erk and phospho-Mek. At the same time, expression of Syk resulted in the activation-dependent phosphorylation of three proteins that bound to the src homology 2 (SH2) domains of PI3K p85. The strongest of these bands had an apparent molecular mass of 36-38 kDa on SDS gels, and it was quantitatively removed from the lysates by adsorption to a fusion protein containing the SH2 domain of Grb2. The appearance of this band was Syk dependent, and it was seen only upon triggering of the TCR complex. Thus, p36/38 was phosphorylated by Syk or a Syk-regulated kinase, and this protein may provide a link to the recruitment and activation of PI3K, as well as to the Ras-MAPK pathway, in TCR-triggered T cells.

Differential Responsiveness of Innate-like IL-17- and IFN-γ-Producing γδ T Cells to Homeostatic Cytokines.

PubMed

Corpuz, Theresa M; Stolp, Jessica; Kim, Hee-Ok; Pinget, Gabriela V; Gray, Daniel H D; Cho, Jae-Ho; Sprent, Jonathan; Webster, Kylie E

2016-01-15

γδ T cells respond to molecules upregulated following infection or cellular stress using both TCR and non-TCR molecules. The importance of innate signals versus TCR ligation varies greatly. Both innate-like IL-17-producing γδ T (γδT-17) and IFN-γ-producing γδ T (γδT-IFNγ) subsets tune the sensitivity of their TCR following thymic development, allowing robust responses to inflammatory cytokines in the periphery. The remaining conventional γδ T cells retain high TCR responsiveness. We determined homeostatic mechanisms that govern these various subsets in the peripheral lymphoid tissues. We found that, although innate-like γδT-17 and γδT-IFNγ cells share elements of thymic development, they diverge when it comes to homeostasis. Both exhibit acute sensitivity to cytokines compared with conventional γδ T cells, but they do not monopolize the same cytokine. γδT-17 cells rely exclusively on IL-7 for turnover and survival, aligning them with NKT17 cells; IL-7 ligation triggers proliferation, as well as promotes survival, upregulating Bcl-2 and Bcl-xL. γδT-IFNγ cells instead depend heavily on IL-15. They display traits analogous to memory CD8(+) T cells and upregulate Bcl-xL and Mcl-1 upon cytokine stimulation. The conventional γδ T cells display low sensitivity to cytokine-alone stimulation and favor IL-7 for their turnover, characteristics reminiscent of naive αβ T cells, suggesting that they may also require tonic TCR signaling for population maintenance. These survival constraints suggest that γδ T cell subsets do not directly compete with each other for cytokines, but instead fall into resource niches with other functionally similar lymphocytes. Copyright © 2016 by The American Association of Immunologists, Inc.

Subsurface Event Detection and Classification Using Wireless Signal Networks

PubMed Central

Yoon, Suk-Un; Ghazanfari, Ehsan; Cheng, Liang; Pamukcu, Sibel; Suleiman, Muhannad T.

2012-01-01

Subsurface environment sensing and monitoring applications such as detection of water intrusion or a landslide, which could significantly change the physical properties of the host soil, can be accomplished using a novel concept, Wireless Signal Networks (WSiNs). The wireless signal networks take advantage of the variations of radio signal strength on the distributed underground sensor nodes of WSiNs to monitor and characterize the sensed area. To characterize subsurface environments for event detection and classification, this paper provides a detailed list and experimental data of soil properties on how radio propagation is affected by soil properties in subsurface communication environments. Experiments demonstrated that calibrated wireless signal strength variations can be used as indicators to sense changes in the subsurface environment. The concept of WSiNs for the subsurface event detection is evaluated with applications such as detection of water intrusion, relative density change, and relative motion using actual underground sensor nodes. To classify geo-events using the measured signal strength as a main indicator of geo-events, we propose a window-based minimum distance classifier based on Bayesian decision theory. The window-based classifier for wireless signal networks has two steps: event detection and event classification. With the event detection, the window-based classifier classifies geo-events on the event occurring regions that are called a classification window. The proposed window-based classification method is evaluated with a water leakage experiment in which the data has been measured in laboratory experiments. In these experiments, the proposed detection and classification method based on wireless signal network can detect and classify subsurface events. PMID:23202191

Gating-signal propagation by a feed-forward neural motif

NASA Astrophysics Data System (ADS)

Liang, Xiaoming; Yanchuk, Serhiy; Zhao, Liang

2013-07-01

We study the signal propagation in a feed-forward motif consisting of three bistable neurons: Two input neurons receive input signals and the third output neuron generates the output. We find that a weak input signal can be propagated from the input neurons to the output neuron without amplitude attenuation. We further reveal that the initial states of the input neurons and the coupling strength act as signal gates and determine whether the propagation is enhanced or not. We also investigate the effect of the input signal frequency on enhanced signal propagation.

Research on signal demodulation technology of Mach-Zehnder optical fiber sensor vibration system

NASA Astrophysics Data System (ADS)

Liu, Juncheng; Cheng, Pengshen; Hu, Tong

2017-08-01

Mach-Zehnder (M-Z) interferometer is frequently used in optical fiber vibration system. And signal demodulation technology plays an important role in the signal processing of M-Z optical fiber vibration system. In order to accurately get the phase information of the vibration signals, the signal demodulation technique based on M-Z interference principle is studied. In this paper, by analyzing the principles of 3 × 3 fiber coupler homodyne demodulation method and phase-generating carrier (PGC) technology, the advantages and disadvantages of the two demodulation methods for different vibration signal are presented. Then the method of judging signal strength is proposed. The correlation between the demodulation effects and strength of the perturbation signals is analyzed. Finally, the simulation experiments are carried out to compare the demodulation effects of the two demodulation methods, the results demonstrate that PGC demodulation technology has great advantages in weak signals, and the 3 × 3 fiber coupler is more suitable for strong signals.

Cooperation in the dark: signalling and collective action in quorum-sensing bacteria.

PubMed

Brown, S P; Johnstone, R A

2001-05-07

The study of quorum-sensing bacteria has revealed a widespread mechanism of coordinating bacterial gene expression with cell density. By monitoring a constitutively produced signal molecule, individual bacteria can limit their expression of group-beneficial phenotypes to cell densities that guarantee an effective group outcome. In this paper, we attempt to move away from a commonly expressed view that these impressive feats of coordination are examples of multicellularity in prokaryotic populations. Here, we look more closely at the individual conflict underlying this cooperation, illustrating that, even under significant levels of genetic conflict, signalling and resultant cooperative behaviour can stably exist. A predictive two-trait model of signal strength and of the extent of cooperation is developed as a function of relatedness (reflecting multiplicity of infection) and basic population demographic parameters. The model predicts that the strength of quorum signalling will increase as conflict (multiplicity of infecting strains) increases, as individuals attempt to coax more cooperative contributions from their competitors, leading to a devaluation of the signal as an indicator of density. Conversely, as genetic conflict increases, the model predicts that the threshold density for cooperation will increase and the subsequent strength of group cooperation will be depressed.

Retrieval Search and Strength Evoke Dissociable Brain Activity during Episodic Memory Recall

PubMed Central

Reas, Emilie T.; Brewer, James B.

2014-01-01

Neuroimaging studies of episodic memory retrieval have revealed activations in the human frontal, parietal, and medial-temporal lobes that are associated with memory strength. However, it remains unclear whether these brain responses are veritable signals of memory strength or are instead regulated by concomitant subcomponents of retrieval such as retrieval effort or mental search. This study used event-related fMRI during cued recall of previously memorized word-pair associates to dissociate brain responses modulated by memory search from those modulated by the strength of a recalled memory. Search-related deactivations, dissociated from activity due to memory strength, were observed in regions of the default network, whereas distinctly strength-dependent activations were present in superior and inferior parietal and dorsolateral PFC. Both search and strength regulated activity in dorsal anterior cingulate and anterior insula. These findings suggest that, although highly correlated and partially subserved by overlapping cognitive control mechanisms, search and memory strength engage dissociable regions of frontoparietal attention and default networks. PMID:23190328

A generalizable platform for interrogating target- and signal-specific consequences of electrophilic modifications in redox-dependent cell signaling.

