Science.gov

Sample records for activated effector memory

  1. Curcumin serves as a human kv1.3 blocker to inhibit effector memory T lymphocyte activities.

    PubMed

    Lian, Yi-Tian; Yang, Xiao-Fang; Wang, Zhao-Hui; Yang, Yong; Yang, Ying; Shu, Yan-Wen; Cheng, Long-Xian; Liu, Kun

    2013-09-01

    Curcumin, the principal active component of turmeric, has long been used to treat various diseases in India and China. Recent studies show that curcumin can serve as a therapeutic agent for autoimmune diseases via a variety of mechanisms. Effector memory T cells (T(EM), CCR7⁻ CD45RO⁺ T lymphocyte) have been demonstrated to play a crucial role in the pathogenesis of T cell-mediated autoimmune diseases, such as multiple sclerosis (MS) or rheumatoid arthritis (RA). Kv1.3 channels are predominantly expressed in T(EM) cells and control T(EM) activities. In the present study, we examined the effect of curcumin on human Kv1.3 (hKv1.3) channels stably expressed in HEK-293 cells and its ability to inhibit proliferation and cytokine secretion of T(EM) cells isolated from patients with MS or RA. Curcumin exhibited a direct blockage of hKv1.3 channels in a time-dependent and concentration-dependent manner. Moreover, the activation curve was shifted to a more positive potential, which was consistent with an open-channel blockade. Paralleling hKv1.3 inhibition, curcumin significantly inhibited proliferation and interferon-γ secretion of T(EM) cells. Our findings demonstrate that curcumin is able to inhibit proliferation and proinflammatory cytokine secretion of T(EM) cells probably through inhibition of hKv1.3 channels, which contributes to the potency of curcumin for the treatment of autoimmune diseases. This is probably one of pharmacological mechanisms of curcumin used to treat autoimmune diseases. PMID:23132777

  2. Early effector cells survive the contraction phase in malaria infection and generate both central and effector memory T cells.

    PubMed

    Opata, Michael M; Carpio, Victor H; Ibitokou, Samad A; Dillon, Brian E; Obiero, Joshua M; Stephens, Robin

    2015-06-01

    CD4 T cells orchestrate immunity against blood-stage malaria. However, a major challenge in designing vaccines to the disease is poor understanding of the requirements for the generation of protective memory T cells (Tmem) from responding effector T cells (Teff) in chronic parasite infection. In this study, we use a transgenic mouse model with T cells specific for the merozoite surface protein (MSP)-1 of Plasmodium chabaudi to show that activated T cells generate three distinct Teff subsets with progressive activation phenotypes. The earliest observed Teff subsets (CD127(-)CD62L(hi)CD27(+)) are less divided than CD62L(lo) Teff and express memory genes. Intermediate (CD62L(lo)CD27(+)) effector subsets include the most multicytokine-producing T cells, whereas fully activated (CD62L(lo)CD27(-)) late effector cells have a terminal Teff phenotype (PD-1(+), Fas(hi), AnnexinV(+)). We show that although IL-2 promotes expansion, it actually slows terminal effector differentiation. Using adoptive transfer, we show that only early Teff survive the contraction phase and generate the terminal late Teff subsets, whereas in uninfected recipients, they become both central and effector Tmem. Furthermore, we show that progression toward full Teff activation is promoted by increased duration of infection, which in the long-term promotes Tem differentiation. Therefore, we have defined markers of progressive activation of CD4 Teff at the peak of malaria infection, including a subset that survives the contraction phase to make Tmem, and show that Ag and cytokine levels during CD4 T cell expansion influence the proportion of activated cells that can survive contraction and generate memory in malaria infection.

  3. Early effector cells survive the contraction phase in malaria infection and generate both central and effector memory T cells.

    PubMed

    Opata, Michael M; Carpio, Victor H; Ibitokou, Samad A; Dillon, Brian E; Obiero, Joshua M; Stephens, Robin

    2015-06-01

    CD4 T cells orchestrate immunity against blood-stage malaria. However, a major challenge in designing vaccines to the disease is poor understanding of the requirements for the generation of protective memory T cells (Tmem) from responding effector T cells (Teff) in chronic parasite infection. In this study, we use a transgenic mouse model with T cells specific for the merozoite surface protein (MSP)-1 of Plasmodium chabaudi to show that activated T cells generate three distinct Teff subsets with progressive activation phenotypes. The earliest observed Teff subsets (CD127(-)CD62L(hi)CD27(+)) are less divided than CD62L(lo) Teff and express memory genes. Intermediate (CD62L(lo)CD27(+)) effector subsets include the most multicytokine-producing T cells, whereas fully activated (CD62L(lo)CD27(-)) late effector cells have a terminal Teff phenotype (PD-1(+), Fas(hi), AnnexinV(+)). We show that although IL-2 promotes expansion, it actually slows terminal effector differentiation. Using adoptive transfer, we show that only early Teff survive the contraction phase and generate the terminal late Teff subsets, whereas in uninfected recipients, they become both central and effector Tmem. Furthermore, we show that progression toward full Teff activation is promoted by increased duration of infection, which in the long-term promotes Tem differentiation. Therefore, we have defined markers of progressive activation of CD4 Teff at the peak of malaria infection, including a subset that survives the contraction phase to make Tmem, and show that Ag and cytokine levels during CD4 T cell expansion influence the proportion of activated cells that can survive contraction and generate memory in malaria infection. PMID:25911759

  4. Non-random pairing of CD46 isoforms with skewing towards BC2 and C2 in activated and memory/effector T cells

    PubMed Central

    Hansen, Aida S.; Bundgaard, Bettina B.; Møller, Bjarne K.; Höllsberg, Per

    2016-01-01

    CD46 is a glycoprotein with important functions in innate and adaptive immune responses. Functionally different isoforms are generated by alternative splicing at exons 7–9 (BC and C isoforms) and exon 13 (CYT-1 and CYT-2 isoforms) giving rise to BC1, BC2, C1 and C2. We developed a novel real-time PCR assay that allows quantitative comparisons between these isoforms. Their relative frequency in CD4+ T cells from 100 donors revealed a distribution with high interpersonally variability. Importantly, the distribution between the isoforms was not random and although splicing favoured inclusion of exon 8 (BC isoforms), exclusion of exon 8 (C isoforms) was significantly linked to exclusion of exon 13 (CYT-2 isoforms). Despite inter-individual differences, CD4+ and CD8+ T cells, B cells, NK cells and monocytes expressed similar isoform profiles intra-individually. However, memory/effector CD4+ T cells had a significantly higher frequency of CYT-2 when compared with naïve CD4+ T cells. Likewise, in vitro activation of naïve and total CD4+ T cells increased the expression of CYT-2. This indicates that although splicing factors determine a certain expression profile in an individual, the profile can be modulated by external stimuli. This suggests a mechanism by which alterations in CD46 isoforms may temporarily regulate the immune response. PMID:27739531

  5. IL-15 Trans-Signaling with the Superagonist RLI Promotes Effector/Memory CD8+ T Cell Responses and Enhances Antitumor Activity of PD-1 Antagonists.

    PubMed

    Desbois, Mélanie; Le Vu, Pauline; Coutzac, Clélia; Marcheteau, Elie; Béal, Coralie; Terme, Magali; Gey, Alain; Morisseau, Sébastien; Teppaz, Géraldine; Boselli, Lisa; Jacques, Yannick; Béchard, David; Tartour, Eric; Cassard, Lydie; Chaput, Nathalie

    2016-07-01

    Tumors with the help of the surrounding environment facilitate the immune suppression in patients, and immunotherapy can counteract this inhibition. Among immunotherapeutic strategies, the immunostimulatory cytokine IL-15 could represent a serious candidate for the reactivation of antitumor immunity. However, exogenous IL-15 may have a limited impact on patients with cancer due to its dependency on IL-15Rα frequently downregulated in cancer patients. In this work, we studied the antitumor activity of the IL-15 superagonist receptor-linker-IL-15 (RLI), designed to bypass the need of endogenous IL-15Rα. RLI consists of human IL-15 covalently linked to the human IL-15Rα sushi(+) domain. In a mouse model of colorectal carcinoma, RLI as a stand-alone treatment could limit tumor outgrowth only when initiated at an early time of tumor development. At a later time, RLI was not effective, coinciding with the strong accumulation of terminally exhausted programmed cell death-1 (PD-1)(high) T cell Ig mucin-3(+) CD8(+) T cells, suggesting that RLI was not able to reactivate terminally exhausted CD8(+) T cells. Combination with PD-1 blocking Ab showed synergistic activity with RLI, but not with IL-15. RLI could induce a greater accumulation of memory CD8(+) T cells and a stronger effector function in comparison with IL-15. Ex vivo stimulation of tumor-infiltrated lymphocytes from 16 patients with renal cell carcinoma demonstrated 56% of a strong tumor-infiltrated lymphocyte reactivation with the combination anti-PD-1/RLI compared with 43 and 6% with RLI or anti-PD-1, respectively. Altogether, this work provides evidence that the sushi-IL-15Rα/IL-15 fusion protein RLI enhances antitumor activity of anti-PD-1 treatment and is a promising approach to stimulate host immunity.

  6. IL-15 Trans-Signaling with the Superagonist RLI Promotes Effector/Memory CD8+ T Cell Responses and Enhances Antitumor Activity of PD-1 Antagonists.

    PubMed

    Desbois, Mélanie; Le Vu, Pauline; Coutzac, Clélia; Marcheteau, Elie; Béal, Coralie; Terme, Magali; Gey, Alain; Morisseau, Sébastien; Teppaz, Géraldine; Boselli, Lisa; Jacques, Yannick; Béchard, David; Tartour, Eric; Cassard, Lydie; Chaput, Nathalie

    2016-07-01

    Tumors with the help of the surrounding environment facilitate the immune suppression in patients, and immunotherapy can counteract this inhibition. Among immunotherapeutic strategies, the immunostimulatory cytokine IL-15 could represent a serious candidate for the reactivation of antitumor immunity. However, exogenous IL-15 may have a limited impact on patients with cancer due to its dependency on IL-15Rα frequently downregulated in cancer patients. In this work, we studied the antitumor activity of the IL-15 superagonist receptor-linker-IL-15 (RLI), designed to bypass the need of endogenous IL-15Rα. RLI consists of human IL-15 covalently linked to the human IL-15Rα sushi(+) domain. In a mouse model of colorectal carcinoma, RLI as a stand-alone treatment could limit tumor outgrowth only when initiated at an early time of tumor development. At a later time, RLI was not effective, coinciding with the strong accumulation of terminally exhausted programmed cell death-1 (PD-1)(high) T cell Ig mucin-3(+) CD8(+) T cells, suggesting that RLI was not able to reactivate terminally exhausted CD8(+) T cells. Combination with PD-1 blocking Ab showed synergistic activity with RLI, but not with IL-15. RLI could induce a greater accumulation of memory CD8(+) T cells and a stronger effector function in comparison with IL-15. Ex vivo stimulation of tumor-infiltrated lymphocytes from 16 patients with renal cell carcinoma demonstrated 56% of a strong tumor-infiltrated lymphocyte reactivation with the combination anti-PD-1/RLI compared with 43 and 6% with RLI or anti-PD-1, respectively. Altogether, this work provides evidence that the sushi-IL-15Rα/IL-15 fusion protein RLI enhances antitumor activity of anti-PD-1 treatment and is a promising approach to stimulate host immunity. PMID:27217584

  7. Active Flow Effectors for Noise and Separation Control

    NASA Technical Reports Server (NTRS)

    Turner, Travis L.

    2011-01-01

    New flow effector technology for separation control and enhanced mixing is based upon shape memory alloy hybrid composite (SMAHC) technology. The technology allows for variable shape control of aircraft structures through actively deformable surfaces. The flow effectors are made by embedding shape memory alloy actuator material in a composite structure. When thermally actuated, the flow effector def1ects into or out of the flow in a prescribed manner to enhance mixing or induce separation for a variety of applications, including aeroacoustic noise reduction, drag reduction, and f1ight control. The active flow effectors were developed for noise reduction as an alternative to fixed-configuration effectors, such as static chevrons, that cannot be optimized for airframe installation effects or variable operating conditions and cannot be retracted for off-design or fail-safe conditions. Benefits include: Increased vehicle control, overall efficiency, and reduced noise throughout all f1ight regimes, Reduced flow noise, Reduced drag, Simplicity of design and fabrication, Simplicity of control through direct current stimulation, autonomous re sponse to environmental heating, fast re sponse, and a high degree of geometric stability. The concept involves embedding prestrained SMA actuators on one side of the chevron neutral axis in order to generate a thermal moment and def1ect the structure out of plane when heated. The force developed in the host structure during def1ection and the aerodynamic load is used for returning the structure to the retracted position. The chevron design is highly scalable and versatile, and easily affords active and/or autonomous (environmental) control. The technology offers wide-ranging market applications, including aerospace, automotive, and any application that requires flow separation or noise control.

  8. TLR4 ligands lipopolysaccharide and monophosphoryl lipid a differentially regulate effector and memory CD8+ T Cell differentiation.