PubMed

Lin, Hong-Yu; Haegele, Joseph A; Disare, Michael T; Lin, Qishan; Aye, Yimon

2015-05-20

Despite the known propensity of small-molecule electrophiles to react with numerous cysteine-active proteins, biological actions of individual signal inducers have emerged to be chemotype-specific. To pinpoint and quantify the impacts of modifying one target out of the whole proteome, we develop a target-protein-personalized "electrophile toolbox" with which specific intracellular targets can be selectively modified at a precise time by specific reactive signals. This general methodology, T-REX (targetable reactive electrophiles and oxidants), is established by (1) constructing a platform that can deliver a range of electronic and sterically different bioactive lipid-derived signaling electrophiles to specific proteins in cells; (2) probing the kinetics of targeted delivery concept, which revealed that targeting efficiency in cells is largely driven by initial on-rate of alkylation; and (3) evaluating the consequences of protein-target- and small-molecule-signal-specific modifications on the strength of downstream signaling. These data show that T-REX allows quantitative interrogations into the extent to which the Nrf2 transcription factor-dependent antioxidant response element (ARE) signaling is activated by selective electrophilic modifications on Keap1 protein, one of several redox-sensitive regulators of the Nrf2-ARE axis. The results document Keap1 as a promiscuous electrophile-responsive sensor able to respond with similar efficiencies to discrete electrophilic signals, promoting comparable strength of Nrf2-ARE induction. T-REX is also able to elicit cell activation in cases in which whole-cell electrophile flooding fails to stimulate ARE induction prior to causing cytotoxicity. The platform presents a previously unavailable opportunity to elucidate the functional consequences of small-molecule-signal- and protein-target-specific electrophilic modifications in an otherwise unaffected cellular background.

Phosphoproteomics Reveals Regulatory T Cell-Mediated DEF6 Dephosphorylation That Affects Cytokine Expression in Human Conventional T Cells

PubMed Central

Joshi, Rubin N.; Binai, Nadine A.; Marabita, Francesco; Sui, Zhenhua; Altman, Amnon; Heck, Albert J. R.; Tegnér, Jesper; Schmidt, Angelika

2017-01-01

Regulatory T cells (Tregs) control key events of immune tolerance, primarily by suppression of effector T cells. We previously revealed that Tregs rapidly suppress T cell receptor (TCR)-induced calcium store depletion in conventional CD4+CD25− T cells (Tcons) independently of IP3 levels, consequently inhibiting NFAT signaling and effector cytokine expression. Here, we study Treg suppression mechanisms through unbiased phosphoproteomics of primary human Tcons upon TCR stimulation and Treg-mediated suppression, respectively. Tregs induced a state of overall decreased phosphorylation as opposed to TCR stimulation. We discovered novel phosphosites (T595_S597) in the DEF6 (SLAT) protein that were phosphorylated upon TCR stimulation and conversely dephosphorylated upon coculture with Tregs. Mutation of these DEF6 phosphosites abrogated interaction of DEF6 with the IP3 receptor and affected NFAT activation and cytokine transcription in primary Tcons. This novel mechanism and phosphoproteomics data resource may aid in modifying sensitivity of Tcons to Treg-mediated suppression in autoimmune disease or cancer. PMID:28993769

Impact of lipid rafts on the T -cell-receptor and peptide-major-histocompatibility-complex interactions under different measurement conditions

NASA Astrophysics Data System (ADS)

Li, Long; Xu, Guang-Kui; Song, Fan

2017-01-01

The interactions between T-cell receptor (TCR) and peptide-major-histocompatibility complex (pMHC), which enable T-cell development and initiate adaptive immune responses, have been intensively studied. However, a central issue of how lipid rafts affect the TCR-pMHC interactions remains unclear. Here, by using a statistical-mechanical membrane model, we show that the binding affinity of TCR and pMHC anchored on two apposing cell membranes is significantly enhanced because of the lipid raft-induced signaling protein aggregation. This finding may provide an alternative insight into the mechanism of T-cell activation triggered by very low densities of pMHC. In the case of cell-substrate adhesion, our results indicate that the loss of lateral mobility of the proteins on the solid substrate leads to the inhibitory effect of lipid rafts on TCR-pMHC interactions. Our findings help to understand why different experimental methods for measuring the impact of lipid rafts on the receptor-ligand interactions have led to contradictory conclusions.

Disregulated expression of the transcription factor ThPOK during T-cell development leads to high incidence of T-cell lymphomas.

PubMed

Lee, Hyung-Ok; He, Xiao; Mookerjee-Basu, Jayati; Zhongping, Dai; Hua, Xiang; Nicolas, Emmanuelle; Sulis, Maria Luisa; Ferrando, Adolfo A; Testa, Joseph R; Kappes, Dietmar J

2015-06-23

The transcription factor T-helper-inducing POZ/Krueppel-like factor (ThPOK, encoded by the Zbtb7b gene) plays widespread and critical roles in T-cell development, particularly as the master regulator of CD4 commitment. Here we show that mice expressing a constitutive T-cell-specific ThPOK transgene (ThPOK(const) mice) develop thymic lymphomas. These tumors resemble human T-cell acute lymphoblastic leukemia (T-ALL), in that they predominantly exhibit activating Notch1 mutations. Lymphomagenesis is prevented if thymocyte development is arrested at the DN3 stage by recombination-activating gene (RAG) deficiency, but restored by introduction of a T-cell receptor (TCR) transgene or by a single injection of anti-αβTCR antibody into ThPOK(const) RAG-deficient mice, which promotes development to the CD4(+)8(+) (DP) stage. Hence, TCR signals and/or traversal of the DN (double negative) > DP (double positive) checkpoint are required for ThPOK-mediated lymphomagenesis. These results demonstrate a novel link between ThPOK, TCR signaling, and lymphomagenesis. Finally, we present evidence that ectopic ThPOK expression gives rise to a preleukemic and self-perpetuating DN4 lymphoma precursor population. Our results collectively define a novel role for ThPOK as an oncogene and precisely map the stage in thymopoiesis susceptible to ThPOK-dependent tumor initiation.

Targeted Sos1 deletion reveals its critical role in early T-cell development

PubMed Central

Kortum, Robert L.; Sommers, Connie L.; Alexander, Clayton P.; Pinski, John M.; Li, Wenmei; Grinberg, Alex; Lee, Jan; Love, Paul E.; Samelson, Lawrence E.

2011-01-01

Activation of the small G protein Ras is required for thymocyte differentiation. In thymocytes, Ras is activated by the Ras guanine exchange factors (RasGEFs) Sos1, Sos2, and RasGRP1. We report the development of a floxed allele of sos1 to assess the role of Sos1 during thymocyte development. Sos1 was required for pre–T-cell receptor (pre-TCR)– but not TCR-stimulated developmental signals. Sos1 deletion led to a partial block at the DN-to-DP transition. Sos1-deficient thymocytes showed reduced pre-TCR–stimulated proliferation, differentiation, and ERK phosphorylation. In contrast, TCR-stimulated positive selection, and negative selection under strong stimulatory conditions, remained intact in Sos1-deficient mice. Comparison of RasGEF expression at different developmental stages showed that relative to Sos2 and RasGRP1, Sos1 is most abundant in DN thymocytes, but least abundant in DP thymocytes. These data reveal that Sos1 is uniquely positioned to affect signal transduction early in thymocyte development. PMID:21746917

On the Contribution of Curl-Free Current Patterns to the Ultimate Intrinsic Signal-to-Noise Ratio at Ultra-High Field Strength.

PubMed

Pfrommer, Andreas; Henning, Anke

2017-05-01

The ultimate intrinsic signal-to-noise ratio (SNR) is a coil independent performance measure to compare different receive coil designs. To evaluate this benchmark in a sample, a complete electromagnetic basis set is required. The basis set can be obtained by curl-free and divergence-free surface current distributions, which excite linearly independent solutions to Maxwell's equations. In this work, we quantitatively investigate the contribution of curl-free current patterns to the ultimate intrinsic SNR in a spherical head-sized model at 9.4 T. Therefore, we compare the ultimate intrinsic SNR obtained with having only curl-free or divergence-free current patterns, with the ultimate intrinsic SNR obtained from a combination of curl-free and divergence-free current patterns. The influence of parallel imaging is studied for various acceleration factors. Moreover results for different field strengths (1.5 T up to 11.7 T) are presented at specific voxel positions and acceleration factors. The full-wave electromagnetic problem is analytically solved using dyadic Green's functions. We show, that at ultra-high field strength (B 0 ⩾7T) a combination of curl-free and divergence-free current patterns is required to achieve the best possible SNR at any position in a spherical head-sized model. On 1.5- and 3T platforms, divergence-free current patterns are sufficient to cover more than 90% of the ultimate intrinsic SNR. Copyright © 2017 John Wiley & Sons, Ltd.

Tumor vessel-injuring ability improves antitumor effect of cytotoxic T lymphocytes in adoptive immunotherapy.

PubMed

Kanagawa, N; Yanagawa, T; Nakagawa, T; Okada, N; Nakagawa, S

2013-01-01

Angiogenesis is required for normal physiologic processes, but it is also involved in tumor growth, progression and metastasis. Here, we report the development of an immune-based antiangiogenic strategy based on the generation of T lymphocytes that possess killing specificity for cells expressing vascular endothelial growth factor receptor 2 (VEGFR2). To target VEGFR2-expressing cells, we engineered cytotoxic T lymphocyte (CTL) expressing chimeric T-cell receptors (cTCR-CTL) comprised of a single-chain variable fragment (scFv) against VEGFR2 linked to an intracellular signaling sequence derived from the CD3ζ chain of the TCR and CD28 by retroviral gene transduction methods. The cTCR-CTL exhibited efficient killing specificity against VEGFR2 and a tumor-targeting function in vitro and in vivo. Reflecting such abilities, we confirmed that the cTCR-CTL strongly inhibited the growth of a variety of syngeneic tumors after adoptive transfer into tumor-bearing mice without consequent damage to normal tissue. In addition, CTL expressing both cTCR and tumor-specific TCR induced complete tumor regression due to enhanced tumor infiltration by the CTL and long-term antigen-specific function. These findings provide evidence that the tumor vessel-injuring ability improved the antitumor effect of CTLs in adoptive immunotherapy for a broad range of cancers by inducing immune-mediated destruction of the tumor neovasculature.