    PubMed

    Cui, Weiguo; Joshi, Nikhil S; Liu, Ying; Meng, Hailong; Kleinstein, Steven H; Kaech, Susan M

    2014-05-01

    Vaccines formulated with nonreplicating pathogens require adjuvants to help bolster immunogenicity. The role of adjuvants in Ab production has been well studied, but how they influence memory CD8(+) T cell differentiation remains poorly defined. In this study we implemented dendritic cell-mediated immunization to study the effects of commonly used adjuvants, TLR ligands, on effector and memory CD8(+) T cell differentiation in mice. Intriguingly, we found that the TLR4 ligand LPS was far more superior to other TLR ligands in generating memory CD8(+) T cells upon immunization. LPS boosted clonal expansion similar to the other adjuvants, but fewer of the activated CD8(+) T cells died during contraction, generating a larger pool of memory cells. Surprisingly, monophosphoryl lipid A (MPLA), another TLR4 ligand, enhanced clonal expansion of effector CD8(+) T cells, but it also promoted their terminal differentiation and contraction; thus, fewer memory CD8(+) T cells formed, and MPLA-primed animals were less protected against secondary infection compared with those primed with LPS. Furthermore, gene expression profiling revealed that LPS-primed effector cells displayed a stronger pro-memory gene expression signature, whereas the gene expression profile of MPLA-primed effector cells aligned closer with terminal effector CD8(+) T cells. Lastly, we demonstrated that the LPS-TLR4-derived "pro-memory" signals were MyD88, but not Toll/IL-1R domain-containing adapter inducing IFN-β, dependent. This study reveals the influential power of adjuvants on the quantity and quality of CD8(+) T cell memory, and that attention to adjuvant selection is crucial because boosting effector cell expansion may not always equate with more memory T cells or greater protection.

  9. Different Subsets of T Cells, Memory, Effector Functions, and CAR-T Immunotherapy.

    PubMed

    Golubovskaya, Vita; Wu, Lijun

    2016-01-01

    This review is focused on different subsets of T cells: CD4 and CD8, memory and effector functions, and their role in CAR-T therapy--a cellular adoptive immunotherapy with T cells expressing chimeric antigen receptor. The CAR-T cells recognize tumor antigens and induce cytotoxic activities against tumor cells. Recently, differences in T cell functions and the role of memory and effector T cells were shown to be important in CAR-T cell immunotherapy. The CD4⁺ subsets (Th1, Th2, Th9, Th17, Th22, Treg, and Tfh) and CD8⁺ memory and effector subsets differ in extra-cellular (CD25, CD45RO, CD45RA, CCR-7, L-Selectin [CD62L], etc.); intracellular markers (FOXP3); epigenetic and genetic programs; and metabolic pathways (catabolic or anabolic); and these differences can be modulated to improve CAR-T therapy. In addition, CD4⁺ Treg cells suppress the efficacy of CAR-T cell therapy, and different approaches to overcome this suppression are discussed in this review. Thus, next-generation CAR-T immunotherapy can be improved, based on our knowledge of T cell subsets functions, differentiation, proliferation, and signaling pathways to generate more active CAR-T cells against tumors.

  10. Different Subsets of T Cells, Memory, Effector Functions, and CAR-T Immunotherapy

    PubMed Central

    Golubovskaya, Vita; Wu, Lijun

    2016-01-01

    This review is focused on different subsets of T cells: CD4 and CD8, memory and effector functions, and their role in CAR-T therapy––a cellular adoptive immunotherapy with T cells expressing chimeric antigen receptor. The CAR-T cells recognize tumor antigens and induce cytotoxic activities against tumor cells. Recently, differences in T cell functions and the role of memory and effector T cells were shown to be important in CAR-T cell immunotherapy. The CD4+ subsets (Th1, Th2, Th9, Th17, Th22, Treg, and Tfh) and CD8+ memory and effector subsets differ in extra-cellular (CD25, CD45RO, CD45RA, CCR-7, L-Selectin [CD62L], etc.); intracellular markers (FOXP3); epigenetic and genetic programs; and metabolic pathways (catabolic or anabolic); and these differences can be modulated to improve CAR-T therapy. In addition, CD4+ Treg cells suppress the efficacy of CAR-T cell therapy, and different approaches to overcome this suppression are discussed in this review. Thus, next-generation CAR-T immunotherapy can be improved, based on our knowledge of T cell subsets functions, differentiation, proliferation, and signaling pathways to generate more active CAR-T cells against tumors. PMID:26999211

  11. Dicer Regulates the Balance of Short-Lived Effector and Long-Lived Memory CD8 T Cell Lineages

    PubMed Central

    Baumann, Florian M.; Yuzefpolskiy, Yevgeniy; Sarkar, Surojit; Kalia, Vandana

    2016-01-01

    MicroRNAs constitute a major post-transcriptional mechanism for controlling protein expression, and are emerging as key regulators during T cell development and function. Recent reports of augmented CD8 T cell activation and effector differentiation, and aberrant migratory properties upon ablation of Dicer/miRNAs in naïve cells have established a regulatory role of miRNAs during priming. Whether miRNAs continue to exert similar functions or are dispensable during later stages of CD8 T cell expansion and memory differentiation remains unclear. Here, we report a critical role of Dicer/miRNAs in regulating the balance of long-lived memory and short-lived terminal effector fates during the post-priming stages when CD8 T cells undergo clonal expansion to generate a large cytotoxic T lymphocyte (CTL) pool and subsequently differentiate into a quiescent memory state. Conditional ablation of Dicer/miRNAs in early effector CD8 T cells following optimal activation and expression of granzyme B, using unique dicerfl/fl gzmb-cre mice, led to a strikingly diminished peak effector size relative to wild-type antigen-specific cells in the same infectious milieu. Diminished expansion of Dicer-ablated CD8 T cells was associated with lack of sustained antigen-driven proliferation and reduced accumulation of short-lived effector cells. Additionally, Dicer-ablated CD8 T cells exhibited more pronounced contraction after pathogen clearance and comprised a significantly smaller proportion of the memory pool, despite significantly higher proportions of CD127Hi memory precursors at the effector peak. Combined with previous reports of dynamic changes in miRNA expression as CD8 T cells differentiate from naïve to effector and memory states, these findings support distinct stage-specific roles of miRNA-dependent gene regulation during CD8 T cell differentiation. PMID:27627450

  12. Dicer Regulates the Balance of Short-Lived Effector and Long-Lived Memory CD8 T Cell Lineages.

    PubMed

    Baumann, Florian M; Yuzefpolskiy, Yevgeniy; Sarkar, Surojit; Kalia, Vandana

    2016-01-01

    MicroRNAs constitute a major post-transcriptional mechanism for controlling protein expression, and are emerging as key regulators during T cell development and function. Recent reports of augmented CD8 T cell activation and effector differentiation, and aberrant migratory properties upon ablation of Dicer/miRNAs in naïve cells have established a regulatory role of miRNAs during priming. Whether miRNAs continue to exert similar functions or are dispensable during later stages of CD8 T cell expansion and memory differentiation remains unclear. Here, we report a critical role of Dicer/miRNAs in regulating the balance of long-lived memory and short-lived terminal effector fates during the post-priming stages when CD8 T cells undergo clonal expansion to generate a large cytotoxic T lymphocyte (CTL) pool and subsequently differentiate into a quiescent memory state. Conditional ablation of Dicer/miRNAs in early effector CD8 T cells following optimal activation and expression of granzyme B, using unique dicerfl/fl gzmb-cre mice, led to a strikingly diminished peak effector size relative to wild-type antigen-specific cells in the same infectious milieu. Diminished expansion of Dicer-ablated CD8 T cells was associated with lack of sustained antigen-driven proliferation and reduced accumulation of short-lived effector cells. Additionally, Dicer-ablated CD8 T cells exhibited more pronounced contraction after pathogen clearance and comprised a significantly smaller proportion of the memory pool, despite significantly higher proportions of CD127Hi memory precursors at the effector peak. Combined with previous reports of dynamic changes in miRNA expression as CD8 T cells differentiate from naïve to effector and memory states, these findings support distinct stage-specific roles of miRNA-dependent gene regulation during CD8 T cell differentiation. PMID:27627450

  13. IL-15 induces CD4+ effector memory T cell production and tissue emigration in nonhuman primates

    PubMed Central

    Picker, Louis J.; Reed-Inderbitzin, Edward F.; Hagen, Shoko I.; Edgar, John B.; Hansen, Scott G.; Legasse, Alfred; Planer, Shannon; Piatak, Michael; Lifson, Jeffrey D.; Maino, Vernon C.; Axthelm, Michael K.; Villinger, Francois

    2006-01-01

    HIV infection selectively targets CD4+ effector memory T (TEM) cells, resulting in dramatic depletion of CD4+ T cells in mucosal effector sites in early infection. Regeneration of the TEM cell compartment is slow and incomplete, even when viral replication is controlled by antiretroviral therapy (ART). Here, we demonstrate that IL-15 dramatically increases in vivo proliferation of rhesus macaque (RM) CD4+ and CD8+ TEM cells with little effect on the naive or central memory T (TCM) cell subsets, a response pattern that is quite distinct from that of either IL-2 or IL-7. TEM cells produced in response to IL-15 did not accumulate in blood. Rather, 5-bromo-2′-deoxyuridine (BrdU) labeling studies suggest that many of these cells rapidly disperse to extralymphoid effector sites, where they manifest (slow) decay kinetics indistinguishable from that of untreated controls. In RMs with uncontrolled SIV infection and highly activated immune systems, IL-15 did not significantly increase CD4+ TEM cell proliferation, but with virologic control and concomitant reduction in immune activation by ART, IL-15 responsiveness was again observed. These data suggest that therapeutic use of IL-15 in the setting of ART might facilitate specific restoration of the CD4+ T cell compartment that is the primary target of HIV with less risk of exhausting precursor T cell compartments or generating potentially deleterious regulatory subsets. PMID:16691294

  14. Active membrane cholesterol as a physiological effector.

    PubMed

    Lange, Yvonne; Steck, Theodore L

    2016-09-01

    Sterols associate preferentially with plasma membrane sphingolipids and saturated phospholipids to form stoichiometric complexes. Cholesterol in molar excess of the capacity of these polar bilayer lipids has a high accessibility and fugacity; we call this fraction active cholesterol. This review first considers how active cholesterol serves as an upstream regulator of cellular sterol homeostasis. The mechanism appears to utilize the redistribution of active cholesterol down its diffusional gradient to the endoplasmic reticulum and mitochondria, where it binds multiple effectors and directs their feedback activity. We have also reviewed a broad literature in search of a role for active cholesterol (as opposed to bulk cholesterol or lipid domains such as rafts) in the activity of diverse membrane proteins. Several systems provide such evidence, implicating, in particular, caveolin-1, various kinds of ABC-type cholesterol transporters, solute transporters, receptors and ion channels. We suggest that this larger role for active cholesterol warrants close attention and can be tested easily.

  15. Transcriptional regulation of effector and memory CD8+ T cell fates

    PubMed Central

    Thaventhiran, James E. D.; Fearon, Douglas T.; Gattinoni, Luca

    2013-01-01

    Immunity to intracellular pathogens and cancer relies on the generation of robust CD8+ T cell effector responses as well as the establishment of immunological memory. During a primary immune response CD8+ T cells experience diverse extracellular environmental cues and cell-cell interactions that trigger downstream transcriptional programs ultimately guiding a CD8+ T cell to undertake either an effector or a memory cell fate. Here, we discuss our current understanding of the signaling pathways and transcriptional networks that regulate effector and memory commitment in CD8+ T lymphocytes. PMID:23747000

  16. The dual targeting of immunosuppressive cells and oxidants promotes effector and memory T-cell functions against lung cancer

    PubMed Central

    Sawant, Anandi; Schafer, Cara C; Ponnazhagan, Selvarangan; Deshane, Jessy S

    2014-01-01

    We have recently demonstrated that the combination of gemcitabine and a superoxide dismutase mimetic protects mice against lung cancer by suppressing the functions of myeloid-derived suppressor cells and by activating memory CD8+ T-cell responses. Persistent memory cells exhibited a glycolytic metabolism, which may have directly enhanced their effector functions. This combinatorial therapeutic regimen may reduce the propensity of some cancer patients to relapse. PMID:24711958

  17. Co-administration of α-GalCer analog and TLR4 agonist induces robust CD8+ T-cell responses to PyCS protein and WT-1 antigen and activates memory-like effector NKT cells

    PubMed Central

    Coelho-dos-Reis, Jordana G.; Huang, Jing; Tsao, Tiffany; Pereira, Felipe V.; Funakoshi, Ryota; Nakajima, Hiroko; Sugiyama, Haruo; Tsuji, Moriya

    2016-01-01

    In the present study, the combined adjuvant effect of 7DW8-5, a potent α-GalCer-analog, and monophosphoryl lipid A (MPLA), a TLR4 agonist, on the induction of vaccine-induced CD8+ T-cell responses and protective immunity was evaluated. Mice were immunized with peptides corresponding to the CD8+ T-cell epitopes of a malaria antigen, a circumsporozoite protein of Plasmodium yoelii, and a tumor antigen, a Wilms Tumor antigen-1 (WT-1), together with 7DW8-5 and MPLA, as an adjuvant. These immunization regimens were able to induce higher levels of CD8+ T-cell responses and, ultimately, enhanced levels of protection against malaria and tumor challenges compared to the levels induced by immunization with peptides mixed with 7DW8-5 or MPLA alone. Co-administration of 7DW8-5 and MPLA induces activation of memory-like effector natural killer T (NKT) cells, i.e. CD44+CD62L−NKT cells. Our study indicates that 7DW8-5 greatly enhances important synergistic pathways associated to memory immune responses when co-administered with MPLA, thus rendering this combination of adjuvants a novel vaccine adjuvant formulation. PMID:27132023

  18. Co-administration of α-GalCer analog and TLR4 agonist induces robust CD8(+) T-cell responses to PyCS protein and WT-1 antigen and activates memory-like effector NKT cells.