Single-chain antigen recognition receptors that costimulate potent rejection of established experimental tumors.

PubMed

Haynes, Nicole M; Trapani, Joseph A; Teng, Michèle W L; Jackson, Jacob T; Cerruti, Loretta; Jane, Stephen M; Kershaw, Michael H; Smyth, Mark J; Darcy, Phillip K

2002-11-01

Tumor cells are usually weakly immunogenic as they largely express self-antigens and can down-regulate major histocompatability complex/peptide molecules and critical costimulatory ligands. The challenge for immunotherapies has been to provide vigorous immune effector cells that circumvent these tumor escape mechanisms and eradicate established tumors. One promising approach is to engineer T cells with single-chain antibody receptors, and since T cells require 2 distinct signals for optimal activation, we have compared the therapeutic efficacy of erbB2-reactive chimeric receptors that contain either T-cell receptor zeta (TCR-zeta) or CD28/TCR-zeta signaling domains. We have demonstrated that primary mouse CD8(+) T lymphocytes expressing the single-chain Fv (scFv)-CD28-zeta receptor have a greater capacity to secrete Tc1 cytokines, induce T-cell proliferation, and inhibit established tumor growth and metastases in vivo. The suppression of established tumor burden by cytotoxic T cells expressing the CD28/TCR-zeta chimera was critically dependent upon their interferon gamma (IFN-gamma) secretion. Our study has illustrated the practical advantage of engineering a T-cell signaling complex that codelivers CD28 activation, dependent only upon the tumor's expression of the appropriate tumor associated antigen.

MUC1 (CD227) interacts with lck tyrosine kinase in Jurkat lymphoma cells and normal T cells.

PubMed

Mukherjee, P; Tinder, T L; Basu, G D; Gendler, S J

2005-01-01

MUC1 (CD227) is a large transmembrane epithelial mucin glycoprotein, which is aberrantly overexpressed in most adenocarcinomas and is a target for immune therapy for epithelial tumors. Recently, MUC1 has been detected in a variety of hematopoietic cell malignancies including T and B cell lymphomas and myelomas; however, its function in these cells is not clearly defined. Using the Jurkat T cell lymphoma cell line and normal human T cells, we demonstrate that MUC1 is not only expressed in these cells but is also phosphorylated upon T cell receptor (TCR) ligation and associates with the Src-related T cell tyrosine kinase, p56lck. Upon TCR-mediated activation of Jurkat cells, MUC1 is found in the low-density membrane fractions, where linker of T cell activation is contained. Abrogation of MUC1 expression in Jurkat cells by MUC1-specific small interfering RNA resulted in defects in TCR-mediated downstream signaling events associated with T cell activation. These include reduction in Ca2+ influx and extracellular signal-regulated kinase 1/2 phosphorylation, leading to a decrease in CD69 expression, proliferation, and interleukin-2 production. These results suggest a regulatory role of MUC1 in modulating proximal signal transduction events through its interaction with proteins of the activation complex.

How nonuniform contact profiles of T cell receptors modulate thymic selection outcomes

NASA Astrophysics Data System (ADS)

Chen, Hanrong; Chakraborty, Arup K.; Kardar, Mehran

2018-03-01

T cell receptors (TCRs) bind foreign or self-peptides attached to major histocompatibility complex (MHC) molecules, and the strength of this interaction determines T cell activation. Optimizing the ability of T cells to recognize a diversity of foreign peptides yet be tolerant of self-peptides is crucial for the adaptive immune system to properly function. This is achieved by selection of T cells in the thymus, where immature T cells expressing unique, stochastically generated TCRs interact with a large number of self-peptide-MHC; if a TCR does not bind strongly enough to any self-peptide-MHC, or too strongly with at least one self-peptide-MHC, the T cell dies. Past theoretical work cast thymic selection as an extreme value problem and characterized the statistical enrichment or depletion of amino acids in the postselection TCR repertoire, showing how T cells are selected to be able to specifically recognize peptides derived from diverse pathogens yet have limited self-reactivity. Here, we investigate how the diversity of the postselection TCR repertoire is modified when TCRs make nonuniform contacts with peptide-MHC. Specifically, we were motivated by recent experiments showing that amino acids at certain positions of a TCR sequence have large effects on thymic selection outcomes, and crystal structure data that reveal a nonuniform contact profile between a TCR and its peptide-MHC ligand. Using a representative TCR contact profile as an illustration, we show via simulations that the statistical enrichment or depletion of amino acids now varies by position according to the contact profile, and, importantly, it depends on the implementation of nonuniform contacts during thymic selection. We explain these nontrivial results analytically. Our study has implications for understanding the selection forces that shape the functionality of the postselection TCR repertoire.

Application of Photoshop and Scion Image analysis to quantification of signals in histochemistry, immunocytochemistry and hybridocytochemistry.

PubMed

Tolivia, Jorge; Navarro, Ana; del Valle, Eva; Perez, Cristina; Ordoñez, Cristina; Martínez, Eva

2006-02-01

To describe a simple method to achieve the differential selection and subsequent quantification of the strength signal using only one section. Several methods for performing quantitative histochemistry, immunocytochemistry or hybridocytochemistry, without use of specific commercial image analysis systems, rely on pixel-counting algorithms, which do not provide information on the amount of chromogen present in the section. Other techniques use complex algorithms to calculate the cumulative signal strength using two consecutive sections. To separate the chromogen signal we used the "Color range" option of the Adobe Photoshop program, which provides a specific file for a particular chromogen selection that could be applied on similar sections. The measurement of the chromogen signal strength of the specific staining is achieved with the Scion Image software program. The method described in this paper can also be applied to simultaneous detection of different signals on the same section or different parameters (area of particles, number of particles, etc.) when the "Analyze particles" tool of the Scion program is used.

Mitochondrial reactive oxygen species regulate the strength of inhibitory GABA-mediated synaptic transmission

NASA Astrophysics Data System (ADS)

Accardi, Michael V.; Daniels, Bryan A.; Brown, Patricia M. G. E.; Fritschy, Jean-Marc; Tyagarajan, Shiva K.; Bowie, Derek

2014-01-01

Neuronal communication imposes a heavy metabolic burden in maintaining ionic gradients essential for action potential firing and synaptic signalling. Although cellular metabolism is known to regulate excitatory neurotransmission, it is still unclear whether the brain’s energy supply affects inhibitory signalling. Here we show that mitochondrial-derived reactive oxygen species (mROS) regulate the strength of postsynaptic GABAA receptors at inhibitory synapses of cerebellar stellate cells. Inhibition is strengthened through a mechanism that selectively recruits α3-containing GABAA receptors into synapses with no discernible effect on resident α1-containing receptors. Since mROS promotes the emergence of postsynaptic events with unique kinetic properties, we conclude that newly recruited α3-containing GABAA receptors are activated by neurotransmitter released onto discrete postsynaptic sites. Although traditionally associated with oxidative stress in neurodegenerative disease, our data identify mROS as a putative homeostatic signalling molecule coupling cellular metabolism to the strength of inhibitory transmission.

A Generalizable Platform for Interrogating Target- and Signal-Specific Consequences of Electrophilic Modifications in Redox-Dependent Cell Signaling

PubMed Central

Lin, Hong-Yu; Haegele, Joseph A.; Disare, Michael T.; Lin, Qishan; Aye, Yimon

2015-01-01

Despite the known propensity of small-molecule electrophiles to react with numerous cysteine-active proteins, biological actions of individual signal inducers have emerged to be chemotype-specific. To pinpoint and quantify the impacts of modifying one target out of the whole proteome, we develop a target-protein-personalized “electrophile toolbox” with which specific intracellular targets can be selectively modified at a precise time by specific reactive signals. This general methodology—T-REX (targetable reactive electrophiles & oxidants)—is established by: (1) constructing a platform that can deliver a range of electronic and sterically different bioactive lipid-derived signaling electrophiles to specific proteins in cells; (2) probing the kinetics of targeted delivery concept which revealed that targeting efficiency in cells is largely driven by initial on-rate of alkylation; and (3) evaluating the consequences of protein-target- and small-molecule-signal-specific modifications on the strength of downstream signaling. These data show that T-REX allows quantitative interrogations into the extent to which the Nrf2 transcription factor-dependent antioxidant response element (ARE) signaling is activated by selective electrophilic modifications on Keap1 protein—one of several redox-sensitive regulators of the Nrf2–ARE axis. The results document Keap1 as a promiscuous electrophile-responsive sensor able to respond with similar efficiencies to discrete electrophilic signals, promoting comparable strength of Nrf2–ARE induction. T-REX is also able to elicit cell activation in cases in which whole-cell electrophile flooding fails to stimulate ARE induction prior to causing cytotoxicity. The platform presents a previously unavailable opportunity to elucidate the functional consequences of small-molecule-signal- and protein-target-specific electrophilic modifications in an otherwise unaffected cellular background. PMID:25909755

Spectral characterization of plastic scintillation detector response as a function of magnetic field strength

NASA Astrophysics Data System (ADS)

Simiele, E.; Kapsch, R.-P.; Ankerhold, U.; Culberson, W.; DeWerd, L.