    PubMed

    Coelho-Dos-Reis, Jordana G; Huang, Jing; Tsao, Tiffany; Pereira, Felipe V; Funakoshi, Ryota; Nakajima, Hiroko; Sugiyama, Haruo; Tsuji, Moriya

    2016-07-01

    In the present study, the combined adjuvant effect of 7DW8-5, a potent α-GalCer-analog, and monophosphoryl lipid A (MPLA), a TLR4 agonist, on the induction of vaccine-induced CD8(+) T-cell responses and protective immunity was evaluated. Mice were immunized with peptides corresponding to the CD8(+) T-cell epitopes of a malaria antigen, a circumsporozoite protein of Plasmodium yoelii, and a tumor antigen, a Wilms Tumor antigen-1 (WT-1), together with 7DW8-5 and MPLA, as an adjuvant. These immunization regimens were able to induce higher levels of CD8(+) T-cell responses and, ultimately, enhanced levels of protection against malaria and tumor challenges compared to the levels induced by immunization with peptides mixed with 7DW8-5 or MPLA alone. Co-administration of 7DW8-5 and MPLA induces activation of memory-like effector natural killer T (NKT) cells, i.e. CD44(+)CD62L(-)NKT cells. Our study indicates that 7DW8-5 greatly enhances important synergistic pathways associated to memory immune responses when co-administered with MPLA, thus rendering this combination of adjuvants a novel vaccine adjuvant formulation. PMID:27132023

  19. Differential contributions of central and effector memory T cells to recall responses

    PubMed Central

    Roberts, Alan D.; Ely, Kenneth H.; Woodland, David L.

    2005-01-01

    Although the absolute number of memory CD8+ T cells established in the spleen following antigen encounter remains stable for many years, the relative capacity of these cells to mediate recall responses is not known. Here we used a dual adoptive transfer approach to demonstrate a progressive increase in the quality of memory T cell pools in terms of their ability to proliferate and accumulate at effector sites in response to secondary pathogen challenge. This temporal increase in efficacy occurred in CD62Llo (effector memory) and CD62Lhi (central memory) subpopulations, but was most prominent in the CD62Lhi subpopulation. These data indicate that the contribution of effector memory and central memory T cells to the recall response changes substantially over time. PMID:15983064

  20. Acute exercise mobilises CD8+ T lymphocytes exhibiting an effector-memory phenotype.

    PubMed

    Campbell, John P; Riddell, Natalie E; Burns, Victoria E; Turner, Mark; van Zanten, Jet J C S Veldhuijzen; Drayson, Mark T; Bosch, Jos A

    2009-08-01

    An acute bout of exercise evokes mobilisation of lymphocytes into the bloodstream, which can be largely attributed to increases in CD8+ T lymphocytes (CD8TLs) and natural killer (NK) cells. Evidence further suggests that, even within these lymphocyte subsets, there is preferential mobilisation of cells that share certain functional and phenotypic characteristics, such as high cytotoxicity, low proliferative ability, and high tissue-migrating potential. These features are characteristic of effector-memory CD8TL subsets. The current study therefore investigated the effect of exercise on these newly-identified subsets. Thirteen healthy and physically active males (mean+/-SD: age 20.9+/-1.5 yr) attended three sessions: a control session (no exercise); cycling at 35% Watt(max) (low intensity exercise); and 85% Watt(max) (high intensity exercise). Each bout lasted 20 min. Blood samples were obtained before exercise, during the final min of exercise, and +15, and +60 min post-exercise. CD8TLs were classified into naïve, central memory (CM), effector-memory (EM), and CD45RA+ effector-memory (RAEM) using combinations of the cell surface markers CCR7, CD27, CD62L, CD57, and CD45RA. In parallel, the phenotypically distinct CD56(bright) 'regulatory' and CD56(dim) 'cytotoxic' NK subsets were quantified. The results show a strong differential mobilisation of CD8TL subsets (RAEM>EM>CM>naïve); during high intensity exercise the greatest increase was observed for RAEM CD8Tls (+450%) and the smallest for naïve cells (+84%). Similarly, CD56(dim) NK cells (+995%) were mobilised to a greater extent than CD56(bright) (+153%) NK cells. In conclusion, memory CD8TL that exhibit a high effector and tissue-migrating potential are preferentially mobilised during exercise. This finding unifies a range of independent observations regarding exercise-induced phenotypic and functional changes in circulating lymphocytes. The selective mobilisation of cytotoxic tissue-migrating subsets, both

  1. Ultraviolet B suppresses immunity by inhibiting effector and memory T cells.

    PubMed

    Rana, Sabita; Byrne, Scott Napier; MacDonald, Linda Joanne; Chan, Carling Yan-Yan; Halliday, Gary Mark

    2008-04-01

    Contact hypersensitivity is a T-cell-mediated response to a hapten. Exposing C57BL/6 mice to UV B radiation systemically suppresses both primary and secondary contact hypersensitivity responses. The effects of UVB on in vivo T-cell responses during UVB-induced immunosuppression are unknown. We show here that UVB exposure, before contact sensitization, inhibits the expansion of effector CD4+ and CD8+ T cells in skin-draining lymph nodes and reduces the number of CD4+ and IFN-gamma+ CD8+ T cells infiltrating challenged ear skin. In the absence of UVB, at 10 weeks after initial hapten exposure, the ear skin of sensitized mice was infiltrated by dermal effector memory CD8+ T cells at the site of challenge. However, if mice were previously exposed to UVB, this cell population was absent, suggesting an impaired development of peripheral memory T cells. This finding occurred in the absence of UVB-induced regulatory CD4+ T cells and did not involve prostaglandin E2, suggesting that the importance of these two factors in mediating or initiating UVB-induced immunosuppression is dependent on UVB dose. Together these data indicate that in vivo T-cell responses are prone to immunoregulation by UVB, including a novel effect on both the activated T-cell pool size and the development of memory T cells in peripheral compartments. PMID:18292235

  2. Mcl-1 regulates effector and memory CD8 T-cell differentiation during acute viral infection.

    PubMed

    Kim, Eui Ho; Neldner, Brandon; Gui, Jingang; Craig, Ruth W; Suresh, M

    2016-03-01

    Mcl-1, an anti-apoptotic member of Bcl-2 family maintains cell viability during clonal expansion of CD8 T cells, but the cell intrinsic role of Mcl-1 in contraction of effectors or the number of memory CD8 T cells is unknown. Mcl-1 levels decline during the contraction phase but rebound to high levels in memory CD8 T cells. Therefore, by overexpressing Mcl-1 in CD8 T cells we asked whether limiting levels of Mcl-1 promote contraction of effectors and constrain CD8 T-cell memory. Mcl-1 overexpression failed to affect CD8 T-cell expansion, contraction or the magnitude of CD8 T-cell memory. Strikingly, high Mcl-1 levels enhanced mTOR phosphorylation and augmented the differentiation of terminal effector cells and effector memory CD8 T cells to the detriment of poly-cytokine-producing central memory CD8 T cells. Taken together, these findings provided unexpected insights into the role of Mcl-1 in the differentiation of effector and memory CD8 T cells.

  3. B7-H1 limits the entry of effector CD8+ T cells to the memory pool by upregulating Bim

    PubMed Central

    Gibbons, Rachel M.; Liu, Xin; Pulko, Vesna; Harrington, Susan M.; Krco, Christopher J.; Kwon, Eugene D.; Dong, Haidong

    2012-01-01

    Protective T‑cell immunity against cancer and infections is dependent on the generation of a durable effector and memory T‑cell pool. Studies from cancer and chronic infections reveal that B7-H1 (PD-L1) engagement with its receptor PD-1 promotes apoptosis of effector T cells. It is not clear how B7-H1 regulates T‑cell apoptosis and the subsequent impact of B7-H1 on the generation of memory T cells. In immunized B7-H1-deficient mice, we detected an increased expansion of effector CD8+ T cells and a delayed T‑cell contraction followed by the emergence of a protective CD8+ T‑cell memory capable of completely rejecting tumor metastases in the lung. Intracellular staining revealed that antigen-primed CD8+ T cells in B7-H1-deficient mice express lower levels of the pro-apoptotic molecule Bim. The engagement of activated CD8+ T cells by a plate-bound B7-H1 fusion protein led to the upregulation of Bim and increased cell death. Assays based on blocking antibodies determined that both PD-1 and CD80 are involved in the B7-H1-mediated regulation of Bim in activated CD8+ T cells. Our results suggest that B7-H1 may negatively regulate CD8+ T‑cell memory by enhancing the depletion of effector CD8+ T cells through the upregulation of Bim. Our findings may provide a new strategy for targeting B7-H1 signaling in effector CD8+ T cells to achieve protective antitumor memory responses. PMID:23170254

  4. CD4⁺ effector and memory cell populations protect against Cryptosporidium parvum infection.

    PubMed

    McNair, Nina N; Mead, Jan R

    2013-01-01

    Cryptosporidium parvum is a protozoan parasite that infects the epithelial cells of the small intestine causing diarrheal illness in humans. While T cells are known to be important in resistance and recovery from infection, little has been characterized as to the phenotypic expression of surface effector and memory markers after infection. We used an acute model of infection (C57BL/6 interleukin-12p40), which develops long-standing resistance to re-infection, to characterize expression of different effector and memory cells. Using flow cytometry, we found that heterogeneous populations were generated after infection, consisting of both CD62L(high) central memory T cells (T(CM)) and CD62L(low) effector memory T cells (T(EM)) that were competent to produce the Th type 1 effector cytokine, IFN-γ. Both CD4⁺ and CD8⁺ T(CM) and T(EM) populations persisted in the absence of infection (up to 60 days post-infection). Additionally, transfer of either CD62L(low)CD4⁺ T(EM) or CD62L(high)CD4⁺ T(CM) into naive recipients resulted in a protective response. Taken together, these studies show that distinct subsets of effector and memory CD4⁺ T cells develop after infection with C. parvum, and mediate protective immunity to re-challenge.

  5. Cutting edge: CCR7+ and CCR7- memory T cells do not differ in immediate effector cell function.

    PubMed

    Unsoeld, Heike; Krautwald, Stefan; Voehringer, David; Kunzendorf, Ulrich; Pircher, Hanspeter

    2002-07-15

    It has been proposed that expression of the chemokine receptor CCR7 represents a defining factor for nonpolarized central (CCR7(+)) and polarized effector memory (CCR7(-)) T cells. In this study, we have tested this hypothesis using in vivo-activated T cells from P14 and SMARTA TCR-transgenic (tg) mice specific for MHC class I- and II-restricted epitopes of the lymphocytic choriomeningitis virus (LCMV) glycoprotein. CCR7 cell surface expression on TCR-tg cells was monitored with a CC chemokine ligand 19-Ig fusion protein. CC chemokine ligand 19-Ig staining separated TCR-tg cells activated by LCMV infection into CCR7(-) and CCR7(+) effector/memory T cell populations. Nonetheless, both T cell populations isolated from spleen and liver produced identical amounts of IFN-gamma after short-term Ag stimulation. Furthermore, CCR7(+) and CCR7(-) CD8 TCR-tg cells from LCMV-infected mice exhibited similar lytic activity against LCMV peptide-coated target cells. These results question the proposed concept of differential effector cell function of CCR7(+) and CCR7(-) memory T cells. PMID:12097363

  6. OX40 costimulatory signals potentiate the memory commitment of effector CD8+ T cells.

    PubMed

    Mousavi, Seyed Fazlollah; Soroosh, Pejman; Takahashi, Takeshi; Yoshikai, Yasunobu; Shen, Hao; Lefrançois, Leo; Borst, Jannie; Sugamura, Kazuo; Ishii, Naoto

    2008-11-01

    A T cell costimulatory molecule, OX40, contributes to T cell expansion, survival, and cytokine production. Although several roles for OX40 in CD8(+) T cell responses to tumors and viral infection have been shown, the precise function of these signals in the generation of memory CD8(+) T cells remains to be elucidated. To address this, we examined the generation and maintenance of memory CD8(+) T cells during infection with Listeria monocytogenes in the presence and absence of OX40 signaling. We used the expression of killer cell lectin-like receptor G1 (KLRG1), a recently reported marker, to distinguish between short-lived effector and memory precursor effector T cells (MPECs). Although OX40 was dispensable for the generation of effector T cells in general, the lack of OX40 signals significantly reduced the number and proportion of KLRG1(low) MPECs, and, subsequently, markedly impaired the generation of memory CD8(+) T cells. Moreover, memory T cells that were generated in the absence of OX40 signals in a host animal did not show self-renewal in a second host, suggesting that OX40 is important for the maintenance of memory T cells. Additional experiments making use of an inhibitory mAb against the OX40 ligand demonstrated that OX40 signals are essential during priming, not only for the survival of KLRG1(low) MPECs, but also for their self-renewing ability, both of which contribute to the homeostasis of memory CD8(+) T cells.

  7. Neem leaf glycoprotein promotes dual generation of central and effector memory CD8(+) T cells against sarcoma antigen vaccine to induce protective anti-tumor immunity.

    PubMed

    Ghosh, Sarbari; Sarkar, Madhurima; Ghosh, Tithi; Guha, Ipsita; Bhuniya, Avishek; Saha, Akata; Dasgupta, Shayani; Barik, Subhasis; Bose, Anamika; Baral, Rathindranath

    2016-03-01

    We have previously shown that Neem Leaf Glycoprotein (NLGP) mediates sustained tumor protection by activating host immune response. Now we report that adjuvant help from NLGP predominantly generates CD44(+)CD62L(high)CCR7(high) central memory (TCM; in lymph node) and CD44(+)CD62L(low)CCR7(low) effector memory (TEM; in spleen) CD8(+) T cells of Swiss mice after vaccination with sarcoma antigen (SarAg). Generated TCM and TEM participated either to replenish memory cell pool for sustained disease free states or in rapid tumor eradication respectively. TCM generated after SarAg+NLGP vaccination underwent significant proliferation and IL-2 secretion following SarAg re-stimulation. Furthermore, SarAg+NLGP vaccination helps in greater survival of the memory precursor effector cells at the peak of the effector response and their maintenance as mature memory cells, in comparison to single modality treatment. Such response is corroborated with the reduced phosphorylation of FOXO in the cytosol and increased KLF2 in the nucleus associated with enhanced CD62L, CCR7 expression of lymph node-resident CD8(+) T cells. However, spleen-resident CD8(+) T memory cells show superior efficacy for immediate memory-to-effector cell conversion. The data support in all aspects that SarAg+NLGP demonstrate superiority than SarAg vaccination alone that benefits the host by rapid effector functions whenever required, whereas, central-memory cells are thought to replenish the memory cell pool for ultimate sustained disease free survival till 60 days following post-vaccination tumor inoculation.

  8. Effector memory and central memory NY-ESO-1-specific re-directed T cells for treatment of multiple myeloma.

    PubMed

    Schuberth, P C; Jakka, G; Jensen, S M; Wadle, A; Gautschi, F; Haley, D; Haile, S; Mischo, A; Held, G; Thiel, M; Tinguely, M; Bifulco, C B; Fox, B A; Renner, C; Petrausch, U

    2013-04-01

    The cancer-testis antigen NY-ESO-1 is a potential target antigen for immune therapy expressed in a subset of patients with multiple myeloma. We generated chimeric antigen receptors (CARs) recognizing the immunodominant NY-ESO-1 peptide 157-165 in the context of HLA-A*02:01 to re-direct autologous CD8(+) T cells towards NY-ESO-1(+) myeloma cells. These re-directed T cells specifically lysed NY-ESO-1(157-165)/HLA-A*02:01-positive cells and secreted IFNγ. A total of 40% of CCR7(-) re-directed T cells had an effector memory phenotype and 5% a central memory phenotype. Based on CCR7 cell sorting, effector and memory CAR-positive T cells were separated and CCR7(+) memory cells demonstrated after antigen-specific re-stimulation downregulation of CCR7 as sign of differentiation towards effector cells accompanied by an increased secretion of memory signature cytokines such as IL-2. To evaluate NY-ESO-1 as potential target antigen, we screened 78 bone marrow biopsies of multiple myeloma patients where NY-ESO-1 protein was found to be expressed by immunohistochemistry in 9.7% of samples. Adoptively transferred NY-ESO-1-specific re-directed T cells protected mice against challenge with endogenously NY-ESO-1-positive myeloma cells in a xenograft model. In conclusion, re-directed effector- and central memory T cells specifically recognized NY-ESO-1(157-165)/ HLA-A*02:01-positive cells resulting in antigen-specific functionality in vitro and in vivo.

  9. Effector-memory T cells develop in islets and report islet pathology in type 1 diabetes.

    PubMed

    Chee, Jonathan; Ko, Hyun-Ja; Skowera, Ania; Jhala, Gaurang; Catterall, Tara; Graham, Kate L; Sutherland, Robyn M; Thomas, Helen E; Lew, Andrew M; Peakman, Mark; Kay, Thomas W H; Krishnamurthy, Balasubramanian

    2014-01-15

    CD8(+) T cells are critical in human type 1 diabetes and in the NOD mouse. In this study, we elucidated the natural history of islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)-specific CD8(+) T cells in NOD diabetes using MHC-tetramer technology. IGRP206-214-specific T cells in the peripheral lymphoid tissue increased with age, and their numbers correlated with insulitis progression. IGRP206-214-specific T cells in the peripheral lymphoid tissue expressed markers of chronic Ag stimulation, and their numbers were stable after diagnosis of diabetes, consistent with their memory phenotype. IGRP206-214-specific T cells in NOD mice expand, acquire the phenotype of effector-memory T cells in the islets, and emigrate to the peripheral lymphoid tissue. Our observations suggest that enumeration of effector-memory T cells of multiple autoantigen specificities in the periphery of type 1 diabetic subjects could be a reliable reporter for progression of islet pathology.

  10. Current activities of the Yersinia effector protein YopM.

    PubMed

    Höfling, Sabrina; Grabowski, Benjamin; Norkowski, Stefanie; Schmidt, M Alexander; Rüter, Christian

    2015-05-01

    Yersinia outer protein M (YopM) belongs to the group of Yop effector proteins, which are highly conserved among pathogenic Yersinia species. During infection, the effectors are delivered into the host cell cytoplasm via the type 3 secretion system to subvert the host immune response and support the survival of Yersinia. In contrast to the other Yop effectors, YopM does not possess a known enzymatic activity and its molecular mechanism(s) of action remain(s) poorly understood. However, YopM was shown to promote colonization and dissemination of Yersinia, thus being crucial for the pathogen's virulence in vivo. Moreover, YopM interacts with several host cell proteins and might utilize them to execute its anti-inflammatory activities. The results obtained so far indicate that YopM is a multifunctional protein that counteracts the host immune defense by multiple activities, which are at least partially independent of each other. Finally, its functions seem to be also influenced by differences between the specific YopM isoforms expressed by Yersinia subspecies. In this review, we focus on the global as well as more specific contribution of YopM to virulence of Yersinia during infection and point out the various extra- and intracellular molecular functions of YopM. In addition, the novel cell-penetrating ability of recombinant YopM and its potential applications as a self-delivering immunomodulatory therapeutic will be discussed.

  11. Phenotypic Definition of Effector and Memory T-Lymphocyte Subsets in Mice Chronically Infected with Mycobacterium tuberculosis▿

    PubMed Central

    Henao-Tamayo, Marcela I.; Ordway, Diane J.; Irwin, Scott M.; Shang, Shaobin; Shanley, Crystal; Orme, Ian M.

    2010-01-01

    The bacterium Mycobacterium tuberculosis remains one of the world's most successful pathogens, a situation that is aggravated by the fact that the existing vaccine, Mycobacterium bovis BCG, is not effective in adults. As with any vaccine, the purpose of giving BCG vaccination is to establish a long-lived state of memory immunity, but whether this is successfully completely established is still unclear. It is generally accepted that memory T cells can be divided into central and effector memory populations by function and by phenotype; however, the majority of data supporting this division have been generated using transgenic mouse models or mice that have recovered from acute viral infections. Tuberculosis, on the other hand, represents a persistent, chronic state of immunity in which the presence of memory T cells is far less well defined. We show here that mice vaccinated with BCG or chronically infected with M. tuberculosis establish antigen-specific populations of cells within the lungs that predominantly express a cellular phenotype consistent with their being effector or effector memory cells. In contrast, cells with a central memory phenotype exist in much lower numbers in the lungs but can be found in significantly larger numbers in the spleen, where they may represent a potential reservoir. These data suggest that the effector-to-central-memory T-cell transition may well be minimal in these persisting mycobacterial infections, and they support a novel hypothesis that this may explain the fundamental basis of the failure of the BCG vaccine in humans. PMID:20107011

  12. Identification of Pertussis-Specific Effector Memory T Cells in Preschool Children

    PubMed Central

    Schure, Rose-Minke; Öztürk, Kemal; Berbers, Guy; Sanders, Elisabeth; van Twillert, Inonge; Carollo, Maria; Mascart, Françoise; Ausiello, Clara M.; van Els, Cecile A. C. M.; Smits, Kaat; Buisman, Anne-Marie

    2015-01-01

    Whooping cough remains a problem despite vaccination, and worldwide resurgence of pertussis is evident. Since cellular immunity plays a role in long-term protection against pertussis, we studied pertussis-specific T-cell responses. Around the time of the preschool acellular pertussis (aP) booster dose at 4 years of age, T-cell memory responses were compared in children who were primed during infancy with either a whole-cell pertussis (wP) or an aP vaccine. Peripheral blood mononuclear cells (PBMCs) were isolated and stimulated with pertussis vaccine antigens for 5 days. T cells were characterized by flow-based analysis of carboxyfluorescein succinimidyl ester (CFSE) dilution and CD4, CD3, CD45RA, CCR7, gamma interferon (IFN-γ), and tumor necrosis factor alpha (TNF-α) expression. Before the aP preschool booster vaccination, both the proliferated pertussis toxin (PT)-specific CD4+ and CD8+ T-cell fractions (CFSEdim) were higher in aP- than in wP-primed children. Post-booster vaccination, more pertussis-specific CD4+ effector memory cells (CD45RA− CCR7−) were induced in aP-primed children than in those primed with wP. The booster vaccination did not appear to significantly affect the T-cell memory subsets and functionality in aP-primed or wP-primed children. Although the percentages of Th1 cytokine-producing cells were alike in aP- and wP-primed children pre-booster vaccination, aP-primed children produced more Th1 cytokines due to higher numbers of proliferated pertussis-specific effector memory cells. At present, infant vaccinations with four aP vaccines in the first year of life result in pertussis-specific CD4+ and CD8+ effector memory T-cell responses that persist in children until 4 years of age and are higher than those in wP-primed children. The booster at 4 years of age is therefore questionable; this may be postponed to 6 years of age. PMID:25787136

  13. Activation of macrophages for destruction of Francisella tularensis: identification of cytokines, effector cells, and effector molecules.

    PubMed Central

    Fortier, A H; Polsinelli, T; Green, S J; Nacy, C A

    1992-01-01

    Francisella tularensis live vaccine strain (LVS) was grown in culture with nonadherent resident, starch-elicited, or Proteose Peptone-elicited peritoneal cells. Numbers of bacteria increased 4 logs over the input inoculum in 48 to 72 h. Growth rates were faster in inflammatory cells than in resident cells: generation times for the bacterium were 3 h in inflammatory cells and 6 h in resident macrophages. LVS-infected macrophage cultures treated with lymphokines did not support growth of the bacterium, although lymphokines alone had no inhibitory effects on replication of LVS in culture medium devoid of cells. Removal of gamma interferon (IFN-gamma) by immunoaffinity precipitation rendered lymphokines ineffective for induction of macrophage anti-LVS activity, and recombinant IFN-gamma stimulated both resident and inflammatory macrophage populations to inhibit LVS growth in vitro. Inflammatory macrophages were more sensitive to effects of IFN-gamma: half-maximal activity was achieved at 5 U/ml for inflammatory macrophages and 20 U/ml for resident macrophages. IFN-gamma-induced anti-LVS activity correlated with the production of nitrite (NO2-), an oxidative end product of L-arginine-derived nitric oxide (NO). Anti-LVS activity and nitrite production were both completely inhibited by the addition of either the L-arginine analog NG-monomethyl-L-arginine or anti-tumor necrosis factor antibodies to activated macrophage cultures. Thus, macrophages can be activated by IFN-gamma to suppress the growth of F. tularensis by generation of toxic levels of NO, and inflammatory macrophages are substantially more sensitive to activation activities of IFN-gamma for this effector reaction than are more differentiated resident cells. PMID:1541555

  14. Impact of CCR7 on priming and distribution of antiviral effector and memory CTL.

    PubMed

    Junt, Tobias; Scandella, Elke; Förster, Reinhold; Krebs, Philippe; Krautwald, Stefan; Lipp, Martin; Hengartner, Hans; Ludewig, Burkhard

    2004-12-01

    The chemokine receptor CCR7 is a key factor in the coordinate migration of T cells and dendritic cells (DC) into and their localization within secondary lymphoid organs. In this study we investigated the impact of CCR7 on CD8(+) T cell responses by infecting CCR7(-/-) mice with lymphocytic choriomeningitis virus (LCMV). We found that the absence of CCR7 affects the magnitude of an antiviral CTL response during the acute phase, with reduced numbers of virus-specific CTL in all lymphoid and nonlymphoid organs tested. On the single cell level, CCR7-deficient CTL gained full effector function, such that antiviral protection in CCR7-deficient mice was complete, but delayed. Similarly, adoptive transfer experiments using DC from CCR7-deficient or competent mice for the priming of CCR7-positive or CCR7-negative CD8(+) T cells, respectively, revealed that ectopic positioning of DC and CTL outside organized T cell zones results in reduced priming efficacy. In the memory phase, CCR7-deficient mice maintained a stable LCMV-specific CTL population, predominantly in nonlymphoid organs, and rapidly mounted protective CTL responses against a challenge infection with a vaccinia virus recombinant for the gp33 epitope of LCMV. Taken together, the CCR7-dependent organization of the T cell zone does not appear to be a prerequisite for antiviral effector CTL differentiation and the sustenance of antiviral memory responses in lymphoid or peripheral tissues. PMID:15557160