2018-04-01

The purpose of this work was to characterize intensity and spectral response changes in a plastic scintillation detector (PSD) as a function of magnetic field strength. Spectra measurements as a function of magnetic field strength were performed using an optical spectrometer. The response of both a PSD and PMMA fiber were investigated to isolate the changes in response from the scintillator and the noise signal as a function of magnetic field strength. All irradiations were performed in water at a photon beam energy of 6 MV. Magnetic field strengths of (0, ±0.35, ±0.70, ±1.05, and  ±1.40) T were investigated. Four noise subtraction techniques were investigated to evaluate the impact on the resulting noise-subtracted scintillator response with magnetic field strength. The noise subtraction methods included direct spectral subtraction, the spectral method, and variants thereof. The PMMA fiber exhibited changes in response of up to 50% with magnetic field strength due to the directional light emission from \\breve{C} erenkov radiation. The PSD showed increases in response of up to 10% when not corrected for the noise signal, which agrees with previous investigations of scintillator response in magnetic fields. Decreases in the \\breve{C} erenkov light ratio with negative field strength were observed with a maximum change at  ‑1.40 T of 3.2% compared to 0 T. The change in the noise-subtracted PSD response as a function of magnetic field strength varied with the noise subtraction technique used. Even after noise subtraction, the PSD exhibited changes in response of up to 5.5% over the four noise subtraction methods investigated.

Human common acute lymphoblastic leukemia-derived cell lines are competent to recombine their T-cell receptor delta/alpha regions along a hierarchically ordered pathway.

PubMed

Hansen-Hagge, T E; Yokota, S; Reuter, H J; Schwarz, K; Bartram, C R

1992-11-01

Rearrangements of the T-cell receptor (TCR) delta locus are observed in the majority of human B-cell precursor acute lymphoblastic leukemias (ALL) with a striking predominance of V delta 2(D)D delta 3 recombinations in common ALL (cALL) patients. Recently, we and others showed that almost 20% of cALL cases are characterized by further recombination of V delta 2(D)D delta 3 segments to J alpha elements, thereby deleting the TCR delta locus in analogy to the delta Rec/psi J alpha pathway in differentiating alpha/beta-positive T cells. We report here that two human cALL-derived cell lines, REH and Nalm-6, are competent to recombine the TCR delta/alpha locus under standard tissue culture conditions. Analysis of different REH subclones obtained by limiting dilution of the initial culture showed a biased recombination of V delta 2D delta 3 to distinct J alpha elements. During prolonged tissue culture, a subclone acquired growth advantage and displaced parental cells as well as other subclones. Frequently, the DJ junctions of REH subclones contained extended stretches of palindromic sequences derived from modified D delta 3 coding elements. The other cell line, Nalm-6, started the TCR delta/alpha recombination with an unusual signal joint of a cryptic recombinase signal sequence (RSS) upstream of D delta 3 to the 3' RSS of D delta 3. The RSS dimer was subsequently rearranged in all investigated subclones to an identical J alpha element. Both cell lines might become valuable tools to unravel the complex regulation of TCR delta/alpha recombination pathways in malignant and normal lymphopoiesis.

Subcellular distribution of Lck during CD4 T-cell maturation in the thymic medulla regulates the T-cell activation threshold.

PubMed

Stephen, Tom Li; Wilson, Bridget S; Laufer, Terri M

2012-05-08

Mature peripheral T cells respond to foreign but not to self-antigens. During development in the thymus, deletion of high-affinity self-reactive immature thymocytes contributes to tolerance of mature T cells. However, double-positive thymocytes are positively selected to survive if they respond to self-peptide-MHC complexes; thus, there must be mechanisms to prevent overt reactivity to those same complexes in the periphery. "Developmental tuning" is the active process through which T-cell receptor (TCR)-associated signaling pathways of single-positive (SP) thymocytes are attenuated to respond appropriately to self-peptide-MHC complexes in the periphery. We previously showed that MHC class II expression in the thymic medulla was necessary to tune CD4(+) SP (CD4 SP) thymocytes. CD4 SP thymocytes from mice lacking medullary MHC class II expression had inappropriately enhanced proximal TCR signaling to low-affinity self-ligands that was associated with altered cellular distribution of the tyrosine kinase Lck. Now, we report that activation of both tuned and untuned CD4 SP thymocytes is Lck-dependent. Untuned CD4 SP cells contain a pool of Lck with increased basal phosphorylation that is not associated with the CD4 coreceptor. Phosphorylation of this pool of Lck decreases with tuning. Immunogold transmission electron microscopy of membrane sheets permitted direct visualization of Lck. In the absence of tuning, a significant proportion of Lck and the TCR subunit CD3ζ are expressed on the same protein island; this close association of Lck and the TCR probably explains the enhanced activation of untuned CD4 SP cells. Thus, changes in membrane topography during thymic maturation determine the set point for TCR responsiveness.

SRC homology 2 domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76) N-terminal tyrosine residues regulate a dynamic signaling equilibrium involving feedback of proximal T-cell receptor (TCR) signaling.

PubMed

Ji, Qinqin; Ding, Yiyuan; Salomon, Arthur R

2015-01-01

SRC homology 2 domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76) is a cytosolic adaptor protein that plays an important role in the T-cell receptor-mediated T-cell signaling pathway. SLP-76 links proximal receptor stimulation to downstream effectors through interaction with many signaling proteins. Previous studies showed that mutation of three tyrosine residues, Tyr(112), Tyr(128), and Tyr(145), in the N terminus of SLP-76 results in severely impaired phosphorylation and activation of Itk and PLCγ1, which leads to defective calcium mobilization, Erk activation, and NFAT activation. To expand our knowledge of the role of N-terminal phosphorylation of SLP-76 from these three tyrosine sites, we characterized nearly 1000 tyrosine phosphorylation sites via mass spectrometry in SLP-76 reconstituted wild-type cells and SLP-76 mutant cells in which three tyrosine residues were replaced with phenylalanines (Y3F mutant). Mutation of the three N-terminal tyrosine residues of SLP-76 phenocopied SLP-76-deficient cells for the majority of tyrosine phosphorylation sites observed, including feedback on proximal T-cell receptor signaling proteins. Meanwhile, reversed phosphorylation changes were observed on Tyr(192) of Lck when we compared mutants to the complete removal of SLP-76. In addition, N-terminal tyrosine sites of SLP-76 also perturbed phosphorylation of Tyr(440) of Fyn, Tyr(702) of PLCγ1, Tyr(204), Tyr(397), and Tyr(69) of ZAP-70, revealing new modes of regulation on these sites. All these findings confirmed the central role of N-terminal tyrosine sites of SLP-76 in the pathway and also shed light on novel signaling events that are uniquely regulated by SLP-76 N-terminal tyrosine residues. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

SRC Homology 2 Domain-containing Leukocyte Phosphoprotein of 76 kDa (SLP-76) N-terminal Tyrosine Residues Regulate a Dynamic Signaling Equilibrium Involving Feedback of Proximal T-cell Receptor (TCR) Signaling*

PubMed Central

Ji, Qinqin; Ding, Yiyuan; Salomon, Arthur R.

2015-01-01

SRC homology 2 domain-containing leukocyte phosphoprotein of 76 kDa (SLP-76) is a cytosolic adaptor protein that plays an important role in the T-cell receptor–mediated T-cell signaling pathway. SLP-76 links proximal receptor stimulation to downstream effectors through interaction with many signaling proteins. Previous studies showed that mutation of three tyrosine residues, Tyr112, Tyr128, and Tyr145, in the N terminus of SLP-76 results in severely impaired phosphorylation and activation of Itk and PLCγ1, which leads to defective calcium mobilization, Erk activation, and NFAT activation. To expand our knowledge of the role of N-terminal phosphorylation of SLP-76 from these three tyrosine sites, we characterized nearly 1000 tyrosine phosphorylation sites via mass spectrometry in SLP-76 reconstituted wild-type cells and SLP-76 mutant cells in which three tyrosine residues were replaced with phenylalanines (Y3F mutant). Mutation of the three N-terminal tyrosine residues of SLP-76 phenocopied SLP-76-deficient cells for the majority of tyrosine phosphorylation sites observed, including feedback on proximal T-cell receptor signaling proteins. Meanwhile, reversed phosphorylation changes were observed on Tyr192 of Lck when we compared mutants to the complete removal of SLP-76. In addition, N-terminal tyrosine sites of SLP-76 also perturbed phosphorylation of Tyr440 of Fyn, Tyr702 of PLCγ1, Tyr204, Tyr397, and Tyr69 of ZAP-70, revealing new modes of regulation on these sites. All these findings confirmed the central role of N-terminal tyrosine sites of SLP-76 in the pathway and also shed light on novel signaling events that are uniquely regulated by SLP-76 N-terminal tyrosine residues. PMID:25316710

RAFTK, a novel member of the focal adhesion kinase family, is phosphorylated and associates with signaling molecules upon activation of mature T lymphocytes.