  15. Serine protease inhibitor-6 differentially affects the survival of effector and memory alloreactive CD8-T cells.

    PubMed

    Azzi, J; Ohori, S; Ting, C; Uehara, M; Abdoli, R; Smith, B D; Safa, K; Solhjou, Z; Lukyanchykov, P; Patel, J; McGrath, M; Abdi, R

    2015-01-01

    The clonal expansion of effector T cells and subsequent generation of memory T cells are critical in determining the outcome of transplantation. While cytotoxic T lymphocytes induce direct cytolysis of target cells through secretion of Granzyme-B (GrB), they also express cytoplasmic serine protease inhibitor-6 (Spi6) to protect themselves from GrB that has leaked from granules. Here, we studied the role of GrB/Spi6 axis in determining clonal expansion of alloreactive CD8-T cells and subsequent generation of memory CD8-T cells in transplantation. CD8-T cells from Spi6(-/-) mice underwent more GrB mediated apoptosis upon alloantigen stimulation in vitro and in vivo following adoptive transfer into an allogeneic host. Interestingly, while OT1.Spi6(-/-) CD8 T cells showed significantly lower clonal expansion following skin transplants from OVA mice, there was no difference in the size of the effector memory CD8-T cells long after transplantation. Furthermore, lack of Spi6 resulted in a decrease of short-lived-effector-CD8-cells but did not impact the pool of memory-precursor-effector-CD8-cells. Similar results were found in heart transplant models. Our findings suggest that the final alloreactive CD8-memory-pool-size is independent from the initial clonal-proliferation as memory precursors express low levels of GrB and therefore are independent of Spi6 for survival. These data advance our understanding of memory T cells generation in transplantation and provide basis for Spi6 based strategies to target effector T cells.

  16. Serine Protease Inhibitor-6 Differentially Affects the Survival of Effector and Memory Alloreactive CD8-T Cells

    PubMed Central

    Azzi, J.; Ohori, S.; Ting, C.; Uehara, M.; Abdoli, R.; Smith, B. D.; Safa, K.; Solhjou, Z.; Lukyanchykov, P.; Patel, J.; McGrath, M.; Abdi, R.

    2016-01-01

    The clonal expansion of effector T cells and subsequent generation of memory T cells are critical in determining the outcome of transplantation. While cytotoxic T lymphocytes induce direct cytolysis of target cells through secretion of Granzyme-B (GrB), they also express cytoplasmic serine protease inhibitor-6 (Spi6) to protect themselves from GrB that has leaked from granules. Here, we studied the role of GrB/Spi6 axis in determining clonal expansion of alloreactive CD8-T cells and subsequent generation of memory CD8-T cells in transplantation. CD8-T cells from Spi6−/− mice underwent more GrB mediated apoptosis upon alloantigen stimulation in vitro and in vivo following adoptive transfer into an allogeneic host. Interestingly, while OT1.Spi6−/− CD8 T cells showed significantly lower clonal expansion following skin transplants from OVA mice, there was no difference in the size of the effector memory CD8-T cells long after transplantation. Furthermore, lack of Spi6 resulted in a decrease of short-lived-effector-CD8-cells but did not impact the pool of memory-precursor-effector-CD8-cells. Similar results were found in heart transplant models. Our findings suggest that the final alloreactive CD8-memory-pool-size is independent from the initial clonal-proliferation as memory precursors express low levels of GrB and therefore are independent of Spi6 for survival. These data advance our understanding of memory T cells generation in transplantation and provide basis for Spi6 based strategies to target effector T cells. PMID:25534448

  17. Differential impact of CD27 and 4-1BB costimulation on effector and memory CD8 T cell generation following peptide immunization.

    PubMed

    Willoughby, Jane E; Kerr, Jonathan P; Rogel, Anne; Taraban, Vadim Y; Buchan, Sarah L; Johnson, Peter W M; Al-Shamkhani, Aymen

    2014-07-01

    The factors that determine differentiation of naive CD8 T cells into memory cells are not well understood. A greater understanding of how memory cells are generated will inform of ways to improve vaccination strategies. In this study, we analyzed the CD8 T cell response elicited by two experimental vaccines comprising a peptide/protein Ag and an agonist that delivers a costimulatory signal via CD27 or 4-1BB. Both agonists increased expansion of Ag-specific CD8 T cells compared with Ag alone. However, their capacity to stimulate differentiation into effector and memory cells differed. CD27 agonists promoted increased expression of perforin and the generation of short-lived memory cells, whereas stimulation with 4-1BB agonists favored generation of stable memory. The memory-promoting effects of 4-1BB were independent of CD4 T cells and were the result of programing within the first 2 d of priming. Consistent with this conclusion, CD27 and 4-1BB-stimulated CD8 T cells expressed disparate amounts of IL-2, IFN-γ, CD25, CD71, and Gp49b as early as 3 d after in vivo activation. In addition, memory CD8 T cells, generated through priming with CD27 agonists, proliferated more extensively than did 4-1BB-generated memory cells, but these cells failed to persist. These data demonstrate a previously unanticipated link between the rates of homeostatic proliferation and memory cell attrition. Our study highlights a role for these receptors in skewing CD8 T cell differentiation into effector and memory cells and provides an approach to optimize vaccines that elicit CD8 T cell responses.

  18. Transcription Factor Bcl11b Controls Effector and Memory CD8 T cell Fate Decision and Function during Poxvirus Infection

    PubMed Central

    Abboud, Georges; Stanfield, Jessica; Tahiliani, Vikas; Desai, Pritesh; Hutchinson, Tarun E.; Lorentsen, Kyle J.; Cho, Jonathan J.; Avram, Dorina; Salek-Ardakani, Shahram

    2016-01-01

    CD8+ T cells play an important role in host resistance to many viral infections, but the underlying transcriptional mechanisms governing their differentiation and functionality remain poorly defined. By using a highly virulent systemic and respiratory poxvirus infection in mice, we show that the transcription factor Bcl11b provides a dual trigger that sustains the clonal expansion of virus-specific effector CD8+ T cells, while simultaneously suppressing the expression of surface markers associated with short-lived effector cell (SLEC) differentiation. Additionally, we demonstrate that Bcl11b supports the acquisition of memory precursor effector cell (MPEC) phenotype and, thus, its absence causes near complete loss of lymphoid and lung-resident memory cells. Interestingly, despite having normal levels of T-bet and Eomesodermin, Bcl11b-deficient CD8+ T cells failed to execute effector differentiation needed for anti-viral cytokine production and degranulation, suggesting a non-redundant role of Bcl11b in regulation of this program. Thus, Bcl11b is a critical player in fate decision of SLECs and MPECs, as well as effector function and memory formation. PMID:27790219

  19. Airway inflammation and IgE production induced by dust mite allergen-specific memory/effector Th2 cell line can be effectively attenuated by IL-35.

    PubMed

    Huang, Chiung-Hui; Loo, Evelyn Xiu-Ling; Kuo, I-Chun; Soh, Gim Hooi; Goh, Denise Li-Meng; Lee, Bee Wah; Chua, Kaw Yan

    2011-07-01

    CD4(+) memory/effector T cells play a central role in orchestrating the rapid and robust immune responses upon re-encounter with specific Ags. However, the immunologic mechanism(s) underlying these responses are still not fully understood. To investigate this, we generated an allergen (major house dust mite allergen, Blo t 5)-specific murine Th2 cell line that secreted IL-4, IL-5, IL-10, and IL-13, but not IL-9 or TNF-α, upon activation by the cognate Ag. These cells also exhibited CD44(high)CD62L(-) and CD127(+) (IL-7Rα(+)) phenotypes, which are characteristics of memory/effector T cells. Experiments involving adoptive transfer of this Th2 cell line in mice, followed by three intranasal challenges with Blo t 5, induced a dexamethasone-sensitive eosinophilic airway inflammation. This was accompanied by elevation of Th2 cytokines and CC- and CXC-motif chemokines, as well as recruitment of lymphocytes and polymorphic mononuclear cells into the lungs. Moreover, Blo t 5-specific IgE was detected 4 d after the last intranasal challenge, whereas elevation of Blo t 5-specific IgG1 was found at week two. Finally, pulmonary delivery of the pVAX-IL-35 DNA construct effectively downregulated Blo t 5-specific allergic airway inflammation, and i.m. injection of pVAX-IL-35 led to long-lasting suppression of circulating Blo t 5-specific and total IgE. This model provides a robust research tool to elucidate the immunopathogenic role of memory/effector Th2 cells in allergic airway inflammation. Our results suggested that IL-35 could be a potential therapeutic target for allergic asthma through its attenuating effects on allergen-specific CD4(+) memory/effector Th2 cell-mediated airway inflammation.

  20. Innate signals compensate for the absence of PKC-{theta} during in vivo CD8(+) T cell effector and memory responses.

    PubMed

    Marsland, Benjamin J; Nembrini, Chiara; Schmitz, Nicole; Abel, Brian; Krautwald, Stefan; Bachmann, Martin F; Kopf, Manfred

    2005-10-01

    PKC- is central to T-helper (Th) 2 cell differentiation and effector function; however, its importance for antiviral effector, and in particular memory CD8(+) T cell responses, remains unclear. We have investigated the role of PKC- during in vivo and in vitro responses against influenza virus, lymphocytic choriomeningitis virus, vaccinia virus, and replication-deficient virus-like particles. In the absence of PKC-, antiviral CD8(+) T cells presented an unresponsive phenotype in vitro, which could be restored with exogenous IL-2 or by Toll-like receptor ligand-activated dendritic cells. In striking contrast, PKC- appeared to be superfluous for in vivo antiviral responses irrespective of whether the virus infected systemically, was localized to the lung, or did not replicate. In addition, CD8(+) CCR7-effector memory responses were normal in PKC--deficient mice, both in lymphoid and peripheral tissues. Our data show that increased activation signals delivered in vivo by highly activated dendritic cells, as present during viral infections, overcome the requirement for PKC- during CD8(+) T cell antiviral responses. PMID:16186501

  1. Innate signals compensate for the absence of PKC-θ during in vivo CD8+ T cell effector and memory responses

    PubMed Central

    Marsland, Benjamin J.; Nembrini, Chiara; Schmitz, Nicole; Abel, Brian; Krautwald, Stefan; Bachmann, Martin F.; Kopf, Manfred

    2005-01-01

    PKC-θ is central to T-helper (Th) 2 cell differentiation and effector function; however, its importance for antiviral effector, and in particular memory CD8+ T cell responses, remains unclear. We have investigated the role of PKC-θ during in vivo and in vitro responses against influenza virus, lymphocytic choriomeningitis virus, vaccinia virus, and replication-deficient virus-like particles. In the absence of PKC-θ, antiviral CD8+ T cells presented an unresponsive phenotype in vitro, which could be restored with exogenous IL-2 or by Toll-like receptor ligand-activated dendritic cells. In striking contrast, PKC-θ appeared to be superfluous for in vivo antiviral responses irrespective of whether the virus infected systemically, was localized to the lung, or did not replicate. In addition, CD8+ CCR7-effector memory responses were normal in PKC-θ-deficient mice, both in lymphoid and peripheral tissues. Our data show that increased activation signals delivered in vivo by highly activated dendritic cells, as present during viral infections, overcome the requirement for PKC-θ during CD8+ T cell antiviral responses. PMID:16186501

  2. Chemokine receptor CXCR3 facilitates CD8+ T cell differentiation into short-lived effector cells leading to memory degeneration

    PubMed Central

    Kurachi, Makoto; Kurachi, Junko; Suenaga, Fumiko; Tsukui, Tatsuya; Abe, Jun; Ueha, Satoshi; Tomura, Michio; Sugihara, Kei; Takamura, Shiki; Kakimi, Kazuhiro

    2011-01-01

    Strength of inflammatory stimuli during the early expansion phase plays a crucial role in the effector versus memory cell fate decision of CD8+ T cells. But it is not known how early lymphocyte distribution after infection has an impact on this process. We demonstrate that the chemokine receptor CXCR3 is involved in promoting CD8+ T cell commitment to an effector fate rather than a memory fate by regulating T cell recruitment to an antigen/inflammation site. After systemic viral or bacterial infection, the contraction of CXCR3−/− antigen-specific CD8+ T cells is significantly attenuated, resulting in massive accumulation of fully functional memory CD8+ T cells. Early after infection, CXCR3−/− antigen-specific CD8+ T cells fail to cluster at the marginal zone in the spleen where inflammatory cytokines such as IL-12 and IFN-α are abundant, thus receiving relatively weak inflammatory stimuli. Consequently, CXCR3−/− CD8+ T cells exhibit transient expression of CD25 and preferentially differentiate into memory precursor effector cells as compared with wild-type CD8+ T cells. This series of events has important implications for development of vaccination strategies to generate increased numbers of antigen-specific memory CD8+ T cells via inhibition of CXCR3-mediated T cell migration to inflamed microenvironments. PMID:21788406

  3. [Transcription activator-like effectors(TALEs)based genome engineering].

    PubMed

    Zhao, Mei-Wei; Duan, Cheng-Li; Liu, Jiang

    2013-10-01

    Systematic reverse-engineering of functional genome architecture requires precise modifications of gene sequences and transcription levels. The development and application of transcription activator-like effectors(TALEs) has created a wealth of genome engineering possibilities. TALEs are a class of naturally occurring DNA-binding proteins found in the plant pathogen Xanthomonas species. The DNA-binding domain of each TALE typically consists of tandem 34-amino acid repeat modules rearranged according to a simple cipher to target new DNA sequences. Customized TALEs can be used for a wide variety of genome engineering applications, including transcriptional modulation and genome editing. Such "genome engineering" has now been established in human cells and a number of model organisms, thus opening the door to better understanding gene function in model organisms, improving traits in crop plants and treating human genetic disorders.

  4. Unexpected positive control of NFκB and miR-155 by DGKα and ζ ensures effector and memory CD8+ T cell differentiation

    PubMed Central

    Yang, Jialong; Zhang, Ping; Krishna, Sruti; Wang, Jinli; Lin, Xingguang; Huang, Hongxiang; Xie, Danli; Gorentla, Balachandra; Huang, Rick; Gao, Jimin; Li, Qi-Jing; Zhong, Xiao-Ping

    2016-01-01

    Signals from the T-cell receptor (TCR) and γ-chain cytokine receptors play crucial roles in initiating activation and effector/memory differentiation of CD8 T-cells. We report here that simultaneous deletion of both diacylglycerol kinase (DGK) α and ζ (DKO) severely impaired expansion of CD8 effector T cells and formation of memory CD8 T-cells after Listeria monocytogenes infection. Moreover, ablation of both DGKα and ζ in preformed memory CD8 T-cells triggered death and impaired homeostatic proliferation of these cells. DKO CD8 T-cells were impaired in priming due to decreased expression of chemokine receptors and migration to the draining lymph nodes. Moreover, DKO CD8 T-cells were unexpectedly defective in NFκB-mediated miR-155 transcript, leading to excessive SOCS1 expression and impaired γ-chain cytokine signaling. Our data identified a DGK-NFκB-miR-155-SOCS1 axis that bridges TCR and γ-chain cytokine signaling for robust CD8 T-cell primary and memory responses to bacterial infection. PMID:27014906

  5. Exosomes: novel effectors of human platelet lysate activity.

    PubMed

    Torreggiani, E; Perut, F; Roncuzzi, L; Zini, N; Baglìo, S R; Baldini, N

    2014-01-01

    Despite the popularity of platelet-rich plasma (PRP) and platelet lysate (PL) in orthopaedic practice, the mechanism of action and the effectiveness of these therapeutic tools are still controversial. So far, the activity of PRP and PL has been associated with different growth factors (GF) released during platelet degranulation. This study, for the first time, identifies exosomes, nanosized vesicles released in the extracellular compartment by a number of elements, including platelets, as one of the effectors of PL activity. Exosomes were isolated from human PL by differential ultracentrifugation, and analysed by electron microscopy and Western blotting. Bone marrow stromal cells (MSC) treated with three different exosome concentrations (0.6 μg, 5 μg and 50 μg) showed a significant, dose-dependent increase in cell proliferation and migration compared to the control. In addition, osteogenic differentiation assays demonstrated that exosome concentration differently affected the ability of MSC to deposit mineralised matrix. Finally, the analysis of exosome protein content revealed a higher amount of basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF-BB) and transforming growth factor beta 1 (TGF-β1) as compared to PL. In regards to RNA content, an enrichment of small RNAs in exosomes as compared to donor platelets has been found. These results suggest that exosomes consistently contribute to PL activity and could represent an advantageous nanodelivery system for cell-free regeneration therapies. PMID:25241964

  6. Effector, Memory, and Dysfunctional CD8+ T Cell Fates in the Antitumor Immune Response

    PubMed Central

    2016-01-01

    The adaptive immune system plays a pivotal role in the host's ability to mount an effective, antigen-specific immune response against tumors. CD8+ tumor-infiltrating lymphocytes (TILs) mediate tumor rejection through recognition of tumor antigens and direct killing of transformed cells. In growing tumors, TILs are often functionally impaired as a result of interaction with, or signals from, transformed cells and the tumor microenvironment. These interactions and signals can lead to transcriptional, functional, and phenotypic changes in TILs that diminish the host's ability to eradicate the tumor. In addition to effector and memory CD8+ T cells, populations described as exhausted, anergic, senescent, and regulatory CD8+ T cells have been observed in clinical and basic studies of antitumor immune responses. In the context of antitumor immunity, these CD8+ T cell subsets remain poorly characterized in terms of fate-specific biomarkers and transcription factor profiles. Here we discuss the current characterization of CD8+ T cell fates in antitumor immune responses and discuss recent insights into how signals in the tumor microenvironment influence TIL transcriptional networks to promote CD8+ T cell dysfunction. PMID:27314056

  7. Prostaglandin D2-loaded microspheres effectively activate macrophage effector functions.

    PubMed

    Pereira, Priscilla Aparecida Tartari; Bitencourt, Claudia da Silva; dos Santos, Daiane Fernanda; Nicolete, Roberto; Gelfuso, Guilherme Martins; Faccioli, Lúcia Helena

    2015-10-12

    Biodegradable lactic-co-glycolic acid (PLGA) microspheres (MS) improve the stability of biomolecules stability and allow enable their sustained release. Lipid mediators represent a strategy for improving host defense; however, most of these mediators, such as prostaglandin D2 (PGD2), have low water solubility and are unstable. The present study aimed to develop and characterize MS loaded with PGD2 (PGD2-MS) to obtain an innovative tool to activate macrophages. PGD2-MS were prepared using an oil-in-water emulsion solvent extraction-evaporation process, and the size, zeta potential, surface morphology and encapsulation efficiency were determined. It was also evaluated in vitro the phagocytic index, NF-κB activation, as well as nitric oxide and cytokine production by alveolar macrophages (AMs) in response to PGD2-MS. PGD2-MS were spherical with a diameter of 5.0±3.3 μm and regular surface, zeta potential of -13.4±5.6 mV, and 36% of encapsulation efficiency, with 16-26% release of entrapped PGD2 at 4 and 48 h, respectively. PGD2-MS were more efficiently internalized by AMs than unloaded-MS, and activated NF-κB more than free PGD2. Moreover, PGD2-MS stimulated the production of nitric oxide, TNF-α, IL-1β, and TGF-β, more than free PGD2, indicating that microencapsulation increased the activating effect of PGD2 on cells. In LPS-pre-treated AMs, PGD2-MS decreased the release of IL-6 but increased the production of nitric oxide and IL-1β. These results show that the morphological characteristics of PGD2-MS facilitated interaction with, and activation of phagocytic cells; moreover, PGD2-MS retained the biological activities of PGD2 to trigger effector mechanisms in AMs. It is suggested that PGD2-MS represent a strategy for therapeutic intervention in the lungs of immunocompromised subjects.

  8. Memory CD4+ T cells are required for optimal NK cell effector functions against the opportunistic fungal pathogen Pneumocystis murina.

    PubMed

    Kelly, Michelle N; Zheng, Mingquan; Ruan, Sanbao; Kolls, Jay; D'Souza, Alain; Shellito, Judd E

    2013-01-01

    Little is known about the role of NK cells or their interplay with other immune cells during opportunistic infections. Using our murine model of Pneumocystis pneumonia, we found that loss of NK cells during immunosuppression results in substantial Pneumocystis lung burden. During early infection of C57B/6 CD4(+) T cell-depleted mice, there were significantly fewer NK cells in the lung tissue compared with CD4(+) T cell-intact animals, and the NK cells present demonstrated decreased upregulation of the activation marker NKp46 and production of the effector cytokine, IFN-γ. Furthermore, coincubation studies revealed a significant increase in fungal killing when NK cells were combined with CD4(+) T cells compared with either cell alone, which was coincident with a significant increase in perforin production by NK cells. Finally, however, we found through adoptive transfer that memory CD4(+) T cells are required for significant NK cell upregulation of the activation marker NK group 2D and production of IFN-γ, granzyme B, and perforin during Pneumocystis infection. To the best of our knowledge, this study is the first to demonstrate a role for NK cells in immunity to Pneumocystis pneumonia, as well as to establish a functional relationship between CD4(+) T cells and NK cells in the host response to an opportunistic fungal pathogen.

  9. [Advances in transcription activator-like effectors--a review].

    PubMed

    Yu, Tang; Li, Lisha; Lin, Jun

    2015-07-01

    As a protein originally found in plant pathogenic bacteria, transcription activator-like effectors (TALEs) can be fused with the cleaving domain of restriction endonuclease (For example Fok I) to form artificial nucleases named TALENs. These proteins are dependent on variable numbers of tandem Repeats of TALEs to recognize and bind DNA sequences. Each of these repeats consists of a set of approximately 34 amino acids, composed of about 32 conserved amino acids and 2 highly variable amino acids called repeat variant di-residues (RVDs). RVDs distinguish one TALE from another and can make TALEs have a simple cipher for the one-to-one recognition for proteins and DNA bases. Based on this, in theory, artificially constructed TALENs could recognize and break DNA sites specifically and arbitrarily to perform gene knockout, insertion or modification. We reviewed the development of this technology in multi-level and multi species, and its advantages and disadvantages compared with ZFNs and CRISPR/Cas technology. We also address its special advantages in industrial microbe breeding, vector construction, targeting precision, high efficiency of editing and biological safety. PMID:26647578

  10. Epidermal Fatty Acid Binding Protein (E-FABP) Is Not Required for the Generation or Maintenance of Effector and Memory T Cells following Infection with Listeria monocytogenes.

    PubMed

    Li, Bing; Schmidt, Nathan W

    2016-01-01

    Following activation of naïve T cells there are dynamic changes in the metabolic pathways used by T cells to support both the energetic needs of the cell and the macromolecules required for growth and proliferation. Among other changes, lipid metabolism undergoes dynamic transitions between fatty acid oxidation and fatty acid synthesis as cells progress from naïve to effector and effector to memory T cells. The hydrophobic nature of lipids requires that they be bound to protein chaperones within a cell. Fatty acid binding proteins (FABPs) represent a large class of lipid chaperones, with epidermal FABP (E-FABP) expressed in T cells. The objective of this study was to determine the contribution of E-FABP in antigen-specific T cell responses. Following infection with Listeria monocytogenes, we observed similar clonal expansion, contraction and formation of memory CD8 T cells in WT and E-FABP-/- mice, which also exhibited similar phenotypic and functional characteristics. Analysis of Listeria-specific CD4 T cells also revealed no defect in the expansion, contraction, and formation of memory CD4 T cells in E-FABP-/- mice. These data demonstrate that E-FABP is dispensable for antigen-specific T cell responses following a bacterial infection. PMID:27588422

  11. Epidermal Fatty Acid Binding Protein (E-FABP) Is Not Required for the Generation or Maintenance of Effector and Memory T Cells following Infection with Listeria monocytogenes

    PubMed Central

    Li, Bing; Schmidt, Nathan W.

    2016-01-01

    Following activation of naïve T cells there are dynamic changes in the metabolic pathways used by T cells to support both the energetic needs of the cell and the macromolecules required for growth and proliferation. Among other changes, lipid metabolism undergoes dynamic transitions between fatty acid oxidation and fatty acid synthesis as cells progress from naïve to effector and effector to memory T cells. The hydrophobic nature of lipids requires that they be bound to protein chaperones within a cell. Fatty acid binding proteins (FABPs) represent a large class of lipid chaperones, with epidermal FABP (E-FABP) expressed in T cells. The objective of this study was to determine the contribution of E-FABP in antigen-specific T cell responses. Following infection with Listeria monocytogenes, we observed similar clonal expansion, contraction and formation of memory CD8 T cells in WT and E-FABP-/- mice, which also exhibited similar phenotypic and functional characteristics. Analysis of Listeria-specific CD4 T cells also revealed no defect in the expansion, contraction, and formation of memory CD4 T cells in E-FABP-/- mice. These data demonstrate that E-FABP is dispensable for antigen-specific T cell responses following a bacterial infection. PMID:27588422

  12. Polarized granzyme release is required for antigen-driven transendothelial migration of human effector memory CD4 T cells

    PubMed Central

    Manes, Thomas D.; Pober, Jordan S.

    2014-01-01

    Human effector memory (EM) CD4 T cells may transmigrate across endothelial cell (EC) monolayers either in response to inflammatory chemokines or in response to TCR recognition of antigen presented on the surface of the EC. The kinetics, morphologic manifestations, and molecular requirements of chemokine- and TCR-driven transendothelial migration (TEM) differ significantly. Here we report that while the MTOC and cytosolic granules follow the nucleus across the endothelium in a uropod during chemokine-driven TEM, MTOC reorientation to the contact region between the T cell and the EC, accompanied by dynein-driven transport of granzyme-containing granules to and exocytosis at the contact region, are early events in TCR-driven but not chemokine-driven TEM. Inhibitors of either granule function or of granzyme proteolytic activity can arrest TCR-driven TEM, implying a requirement for granule discharge in the process. In the final stages of TCR-driven TEM, the MTOC precedes, rather than follows, the nucleus across the endothelium. Thus TCR-driven TEM of EM CD4 T cells appears to be a novel process that more closely resembles immune synapse formation than it does conventional chemotaxis. PMID:25367116

  13. Parallel Profiles of Inflammatory and Effector Memory T Cells in Visceral Fat and Liver of Obesity-Associated Cancer Patients.