PubMed

Ganju, R K; Hatch, W C; Avraham, H; Ona, M A; Druker, B; Avraham, S; Groopman, J E

1997-03-17

The related adhesion focal tyrosine kinase (RAFTK), a recently discovered member of the focal adhesion kinase family, has previously been reported to participate in signal transduction in neuronal cells, megakaryocytes, and B lymphocytes. We have found that RAFTK is constitutively expressed in human T cells and is rapidly phosphorylated upon the activation of the T cell receptor (TCR). This activation also results in an increase in the autophosphorylation and kinase activity of RAFTK. After its stimulation, there was an increase in the association of the src cytoplasmic tyrosine kinase Fyn and the adapter protein Grb2. This association was mediated through the SH2 domains of Fyn and Grb2. RAFTK also co-immunoprecipitates with the SH2 domain of Lck and with the cytoskeletal protein paxillin through its COOH-terminal proline-rich domain. The tyrosine phosphorylation of RAFTK after T cell receptor-mediated stimulation was reduced by the pretreatment of cells with cytochalasin D, suggesting the role of the cytoskeleton in this process. These observations indicate that RAFTK participates in T cell receptor signaling and may act to link signals from the cell surface to the cytoskeleton and thereby affect the host immune response.

Design and evaluation of a wireless sensor network based aircraft strength testing system.

PubMed

Wu, Jian; Yuan, Shenfang; Zhou, Genyuan; Ji, Sai; Wang, Zilong; Wang, Yang

2009-01-01

The verification of aerospace structures, including full-scale fatigue and static test programs, is essential for structure strength design and evaluation. However, the current overall ground strength testing systems employ a large number of wires for communication among sensors and data acquisition facilities. The centralized data processing makes test programs lack efficiency and intelligence. Wireless sensor network (WSN) technology might be expected to address the limitations of cable-based aeronautical ground testing systems. This paper presents a wireless sensor network based aircraft strength testing (AST) system design and its evaluation on a real aircraft specimen. In this paper, a miniature, high-precision, and shock-proof wireless sensor node is designed for multi-channel strain gauge signal conditioning and monitoring. A cluster-star network topology protocol and application layer interface are designed in detail. To verify the functionality of the designed wireless sensor network for strength testing capability, a multi-point WSN based AST system is developed for static testing of a real aircraft undercarriage. Based on the designed wireless sensor nodes, the wireless sensor network is deployed to gather, process, and transmit strain gauge signals and monitor results under different static test loads. This paper shows the efficiency of the wireless sensor network based AST system, compared to a conventional AST system.

Design and Evaluation of a Wireless Sensor Network Based Aircraft Strength Testing System

PubMed Central

Wu, Jian; Yuan, Shenfang; Zhou, Genyuan; Ji, Sai; Wang, Zilong; Wang, Yang

2009-01-01

The verification of aerospace structures, including full-scale fatigue and static test programs, is essential for structure strength design and evaluation. However, the current overall ground strength testing systems employ a large number of wires for communication among sensors and data acquisition facilities. The centralized data processing makes test programs lack efficiency and intelligence. Wireless sensor network (WSN) technology might be expected to address the limitations of cable-based aeronautical ground testing systems. This paper presents a wireless sensor network based aircraft strength testing (AST) system design and its evaluation on a real aircraft specimen. In this paper, a miniature, high-precision, and shock-proof wireless sensor node is designed for multi-channel strain gauge signal conditioning and monitoring. A cluster-star network topology protocol and application layer interface are designed in detail. To verify the functionality of the designed wireless sensor network for strength testing capability, a multi-point WSN based AST system is developed for static testing of a real aircraft undercarriage. Based on the designed wireless sensor nodes, the wireless sensor network is deployed to gather, process, and transmit strain gauge signals and monitor results under different static test loads. This paper shows the efficiency of the wireless sensor network based AST system, compared to a conventional AST system. PMID:22408521

The adaptor protein SLP-76 regulates HIV-1 release and cell to cell transmission in T-cells

PubMed Central

Nagaraja, Tirumuru; Anand, Appakkudal R.; Zhao, Helong; Ganju, Ramesh K.

2014-01-01

HIV-1 infection in T-cells is regulated by T-cell receptor (TCR) activation. However, the cellular proteins of the TCR pathway that regulate HIV-1 infection are poorly characterized. Here, we elucidated the role of SLP-76, a key adaptor protein of the TCR signaling complex, in HIV-1 infection. We observed a significant reduction of HIV-1 virus production in SLP-76-deficient Jurkat T-cells compared to wild-type and SLP-76-reconstituted Jurkat T-cells. We further confirmed the role of SLP-76 in HIV-1 infection by siRNA-mediated knockdown in MT4 cells and PBMCs. Structural-functional analysis revealed that the amino-terminal domain of SLP-76 was important for regulating HIV-1 infection. Further mechanistic studies revealed that lack of SLP-76 impaired virus release, but did not affect viral entry, integration and transcription. We also showed that SLP-76 plays a critical role in cell-to-cell transmission of HIV-1. Signaling studies revealed that SLP-76 associated with viral Nef protein and multiple signaling molecules during HIV-1 infection. Furthermore, SLP-76 facilitated the association of Nef and F-actin, suggesting that SLP-76 mediates the formation of a signaling complex that may regulate viral release via cytoskeletal changes. Taken together, our studies demonstrate a novel role for the adaptor molecule, SLP-76 in regulating HIV-1 infection in T-cells with potential to develop innovative strategies against HIV-1. PMID:22323535

Normal T lymphocytes can express two different T cell receptor beta chains: implications for the mechanism of allelic exclusion

PubMed Central

1995-01-01

We have examined the extent of allelic exclusion at the T cell receptor (TCR) beta locus using monoclonal antibodies specific for V beta products. A small proportion (approximately 1%) of human peripheral blood T cells express two V beta as determined by flow cytometric analysis, isolation of representative clones, and sequencing of the corresponding V beta chains. Dual beta T cells are present in both the CD45R0+ and CD45R0- subset. These results indicate that dual beta expression is compatible with both central and peripheral selection. They also suggest that the substantial degree of TCR beta allelic exclusion is dependent only on asynchronous rearrangements at the beta locus, whereas the role of the pre-TCR is limited to signaling the presence of at least one functional beta protein. PMID:7699339

Postactivation potentiation biases maximal isometric strength assessment.

PubMed

Lima, Leonardo Coelho Rabello; Oliveira, Felipe Bruno Dias; Oliveira, Thiago Pires; Assumpção, Claudio de Oliveira; Greco, Camila Coelho; Cardozo, Adalgiso Croscato; Denadai, Benedito Sérgio

2014-01-01

Postactivation potentiation (PAP) is known to enhance force production. Maximal isometric strength assessment protocols usually consist of two or more maximal voluntary isometric contractions (MVCs). The objective of this study was to determine if PAP would influence isometric strength assessment. Healthy male volunteers (n = 23) performed two five-second MVCs separated by a 180-seconds interval. Changes in isometric peak torque (IPT), time to achieve it (tPTI), contractile impulse (CI), root mean square of the electromyographic signal during PTI (RMS), and rate of torque development (RTD), in different intervals, were measured. Significant increases in IPT (240.6 ± 55.7 N·m versus 248.9 ± 55.1 N·m), RTD (746 ± 152 N·m·s(-1) versus 727 ± 158 N·m·s(-1)), and RMS (59.1 ± 12.2% RMSMAX  versus 54.8 ± 9.4% RMSMAX) were found on the second MVC. tPTI decreased significantly on the second MVC (2373 ± 1200 ms versus 2784 ± 1226 ms). We conclude that a first MVC leads to PAP that elicits significant enhancements in strength-related variables of a second MVC performed 180 seconds later. If disconsidered, this phenomenon might bias maximal isometric strength assessment, overestimating some of these variables.

Strength and Aerobic Exercises Improve Spatial Memory in Aging Rats Through Stimulating Distinct Neuroplasticity Mechanisms.