    PubMed

    Conroy, Melissa J; Galvin, Karen C; Doyle, Suzanne L; Kavanagh, Maria E; Mongan, Ann-Marie; Cannon, Aoife; Moore, Gillian Y; Reynolds, John V; Lysaght, Joanne

    2016-10-01

    In the midst of a worsening obesity epidemic, the incidence of obesity-associated morbidities, including cancer, diabetes, cardiac and liver disease is increasing. Insights into mechanisms underlying pathological obesity-associated inflammation are lacking. Both the omentum, the principal component of visceral fat, and liver of obese individuals are sites of excessive inflammation, but to date the T cell profiles of both compartments have not been assessed or compared in a patient cohort with obesity-associated disease. We have previously identified that omentum is enriched with inflammatory cytokines, chemokines and T cells. Here, we compared the inflammatory profile of T cells in the omentum and liver of patients with the obesity-associated malignancy oesophageal adenocarcinoma (OAC). Furthermore, we assessed the secreted cytokine profile in OAC patient serum, omentum and liver to assess systemic and local inflammation. We observed parallel T cell cytokine profiles and phenotypes in the omentum and liver of OAC patients, in particular CD69(+) and inflammatory effector memory T cells. This study reflects similar processes of inflammation and T cell activation in the omentum and liver, and may suggest common targets to modulate pathological inflammation at these sites.

  14. Expanded CD8+ T cells of murine and human CLL are driven into a senescent KLRG1+ effector memory phenotype.

    PubMed

    Göthert, Joachim Rudolf; Eisele, Lewin; Klein-Hitpass, Ludger; Weber, Stefanie; Zesewitz, Marie-Louise; Sellmann, Ludger; Röth, Alexander; Pircher, Hanspeter; Dührsen, Ulrich; Dürig, Jan

    2013-11-01

    Altered numbers and functions of T cells have previously been demonstrated in chronic lymphocytic leukemia (CLL) patients. However, dynamics and specific T-cell subset alterations have not been studied in great detail. Therefore, we studied CLL blood lymphocyte subsets of individual patients in a longitudinal manner. Dynamic expansions of blood CD4 + and CD8 + T-cell numbers were consistently associated with a progressively increasing CLL leukemic compartment. Interestingly, the T-cell subset expansion over time was more pronounced in CD38 + CLL. Additionally, we performed gene expression profiling of CD3 + T cells of CLL patients and normal donors. Using gene set enrichment analysis, we found significant enrichment of genes with higher expression in CLL T cells within CD8+ effector memory and terminal effector T-cell gene signatures. In agreement with these data, we observed a marked expansion of phenotypic CD8 + effector memory T cells in CLL by flow cytometry. Moreover, we observed that increments of CD8 + effector memory T cells in human CLL and also mouse CLL (Eμ-TCL1 model) were due to an expansion of the inhibitory killer cell lectin-like receptor G1 (KLRG1) expressing cellular subset. Furthermore, higher plasma levels of the natural KLRG1 ligand E-cadherin were detected in CLL patients compared to normal donor controls. The predominance of KLRG1+ expression within CD8+ T cells in conjunction with increased systemic soluble E-cadherin might significantly contribute to CLL immune dysfunction and might additionally represent an important component of the CLL microenvironment.

  15. Application of Long-term cultured Interferon-γ Enzyme-linked Immunospot Assay for Assessing Effector and Memory T Cell Responses in Cattle.

    PubMed

    Maggioli, Mayara F; Palmer, Mitchell V; Vordermeier, H Martin; Whelan, Adam O; Fosse, James M; Nonnecke, Brian J; Waters, W Ray

    2015-01-01

    Effector and memory T cells are generated through developmental programing of naïve cells following antigen recognition. If the infection is controlled up to 95 % of the T cells generated during the expansion phase are eliminated (i.e., contraction phase) and memory T cells remain, sometimes for a lifetime. In humans, two functionally distinct subsets of memory T cells have been described based on the expression of lymph node homing receptors. Central memory T cells express C-C chemokine receptor 7 and CD45RO and are mainly located in T-cell areas of secondary lymphoid organs. Effector memory T cells express CD45RO, lack CCR7 and display receptors associated with lymphocyte homing to peripheral or inflamed tissues. Effector T cells do not express either CCR7 or CD45RO but upon encounter with antigen produce effector cytokines, such as interferon-γ. Interferon-γ release assays are used for the diagnosis of bovine and human tuberculosis and detect primarily effector and effector memory T cell responses. Central memory T cell responses by CD4(+) T cells to vaccination, on the other hand, may be used to predict vaccine efficacy, as demonstrated with simian immunodeficiency virus infection of non-human primates, tuberculosis in mice, and malaria in humans. Several studies with mice and humans as well as unpublished data on cattle, have demonstrated that interferon-γ ELISPOT assays measure central memory T cell responses. With this assay, peripheral blood mononuclear cells are cultured in decreasing concentration of antigen for 10 to 14 days (long-term culture), allowing effector responses to peak and wane; facilitating central memory T cells to differentiate and expand within the culture. PMID:26275095

  16. Application of Long-term cultured Interferon-γ Enzyme-linked Immunospot Assay for Assessing Effector and Memory T Cell Responses in Cattle.

    PubMed

    Maggioli, Mayara F; Palmer, Mitchell V; Vordermeier, H Martin; Whelan, Adam O; Fosse, James M; Nonnecke, Brian J; Waters, W Ray

    2015-07-11

    Effector and memory T cells are generated through developmental programing of naïve cells following antigen recognition. If the infection is controlled up to 95 % of the T cells generated during the expansion phase are eliminated (i.e., contraction phase) and memory T cells remain, sometimes for a lifetime. In humans, two functionally distinct subsets of memory T cells have been described based on the expression of lymph node homing receptors. Central memory T cells express C-C chemokine receptor 7 and CD45RO and are mainly located in T-cell areas of secondary lymphoid organs. Effector memory T cells express CD45RO, lack CCR7 and display receptors associated with lymphocyte homing to peripheral or inflamed tissues. Effector T cells do not express either CCR7 or CD45RO but upon encounter with antigen produce effector cytokines, such as interferon-γ. Interferon-γ release assays are used for the diagnosis of bovine and human tuberculosis and detect primarily effector and effector memory T cell responses. Central memory T cell responses by CD4(+) T cells to vaccination, on the other hand, may be used to predict vaccine efficacy, as demonstrated with simian immunodeficiency virus infection of non-human primates, tuberculosis in mice, and malaria in humans. Several studies with mice and humans as well as unpublished data on cattle, have demonstrated that interferon-γ ELISPOT assays measure central memory T cell responses. With this assay, peripheral blood mononuclear cells are cultured in decreasing concentration of antigen for 10 to 14 days (long-term culture), allowing effector responses to peak and wane; facilitating central memory T cells to differentiate and expand within the culture.

  17. Distinct dendritic cell subsets dictate the fate decision between effector and memory CD8(+) T cell differentiation by a CD24-dependent mechanism.

    PubMed

    Kim, Taeg S; Gorski, Stacey A; Hahn, Steven; Murphy, Kenneth M; Braciale, Thomas J

    2014-03-20

    The contribution of different DC subsets to effector and memory CD8(+) T cell generation during infection and the mechanism by which DCs controls these fate decisions is unclear. Here we demonstrated that the CD103(+) and CD11b(hi) migratory respiratory DC (RDC) subsets after influenza virus infection activated naive virus-specific CD8(+) T cells differentially. CD103(+) RDCs supported the generation of CD8(+) T effector (Teff) cells, which migrate from lymph nodes to the infected lungs. In contrast, migrant CD11b(hi) RDCs activated CD8(+) T cells characteristic of central memory CD8(+) T (CD8(+) Tcm) cells including retention within the draining lymph nodes. CD103(+) RDCs expressed CD24 at an elevated level, contributing to the propensity of this DC subpopulation to support CD8(+) Teff cell differentiation. Mechanistically, CD24 was shown to regulate CD8(+) T cell activation through HMGB1-mediated engagement of T cell RAGE. Thus, there is distribution of labor among DC subsets in regulating CD8(+) T cell differentiation.

  18. Expansion of effector memory regulatory T cells represents a novel prognostic factor in lower risk myelodysplastic syndrome.

    PubMed

    Mailloux, Adam W; Sugimori, Chiharu; Komrokji, Rami S; Yang, Lili; Maciejewski, Jaroslaw P; Sekeres, Mikkael A; Paquette, Ronald; Loughran, Thomas P; List, Alan F; Epling-Burnette, Pearlie K

    2012-09-15

    Myelodysplastic syndromes are premalignant diseases characterized by cytopenias, myeloid dysplasia, immune dysregulation with association to autoimmunity, and variable risk for acute myeloid leukemia transformation. Studies of FOXP3(+) regulatory T cells (Tregs) indicate that the number and/or activation state may influence cancer progression in these patients. Focusing on patients with a lower risk for leukemia transformation, 18 (34.6%) of 52 patients studied displayed an altered Treg compartment compared with age-matched controls. Delineation of unique Treg subsets revealed that an increase in the absolute number of CD4(+)FOXP3(+)CD25(+)CD127(low)CD45RA(-)CD27(-) Tregs (effector memory Tregs [Treg(EM)]) was significantly associated with anemia (p = 0.046), reduced hemoglobin (p = 0.038), and blast counts ≥5% (p = 0.006). In healthy donors, this Treg(EM) population constitutes only 2% of all Tregs (one to six Tregs per microliter) in peripheral blood but, when isolated, exhibit greater suppressive activity in vitro. With a median follow-up of 3.1 y (range 2.7-4.9 y) from sample acquisition, increased numbers of Treg(EM) cells proved to have independent prognostic importance in survival estimates, suggesting that enumeration of this Treg subset may be a more reliable indicator of immunological escape than FOXP3(+) T cells as a whole. Based on multivariate analyses, Treg(EM) impacted survival independently from myeloblast characteristics, cytopenias, karyotype, and comorbidities. Based on these findings, Treg(EM) cell expansion may be synonymous with human Treg activation and indicate microenvironmental changes conducive to transformation in myelodysplastic syndromes.

  19. Defect in recruiting effector memory CD8+ T-cells in malignant pleural effusions compared to normal pleural fluid

    PubMed Central

    2013-01-01

    Background Malignant pleural effusions (MPE) are a common and fatal complication in cancers including lung or breast cancers, or malignant pleural mesothelioma (MPM). MPE animal models and immunotherapy trials in MPM patients previously suggested defects of the cellular immunity in MPE. However only few observational studies of the immune response were done in MPM patients, using questionable control groups (transudate…). Methods We compared T cell populations evaluated by flow cytometry from blood and pleural effusion of untreated patients with MPM (n = 58), pleural metastasis of adenocarcinoma (n = 30) or with benign pleural lesions associated with asbestos exposure (n = 23). Blood and pleural fluid were also obtained from healthy subjects, providing normal values for T cell populations. Results Blood CD4+ or CD8+ T cells percentages were similar in all groups of patients or healthy subjects. Whereas pleural fluid from healthy controls contained mainly CD8+ T cells, benign or malignant pleural effusions included mainly CD4+ T cells. Effector memory T cells were the main T cell subpopulation in pleural fluid from healthy subjects. In contrast, there was a striking and selective recruitment of central memory CD4+ T cells in MPE, but not of effector cells CD8+ T cells or NK cells in the pleural fluid as one would expect in order to obtain an efficient immune response. Conclusions Comparing for the first time MPE to pleural fluid from healthy subjects, we found a local defect in recruiting effector CD8+ T cells, which may be involved in the escape of tumor cells from immune response. Further studies are needed to characterize which subtypes of effector CD8+ T cells are involved, opening prospects for cell therapy in MPE and MPM. PMID:23816056

  20. Application of long-term cultured interferon-gamma enzyme-linked immunospot assay for assessing effector and memory T cell responses in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Effector and memory T cells are generated through developmental programing of naïve cells following antigen recognition. If the infection is controlled, up to 95% of the T cells generated during the expansion phase are eliminated (i.e., contraction phase) and memory T cells remain, sometimes for a l...

  1. A Meloidogyne incognita effector is imported into the nucleus and exhibits transcriptional activation activity in planta.