PubMed

Vilela, Thais Ceresér; Muller, Alexandre Pastoris; Damiani, Adriani Paganini; Macan, Tamires Pavei; da Silva, Sabrina; Canteiro, Paula Bortoluzzi; de Sena Casagrande, Alisson; Pedroso, Giulia Dos Santos; Nesi, Renata Tiscoski; de Andrade, Vanessa Moraes; de Pinho, Ricardo Aurino

2017-12-01

Aging is associated with impaired cognition and memory and increased susceptibility to neurodegenerative disorders. Physical exercise is neuroprotective; however, the major evidence of this effect involves studies of only aerobic training in young animals. The benefits of other exercise protocols such as strength training in aged animals remains unknown. Here, we investigated the effect of aerobic and strength training on spatial memory and hippocampal plasticity in aging rats. Aging Wistar rats performed aerobic or strength training for 50 min 3 to 4 days/week for 8 weeks. Spatial memory and neurotrophic and glutamatergic signaling in the hippocampus of aged rats were evaluated after aerobic or strength training. Both aerobic and strength training improved cognition during the performance of a spatial memory task. Remarkably, the improvement in spatial memory was accompanied by an increase in synaptic plasticity proteins within the hippocampus after exercise training, with some differences in the intracellular functions of those proteins between the two exercise protocols. Moreover, neurotrophic signaling (CREB, BDNF, and the P75 NTR receptor) increased after training for both exercise protocols, and aerobic exercise specifically increased glutamatergic proteins (NMDA receptor and PSD-95). We also observed a decrease in DNA damage after aerobic training. In contrast, strength training increased levels of PKCα and the proinflammatory factors TNF-α and IL-1β. Overall, our results show that both aerobic and strength training improved spatial memory in aging rats through inducing distinct molecular mechanisms of neuroplasticity. Our findings extend the idea that exercise protocols can be used to improve cognition during aging.

T-cell triggering thresholds are modulated by the number of antigen within individual T-cell receptor clusters

DOE Office of Scientific and Technical Information (OSTI.GOV)

Manz, Boryana N.; Jackson, Bryan L.; Petit, Rebecca S.

2011-05-31

T cells react to extremely small numbers of activating agonist peptides. Spatial organization of T-cell receptors (TCR) and their peptide-major histocompatibility complex (pMHC) ligands into microclusters is correlated with T-cell activation. In this study, we have designed an experimental strategy that enables control over the number of agonist peptides per TCR cluster, without altering the total number engaged by the cell. Supported membranes, partitioned with grids of barriers to lateral mobility, provide an effective way of limiting the total number of pMHC ligands that may be assembled within a single TCR cluster. Observations directly reveal that restriction of pMHC contentmore » within individual TCR clusters can decrease T-cell sensitivity for triggering initial calcium flux at fixed total pMHC density. Further analysis suggests that triggering thresholds are determined by the number of activating ligands available to individual TCR clusters, not by the total number encountered by the cell. Results from a series of experiments in which the overall agonist density and the maximum number of agonist per TCR cluster are independently varied in primary T cells indicate that the most probable minimal triggering unit for calcium signaling is at least four pMHC in a single cluster for this system. In conclusion, this threshold is unchanged by inclusion of coagonist pMHC, but costimulation of CD28 by CD80 can modulate the threshold lower.« less

Proceedings of the Second Annual Symposium for Nondestructive Evaluation of Bond Strength

NASA Technical Reports Server (NTRS)

Roberts, Mark J. (Compiler)

1999-01-01

Ultrasonics, microwaves, optically stimulated electron emission (OSEE), and computational chemistry approaches have shown relevance to bond strength determination. Nonlinear ultrasonic nondestructive evaluation methods, however, have shown the most effectiveness over other methods on adhesive bond analysis. Correlation to changes in higher order material properties due to microstructural changes using nonlinear ultrasonics has been shown related to bond strength. Nonlinear ultrasonic energy is an order of magnitude more sensitive than linear ultrasound to these material parameter changes and to acoustic velocity changes caused by the acoustoelastic effect when a bond is prestressed. Signal correlations between non-linear ultrasonic measurements and initialization of bond failures have been measured. This paper reviews bond strength research efforts presented by university and industry experts at the Second Annual Symposium for Nondestructive Evaluation of Bond Strength organized by the NDE Sciences Branch at NASA Langley in November 1998.

Functional requirements for inhibitory signal transmission by the immunomodulatory receptor CD300a.

PubMed

DeBell, Karen E; Simhadri, Venkateswara R; Mariano, John L; Borrego, Francisco

2012-04-26

Activation signals can be negatively regulated by cell surface receptors bearing immunoreceptor tyrosine-based inhibitory motifs (ITIMs). CD300a, an ITIM bearing type I transmembrane protein, is expressed on many hematopoietic cells, including subsets of lymphocytes. We have taken two approaches to further define the mechanism by which CD300a acts as an inhibitor of immune cell receptor signaling. First, we have expressed in Jurkat T cells a chimeric receptor consisting of the extracellular domains of killer-cell immunoglobulin-like receptor (KIR)2DL2 fused to the transmembrane and cytoplasmic segments of CD300a (KIR-CD300a) to explore surrogate ligand-stimulated inhibition of superantigen stimulated T cell receptor (TCR) mediated cell signaling. We found that intact CD300a ITIMs were essential for inhibition and that the tyrosine phosphorylation of these ITIMs required the src tyrosine kinase Lck. Tyrosine phosphorylation of the CD300a ITIMs created docking sites for both src homology 2 domain containing protein tyrosine phosphatase (SHP)-1 and SHP-2. Suppression of SHP-1 and SHP-2 expression in KIR-CD300a Jurkat T cells with siRNA and the use of DT40 chicken B cell lines expressing CD300a and deficient in several phosphatases revealed that SHP-1, but not SHP-2 or the src homology 2 domain containing inositol 5' phosphatase SHIP, was utilized by CD300a for its inhibitory activity. These studies provide new insights into the function of CD300a in tuning T and B cell responses.

Profiling calcium signals of in vitro polarized human effector CD4+ T cells.

PubMed

Kircher, Sarah; Merino-Wong, Maylin; Niemeyer, Barbara A; Alansary, Dalia

2018-06-01

Differentiation of naïve CD4 + T cells into effector subtypes with distinct cytokine profiles and physiological roles is a tightly regulated process, the imbalance of which can lead to an inadequate immune response or autoimmune disease. The crucial role of Ca 2+ signals, mainly mediated by the store operated Ca 2+ entry (SOCE) in shaping the immune response is well described. However, it is unclear if human effector CD4 + T cell subsets show differential Ca 2+ signatures in response to different stimulation methods. Herein, we provide optimized in vitro culture conditions for polarization of human CD4 + effector T cells and characterize their SOCE following both pharmacological store depletion and direct T-cell receptor (TCR) activation. Moreover, we measured whole cell Ca 2+ release activated Ca 2+ currents (I CRAC ) and investigated whether the observed differences correlate to the expression of CRAC genes. Our results show that Ca 2+ profiles of helper CD4 + Th1, Th2 and Th17 are distinct and in part shaped by the intensity of stimulation. Regulatory T cells (Treg) are unique being the subtype with the most prominent SOCE response. Analysis of in vivo differentiated Treg unraveled the role of differential expression of ORAI2 in fine-tuning signals in Treg vs. conventional CD4 + T cells. Copyright © 2018 The Author(s). Published by Elsevier B.V. All rights reserved.

Signal honesty and predation risk among a closely related group of aposematic species

PubMed Central

María Arenas, Lina; Walter, Dominic; Stevens, Martin

2015-01-01

Many animals have bright colours to warn predators that they have defences and are not worth attacking. However, it remains unclear whether the strength of warning colours reliably indicate levels of defence. Few studies have unambiguously established if warning signals are honest, and have rarely considered predator vision or conspicuousness against the background. Importantly, little data exists either on how differences in signal strength translate into survival advantages. Ladybirds exhibit impressive variation in coloration both among and within species. Here we demonstrate that different levels of toxicity exist among and within ladybird species, and that signal contrast against the background is a good predictor of toxicity, showing that the colours are honest signals. Furthermore, field experiments with ladybird models created with regards to predator vision show that models with lower conspicuousness were attacked more frequently. This provides one of the most comprehensive studies on signal honesty in warning coloration to date. PMID:26046332

The fracture strength and frictional strength of Weber Sandstone

USGS Publications Warehouse

Byerlee, J.D.

1975-01-01

The fracture strength and frictional strength of Weber Sandstone have been measured as a function of confining pressure and pore pressure. Both the fracture strength and the frictional strength obey the law of effective stress, that is, the strength is determined not by the confining pressure alone but by the difference between the confining pressure and the pore pressure. The fracture strength of the rock varies by as much as 20 per cent depending on the cement between the grains, but the frictional strength is independent of lithology. Over the range 0 2 kb, ??=0??5 + 0??6??n. This relationship also holds for other rocks such as gabbro, dunite, serpentinite, granite and limestone. ?? 1975.

Rejection of syngeneic colon carcinoma by CTLs expressing single-chain antibody receptors codelivering CD28 costimulation.

PubMed

Haynes, Nicole M; Trapani, Joseph A; Teng, Michele W L; Jackson, Jacob T; Cerruti, Loretta; Jane, Stephen M; Kershaw, Michael H; Smyth, Mark J; Darcy, Phillip K

2002-11-15

A new strategy to improve the therapeutic utility of redirected T cells for cancer involves the development of novel Ag-specific chimeric receptors capable of stimulating optimal and sustained T cell antitumor activity in vivo. Given that T cells require both primary and costimulatory signals for optimal activation and that many tumors do not express critical costimulatory ligands, modified single-chain Ab receptors have been engineered to codeliver CD28 costimulation. In this study, we have compared the antitumor potency of primary T lymphocytes expressing carcinoembryonic Ag (CEA)-reactive chimeric receptors that incorporate either TCR-zeta or CD28/TCR-zeta signaling. Although both receptor-transduced T cell effector populations demonstrated cytolysis of CEA(+) tumors in vitro, T cells expressing the single-chain variable fragment of Ig (scFv)-CD28-zeta chimera had a far greater capacity to control the growth of CEA(+) xenogeneic and syngeneic colon carcinomas in vivo. The observed enhanced antitumor activity of T cells expressing the scFv-CD28-zeta receptor was critically dependent on perforin and the production of IFN-gamma. Overall, this study has illustrated the ability of a chimeric scFv receptor capable of harnessing the signaling machinery of both TCR-zeta and CD28 to augment T cell immunity against tumors that have lost expression of both MHC/peptide and costimulatory ligands in vivo.

T cell receptor–induced phosphoinositide-3-kinase p110δ activity is required for T cell localization to antigenic tissue in mice

PubMed Central

Jarmin, Sarah J.; David, Rachel; Ma, Liang; Chai, Jan-Guo; Dewchand, Hamlata; Takesono, Aya; Ridley, Anne J.; Okkenhaug, Klaus; Marelli-Berg, Federica M.

2008-01-01

The establishment of T cell–mediated inflammation requires the migration of primed T lymphocytes from the blood stream and their retention in antigenic sites. While naive T lymphocyte recirculation in the lymph and blood is constitutively regulated and occurs in the absence of inflammation, the recruitment of primed T cells to nonlymphoid tissue and their retention at the site are enhanced by various inflammatory signals, including TCR engagement by antigen-displaying endothelium and resident antigen-presenting cells. In this study, we investigated whether signals downstream of TCR ligation mediated by the phosphoinositide-3-kinase (PI3K) subunit p110δ contributed to the regulation of these events. T lymphocytes from mice expressing catalytically inactive p110δ displayed normal constitutive trafficking and migratory responses to nonspecific stimuli. However, these cells lost susceptibility to TCR-induced migration and failed to localize efficiently to antigenic tissue. Importantly, we showed that antigen-induced T cell trafficking and subsequent inflammation was abrogated by selective pharmacological inhibition of PI3K p110δ activity. These observations suggest that pharmacological targeting of p110δ activity is a viable strategy for the therapy of T cell–mediated pathology. PMID:18259608

Non-T cell activation linker (NTAL) proteolytic cleavage as a terminator of activatory intracellular signals.

PubMed

Arbulo-Echevarria, Mikel M; Muñoz-Miranda, Juan Pedro; Caballero-García, Andrés; Poveda-Díaz, José L; Fernández-Ponce, Cecilia; Durán-Ruiz, M Carmen; Miazek, Arkadiusz; García-Cózar, Francisco; Aguado, Enrique

2016-08-01

Non-T cell activation linker is an adaptor protein that is tyrosine phosphorylated upon cross-linking of immune receptors expressed on B lymphocytes, NK cells, macrophages, basophils, or mast cells, allowing the recruitment of cytosolic mediators for downstream signaling pathways. Fas receptor acts mainly as a death receptor, and when cross-linked with Fas ligand, many proteins are proteolytically cleaved, including several signaling molecules in T and B cells. Fas receptor triggering also interferes with TCR intracellular signals, probably by means of proteolytic cleavage of several adaptor proteins. We have previously found that the adaptor linker for activation of T cells, evolutionarily related to non-T cell activation linker, is cleaved upon proapoptotic stimuli in T lymphocytes and thymocytes, in a tyrosine phosphorylation-dependent fashion. Here, we describe non-T cell activation linker proteolytic cleavage triggered in human B cells and monocytes by Fas cross-linking and staurosporine treatment. Non-T cell activation linker is cleaved, producing an N-terminal fragment of ∼22 kDa, and such cleavage is abrogated in the presence of caspase 8/granzyme B and caspase 3 inhibitors. Moreover, we have identified an aspartic acid residue at which non-T cell activation linker is cleaved, which similar to linker for activation of T cells, this aspartic acid residue is located close to tyrosine and serine residues, suggesting an interdependence of phosphorylation and proteolytic cleavage. Consistently, induction of non-T cell activation linker phosphorylation by pervanadate inhibits its cleavage. Interestingly, the truncated isoform of non-T cell activation linker, generated after cleavage, has a decreased signaling ability when compared with the full-length molecule. Altogether, our results suggest that cleavage of transmembrane adaptors constitutes a general mechanism for signal termination of immune receptors. © Society for Leukocyte Biology.

Defining the expression hierarchy of latent T-cell epitopes in Epstein-Barr virus infection with TCR-like antibodies

PubMed Central

Sim, Adrian Chong Nyi; Too, Chien Tei; Oo, Min Zin; Lai, Junyun; Eio, Michelle Yating; Song, Zhenying; Srinivasan, Nalini; Tan, Diane Ai Lin; Pang, Shyue Wei; Gan, Shu Uin; Lee, Kok Onn; Loh, Thomas Kwok Seng; Chen, Jianzhu; Chan, Soh Ha; MacAry, Paul Anthony

2013-01-01

Epstein-Barr virus (EBV) is a gamma herpesvirus that causes a life-long latent infection in human hosts. The latent gene products LMP1, LMP2A and EBNA1 are expressed by EBV-associated tumors and peptide epitopes derived from these can be targeted by CD8 Cytotoxic T-Lymphocyte (CTL) lines. Whilst CTL-based methodologies can be utilized to infer the presence of specific latent epitopes, they do not allow a direct visualization or quantitation of these epitopes. Here, we describe the characterization of three TCR-like monoclonal antibodies (mAbs) targeting the latent epitopes LMP1125–133, LMP2A426–434 or EBNA1562–570 in association with HLA-A0201. These are employed to map the expression hierarchy of endogenously generated EBV epitopes. The dominance of EBNA1562–570 in association with HLA-A0201 was consistently observed in cell lines and EBV-associated tumor biopsies. These data highlight the discordance between MHC-epitope density and frequencies of associated CTL with implications for cell-based immunotherapies and/or vaccines for EBV-associated disease. PMID:24240815

Characterization of the mechanical and thermal interface of copper films on carbon substrates modified by boron based interlayers

PubMed Central

Schäfer, D.; Eisenmenger-Sittner, C.; Chirtoc, Mihai; Kijamnajsuk, P.; Kornfeind, N.; Hutter, H.; Neubauer, E.; Kitzmantel, M.

2011-01-01

The manipulation of mechanical and thermal interfaces is essential for the design of modern composites. Amongst these are copper carbon composites which can exhibit excellent heat conductivities if the Cu/C interface is affected by a suitable interlayer to minimize the Thermal Contact Resistance (TCR) and to maximize the adhesion strength between Cu and C. In this paper we report on the effect of boron based interlayers on wetting, mechanical adhesion and on the TCR of Cu coatings deposited on glassy carbon substrates by magnetron sputtering. The interlayers were 5 nm thick and consisted of pure B and B with additions of the carbide forming metals Mo, Ti and Cr in the range of 5 at.% relative to B. The interlayers were deposited by RF magnetron sputtering from either a pure B target or from a composite target. The interlayer composition was checked by Auger Electron Spectroscopy and found to be homogenous within the whole film. The system C-substrate/interlayer/Cu coating was characterized in as deposited samples and samples heat treated for 30 min at 800 °C under High Vacuum (HV), which mimics typical hot pressing parameters during composite formation. Material transport during heat treatment was investigated by Secondary Ion Mass Spectroscopy (SIMS). The de-wetting and hole formation in the Cu coating upon heat treatment were studied by Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM). The adhesion of the Cu coating was evaluated by mechanical pull-off testing. The TCR was assessed by infrared photothermal radiometry (PTR). A correlation between the adhesion strength and the value of the TCR which was measured by PTR was determined for as deposited as well as for heat treated samples. PMID:22241938

Radio Frequency Signals in Jupiter's Atmosphere

PubMed

Lanzerotti; Rinnert; Dehmel; Gliem; Krider; Uman; Bach

1996-05-10

During the Galileo probe's descent through Jupiter's atmosphere, under the ionosphere, the lightning and radio emission detector measured radio frequency signals at levels significantly above the probe's electromagnetic noise. The signal strengths at 3 and 15 kilohertz were relatively large at the beginning of the descent, decreased with depth to a pressure level of about 5 bars, and then increased slowly until the end of the mission. The 15-kilohertz signals show arrival direction anisotropies. Measurements of radio frequency wave forms show that the probe passed through an atmospheric region that did not support lightning within at least 100 kilometers and more likely a few thousand kilometers of the descent trajectory. The apparent opacity of the jovian atmosphere increases sharply at pressures greater than about 4 bars.

MHC drives TCR repertoire shaping, but not maturation, in recent thymic emigrants1

PubMed Central

Houston, Evan G.; Fink, Pamela J.

2009-01-01

After developing in the thymus, recent thymic emigrants (RTEs) enter the lymphoid periphery and undergo a maturation process as they transition into the mature naïve (MN) T cell compartment. This maturation presumably shapes RTEs into a pool of T cells best fit to function robustly in the periphery without causing autoimmunity; however, the mechanism and consequences of this maturation process remain unknown. Using a transgenic mouse system that specifically labels RTEs, we tested the influence of MHC molecules, key drivers of intrathymic T cell selection and naive peripheral T cell homeostasis, in shaping the RTE pool in the lymphoid periphery. We found that the TCRs expressed by RTEs are skewed to longer CDR3 regions compared to those of MN T cells, suggesting that MHC does streamline the TCR repertoire of T cells as they transition from the RTE to the MN T cell stage. This conclusion is borne out in studies in which the representation of individual TCRs was followed as a function of time since thymic egress. Surprisingly, we found that MHC is dispensable for the phenotypic and functional maturation of RTEs. This is an author-produced version of a manuscript accepted for publication in The Journal of Immunology (The JI). The American Association of Immunologists, Inc. (AAI), publisher of The JI, holds the copyright to this manuscript. This version of the manuscript has not yet been copyedited or subjected to editorial proofreading by The JI; hence, it may differ from the final version published in The JI (online and in print). AAI (The JI) is not liable for errors or omissions in this author-produced version of the manuscript or in any version derived from it by the U.S. National Institutes of Health or any other third party. The final, citable version of record can be found at www.jimmunol.org PMID:19915060

Phospho-control of TGF-β superfamily signaling

PubMed Central

Wrighton, Katharine H; Lin, Xia; Feng, Xin-Hua

2010-01-01

Members of the transforming growth factor-β (TGF-β) family control a broad range of cellular responses in metazoan organisms via autocrine, paracrine, and endocrine modes. Thus, aberrant TGF-β signaling can play a key role in the pathogenesis of several diseases, including cancer. TGF-β signaling pathways are activated by a short phospho-cascade, from receptor phosphorylation to the subsequent phosphorylation and activation of downstream signal transducers called R-Smads. R-Smad phosphorylation state determines Smad complex assembly/disassembly, nuclear import/export, transcriptional activity and stability, and is thus the most critical event in TGF-β signaling. Dephosphorylation of R-Smads by specific phosphatases prevents or terminates TGF-β signaling, highlighting the need to consider Smad (de)phosphorylation as a tightly controlled and dynamic event. This article illustrates the essential roles of reversible phosphorylation in controlling the strength and duration of TGF-β signaling and the ensuing physiological responses. PMID:19114991

Immune selection of tumor cells in TCR β-chain transgenic mice.

PubMed

Silaeva, Yulia Yu; Grinenko, Tatyana S; Vagida, Murad S; Kalinina, Anastasia A; Khromykh, Ludmila M; Kazansky, Dmitry B

2014-10-01

The concept of immunological surveillance implies that immunogenic variants of tumor cells arising in the organism can be recognized by the immune system. Tumor progression is provided by somatic evolution of tumor cells under the pressure of the immune system. The loss of MHC Class I molecules on the surface of tumor cells is one of the most known outcomes of immune selection. This study developed a model of immune selection based on the immune response of TCR 1d1 single β-chain transgenic B10.D2(R101) (K(d)I(d)D(b)) mice to allogeneic EL4 (H-2(b)) thymoma cells. In wild-type B10.D2(R101) mice, immunization with EL4 cells induced a vigorous CTL response targeted to the H-2K(b) molecule and results in full rejection of the tumor cells. In contrast, transgenic mice developed a compromised proliferative response in mixed-lymphocyte response assays and were unable to reject transplanted allogeneic EL4 cells. During the immune response to EL4 cells, CD8(+) T-lymphocytes with endogenous β-chains accumulated predominantly in the spleen of transgenic mice and only a small part of the T-lymphocytes expressing transgenic β-chains became CD8(+)CD44(+)CD62L(-) effectors. Then, instead of a full elimination of tumor cells as in wild-type mice, a reproducible prolonged equilibrium phase and subsequent escape was observed in transgenic mice that resulted in death of 90% of the mice in 40-60 days after grafting. Prolonged exposure of tumor cells to the pressure of the immune system in transgenic mice in vivo resulted in a stable loss of H-2K(b) molecules on the EL4 cell surface. Genetic manipulation of the T-lymphocyte repertoire was sufficient to reproduce the classic pattern of interactions between tumor cells and the immune system, usually observed in reliable syngeneic models of anti-tumor immunity. This newly-developed model could be used in further studies of immunoregulatory circuits common for transplantational and anti-tumor immune responses.

Signal processing for order 10 pm accuracy displacement metrology in real-world scientific applications

NASA Technical Reports Server (NTRS)

Halverson, Peter G.; Loya, Frank M.

2004-01-01

This paper describes heterodyne displacement metrology gauge signal processing methods that achieve satisfactory robustness against low signal strength and spurious signals, and good long-term stability. We have a proven displacement-measuring approach that is useful not only to space-optical projects at JPL, but also to the wider field of distance measurements.

Calcium-mediated shaping of naive CD4 T-cell phenotype and function

PubMed Central

Guichard, Vincent; Bonilla, Nelly; Durand, Aurélie; Audemard-Verger, Alexandra; Guilbert, Thomas; Martin, Bruno

2017-01-01

Continuous contact with self-major histocompatibility complex ligands is essential for the survival of naive CD4 T cells. We have previously shown that the resulting tonic TCR signaling also influences their fate upon activation by increasing their ability to differentiate into induced/peripheral regulatory T cells. To decipher the molecular mechanisms governing this process, we here focus on the TCR signaling cascade and demonstrate that a rise in intracellular calcium levels is sufficient to modulate the phenotype of mouse naive CD4 T cells and to increase their sensitivity to regulatory T-cell polarization signals, both processes relying on calcineurin activation. Accordingly, in vivo calcineurin inhibition leads the most self-reactive naive CD4 T cells to adopt the phenotype of their less self-reactive cell-counterparts. Collectively, our findings demonstrate that calcium-mediated activation of the calcineurin pathway acts as a rheostat to shape both the phenotype and effector potential of naive CD4 T cells in the steady-state. PMID:29239722

Function of the nucleotide exchange activity of vav1 in T cell development and activation.

PubMed

Saveliev, Alexander; Vanes, Lesley; Ksionda, Olga; Rapley, Jonathan; Smerdon, Stephen J; Rittinger, Katrin; Tybulewicz, Victor L J

2009-12-15

The guanine nucleotide exchange factor (GEF) Vav1 is essential for transducing T cell antigen receptor (TCR) signals and therefore plays a critical role in the development and activation of T cells. It has been presumed that the GEF activity of Vav1 is important for its function; however, there has been no direct demonstration of this. Here, we generated mice expressing enzymatically inactive, but normally folded, Vav1 protein. Analysis of these mice showed that the GEF activity of Vav1 was necessary for the selection of thymocytes and for the optimal activation of T cells, including signal transduction to Rac1, Akt, and integrins. In contrast, the GEF activity of Vav1 was not required for TCR-induced calcium flux, activation of extracellular signal-regulated kinase and protein kinase D1, and cell polarization. Thus, in T cells, the GEF activity of Vav1 is essential for some, but not all, of its functions.

Improving RF Transmit Power and Received Signal Strength in 2.4 GHz ZigBee Based Active RFID System with Embedded Method

NASA Astrophysics Data System (ADS)

Po'ad, F. A.; Ismail, W.; Jusoh, J. F.

2017-08-01

This paper describes the experiments and analysis conducted on 2.4 GHz embedded active Radio Frequency Identification (RFID) - Wireless Sensor Network (WSN) based system that has been developed for the purposes of location tracking and monitoring in indoor and outdoor environments. Several experiments are conducted to test the effectiveness and performance of the developed system and two of them is by measuring the Radio Frequency (RF) transmitting power and Received Signal Strength (RSS) to prove that the embedded active RFID tag is capable to generate higher transmit power during data transmission and able to provide better RSS reading compared to standalone RFID tag. Experiments are carried out on two RFID tags which are active RFID tag embedded with GPS and GSM (ER2G); and standalone RFID tag communicating with the same active RFID reader. The developed ER2G contributes 12.26 % transmit power and 6.47 % RSS reading higher than standalone RFID tag. The results conclude that the ER2G gives better performance compared to standalone RFID tag and can be used as guidelines for future design improvements.