    PubMed

    Zhang, Lei; Davies, Laura J; Elling, Axel A

    2015-01-01

    Root-knot nematodes are sedentary biotrophic endoparasites that maintain a complex interaction with their host plants. Nematode effector proteins are synthesized in the oesophageal glands of nematodes and secreted into plant tissue through a needle-like stylet. Effectors characterized to date have been shown to mediate processes essential for nematode pathogenesis. To gain an insight into their site of action and putative function, the subcellular localization of 13 previously isolated Meloidogyne incognita effectors was determined. Translational fusions were created between effectors and EGFP-GUS (enhanced green fluorescent protein-β-glucuronidase) reporter genes, which were transiently expressed in tobacco leaf cells. The majority of effectors localized to the cytoplasm, with one effector, 7H08, imported into the nuclei of plant cells. Deletion analysis revealed that the nuclear localization of 7H08 was mediated by two novel independent nuclear localization domains. As a result of the nuclear localization of the effector, 7H08 was tested for the ability to activate gene transcription. 7H08 was found to activate the expression of reporter genes in both yeast and plant systems. This is the first report of a plant-parasitic nematode effector with transcriptional activation activity.

  2. Post-modern pathogens: surprising activities of translocated effectors from E. coli and Legionella.

    PubMed

    Pearson, Jaclyn S; Zhang, Ying; Newton, Hayley J; Hartland, Elizabeth L

    2015-02-01

    Many bacterial pathogens have the ability to manipulate cellular processes and interfere with host cell function through the translocation of bacterial 'effector' proteins. Dedicated protein secretion machines from Gram-negative pathogens, including type III, type IV and type VI secretion systems, inject virulence proteins into infected cells, altering normal cell physiology, including cell structure, metabolism, trafficking and signalling. While effectors were once thought to exert an effect simply by their localization and binding to host cell proteins, increasingly effectors are being recognised as enzymes, in some cases mediating highly novel post-translational modifications on host proteins. Here we highlight some of the more unusual activities of translocated effectors from enteropathogenic Escherichia coli and Legionella pneumophila.

  3. A diametric role for OX40 in the response of effector/memory CD4+ T cells and regulatory T cells to alloantigen

    PubMed Central

    Kinnear, Gillian; Wood, Kathryn J.; Fallah-Arani, Farnaz; Jones, Nick D.

    2013-01-01

    OX40 is a member of the TNFR superfamily that has potent costimulatory properties. Although the impact of blockade of the OX40-OX40L pathway has been well documented in models of autoimmune disease, its effect on the rejection of allografts is less well defined. Here we show that the alloantigen-mediated activation of naïve and memory CD4+ T cells results in the induction of OX40 expression and that blockade of OX40-OX40L interactions prevents skin allograft rejection mediated by either subset of T cells. Moreover, a blocking anti-OX40 was found to have no effect on the activation and proliferation of T cells, but rather effector T cells failed to accumulate in peripheral lymph nodes and subsequently migrate to skin allografts. This was found to be the result of an enhanced degree of cell death amongst proliferating effector cells. In clear contrast, blockade of OX40-OX40L interactions at the time of exposure to alloantigen enhanced the ability of regulatory T cells to suppress T cell responses to alloantigen by supporting rather than diminishing regulatory T cell survival. These data show that OX40-OX40L signalling contributes to the evolution of the adaptive immune response to an allograft via the differential control of alloreactive effector and regulatory T cell survival. Moreover, these data serve to further highlight OX40 and OX40L as therapeutic targets to assist the induction of tolerance to allografts and self-antigens. PMID:23817421

  4. TLR agonists are highly effective at eliciting functional memory CTLs of effector memory phenotype in peptide immunization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Given the importance of memory cytotoxic T lymphocytes (CTLs) in eliminating altered self-cells, including virus-infected and tumor cells, devising effective vaccination strategies for generating memory CTLs is a priority in the field of immunology. Herein, we elaborate upon a novel boosting approac...

  5. Characterization of Effector and Memory T Cell Subsets in the Immune Response to Bovine Tuberculosis in Cattle

    PubMed Central

    Maggioli, Mayara F.; Palmer, Mitchell V.; Thacker, Tyler C.; Vordermeier, H. Martin; Waters, W. Ray

    2015-01-01

    Cultured IFN-γ ELISPOT assays are primarily a measure of central memory T cell (Tcm) responses with humans; however, this important subset of lymphocytes is poorly characterized in cattle. Vaccine-elicited cultured IFN-γ ELISPOT responses correlate with protection against bovine tuberculosis in cattle. However, whether this assay measures cattle Tcm responses or not is uncertain. The objective of the present study was to characterize the relative contribution of Tcm (CCR7+, CD62Lhi, CD45RO+), T effector memory (Tem, defined as: CCR7-, CD62Llow/int, CD45RO+), and T effector cells (CCR7-, CD62L-/low, CD45RO-), in the immune response to Mycobacterium bovis. Peripheral blood mononuclear cells (PBMC) from infected cattle were stimulated with a cocktail of M. bovis purified protein derivative, rTb10.4 and rAg85A for 13 days with periodic addition of fresh media and rIL-2. On day 13, cultured PBMC were re-stimulated with medium alone, rESAT-6:CFP10 or PPDb with fresh autologous adherent cells for antigen presentation. Cultured cells (13 days) or fresh PBMCs (ex vivo response) from the same calves were analyzed for IFN-γ production, proliferation, and CD4, CD45RO, CD62L, CD44, and CCR7 expression via flow cytometry after overnight stimulation. In response to mycobacterial antigens, ~75% of CD4+ IFN-γ+ cells in long-term cultures expressed a Tcm phenotype while less than 10% of the ex vivo response consisted of Tcm cells. Upon re-exposure to antigen, long-term cultured cells were highly proliferative, a distinctive characteristic of Tcm, and the predominant phenotype within the long-term cultures switched from Tcm to Tem. These findings suggest that proliferative responses of Tcm cells to some extent occurs simultaneously with reversion to effector phenotypes (mostly Tem). The present study characterizes Tcm cells of cattle and their participation in the response to M. bovis infection. PMID:25879774

  6. Reversible regulation of aptamer activity with effector-responsive hairpin oligonucleotides.

    PubMed

    Li, Na

    2013-02-01

    Aptamers are oligonucleotides that can bind to various nonnucleic acid molecular targets in a high affinity and specificity. As an emerging class of therapeutic agents, aptamers offer an unparalleled advantage over other classes of therapeutic agents: the possibility to rationally regulate the therapeutic activity of aptamers. Most existing strategies for regulating the aptamer activity have a limited specificity and/or reversibility. Herein we report a simple, generic strategy to simultaneously achieve specificity and reversibility by exploiting the spontaneous conformational change of hairpin oligonucleotides upon the specific recognition of nucleic acid effectors. The effector-responsive hairpin oligonucleotide consists of a sensing loop that recognizes a particular nucleic acid effector, an aptamer stem that inhibits a certain therapeutic target, and an antidote stem that is complementary to the aptamer. Upon the introduction/removal of the effector, the hairpin oligonucleotide undergoes a conformational change that activates/deactivates the aptamer's inhibiting activity on the therapeutic target. This new strategy has been demonstrated with an anticoagulant aptamer that binds and inhibits human α-thrombin. PMID:22651934

  7. Effector and memory T cell subsets in the response to bovine tuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Long-term (i.e., 14d) cultured IFN-gamma ELISPOT assays of PBMC are used as a correlate of T cell central memory (Tcm) responses in cattle and humans. With bovine tuberculosis, vaccine-elicited Tcm responses correlate with protection against experimental Mycobacterium bovis infection. The objective ...

  8. Effector and memory T cell subsets in the response to bovine tuberculosis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Long-term (i.e., 14 days) cultured IFN-gamma ELISPOT assays of peripheral blood mononuclear cells (PBMC) are used to access T cell central memory (Tcm) responses in both cattle and humans. With bovine tuberculosis, vaccine-elicited long-term IFN-gamma ELISPOT response correlates with protection; how...

  9. Long-Term Central and Effector SHIV-Specific Memory T Cell Responses Elicited after a Single Immunization with a Novel Lentivector DNA Vaccine

    PubMed Central

    Arrode-Brusés, Géraldine; Moussa, Maha; Baccard-Longere, Monique; Villinger, François; Chebloune, Yahia

    2014-01-01

    Prevention of HIV acquisition and replication requires long lasting and effective immunity. Given the state of HIV vaccine development, innovative vectors and immunization strategies are urgently needed to generate safe and efficacious HIV vaccines. Here, we developed a novel lentivirus-based DNA vector that does not integrate in the host genome and undergoes a single-cycle of replication. Viral proteins are constitutively expressed under the control of Tat-independent LTR promoter from goat lentivirus. We immunized six macaques once only with CAL-SHIV-IN− DNA using combined intramuscular and intradermal injections plus electroporation. Antigen-specific T cell responses were monitored for 47 weeks post-immunization (PI). PBMCs were assessed directly ex vivo or after 6 and 12 days of in vitro culture using antigenic and/or homeostatic proliferation. IFN-γ ELISPOT was used to measure immediate cytokine secretion from antigen specific effector cells and from memory precursors with high proliferative capacity (PHPC). The memory phenotype and functions (proliferation, cytokine expression, lytic content) of specific T cells were tested using multiparametric FACS-based assays. All immunized macaques developed lasting peripheral CD8+ and CD4+ T cell responses mainly against Gag and Nef antigens. During the primary expansion phase, immediate effector cells as well as increasing numbers of proliferating cells with limited effector functions were detected which expressed markers of effector (EM) and central (CM) memory phenotypes. These responses contracted but then reemerged later in absence of antigen boost. Strong PHPC responses comprising vaccine-specific CM and EM T cells that readily expanded and acquired immediate effector functions were detected at 40/47 weeks PI. Altogether, our study demonstrated that a single immunization with a replication-limited DNA vaccine elicited persistent vaccine-specific CM and EM CD8+ and CD4+ T cells with immediate and readily

  10. Persistence of Protective Immunity to Malaria Induced by DNA Priming and Poxvirus Boosting: Characterization of Effector and Memory CD8+-T-Cell Populations

    PubMed Central

    Sedegah, Martha; Brice, Gary T.; Rogers, William O.; Doolan, Denise L.; Charoenvit, Yupin; Jones, Trevor R.; Majam, Victoria F.; Belmonte, Arnel; Lu, Minh; Belmonte, Maria; Carucci, Daniel J.; Hoffman, Stephen L.

    2002-01-01

    The persistence of immunity to malaria induced in mice by a heterologous DNA priming and poxvirus boosting regimen was characterized. Mice were immunized by priming with DNA vaccine plasmids encoding the Plasmodium yoelii circumsporozoite protein (PyCSP) and murine granulocyte-macrophage colony-stimulating factor and boosting with recombinant vaccinia encoding PyCSP. BALB/c mice immunized with either high-dose (100 μg of p PyCSP plus 30 μg of pGM-CSF) or low-dose (1 μg of p PyCSP plus 1 μg of pGM-CSF DNA) priming were protected against challenge with 50 P. yoelii sporozoites. Protection 2 weeks after immunization was 70 to 100%, persisted at this level for at least 20 weeks, and declined to 30 to 40% by 28 weeks. Eight of eight mice protected at 20 weeks were still protected when rechallenged at 40 weeks. The antigen (Ag)-specific effector CD8+-T-cell population present 2 weeks after boosting had ex vivo Ag-specific cytolytic activity, expressed both gamma interferon (IFN-γ) and tumor necrosis factor alpha, and constituted 12 to 20% of splenic CD8+ T cells. In contrast, the memory CD8+-Ag-specific-cell population at 28 weeks lacked cytolytic activity and constituted only 6% of splenic CD8+ T cells, but at the single-cell level it produced significantly higher levels of IFN-γ than the effectors. High levels of Ag- or parasite-specific antibodies present 2 weeks after boosting had declined three- to sevenfold by 28 weeks. Low-dose priming was similarly immunogenic and as protective as high-dose priming against a 50-, but not a 250-, sporozoite challenge. These results demonstrate that a heterologous priming and boosting vaccination can provide lasting protection against malaria in this model system. PMID:12065488

  11. Mechanism of IRSp53 inhibition and combinatorial activation by Cdc42 and downstream effectors

    PubMed Central

    Kast, David J; Yang, Changsong; Disanza, Andrea; Boczkowska, Malgorzata; Madasu, Yadaiah; Scita, Giorgio; Svitkina, Tatyana; Dominguez, Roberto

    2014-01-01

    The Rho family GTPase effector IRSp53 has essential roles in filopodia formation and neuronal development, but its regulatory mechanism is poorly understood. IRSp53 contains a membrane-binding BAR domain followed by an unconventional CRIB motif that overlaps with a proline-rich region (CRIB–PR) and an SH3 domain that recruits actin cytoskeleton effectors. Using a fluorescence reporter assay, we show that human IRSp53 adopts a closed inactive conformation that opens synergistically with the binding of human Cdc42 to the CRIB–PR and effector proteins, such as the tumor-promoting factor Eps8, to the SH3 domain. The crystal structure of Cdc42 bound to the CRIB–PR reveals a new mode of effector binding to Rho family GTPases. Structure-inspired mutations disrupt autoinhibition and Cdc42 binding in vitro and decouple Cdc42- and IRSp53-dependent filopodia formation in cells. The data support a combinatorial mechanism of IRSp53 activation. PMID:24584464

  12. Effector and Central Memory Poly-Functional CD4+ and CD8+ T Cells are Boosted upon ZOSTAVAX® Vaccination

    PubMed Central

    Sei, Janet J.; Cox, Kara S.; Dubey, Sheri A.; Antonello, Joseph M.; Krah, David L.; Casimiro, Danilo R.; Vora, Kalpit A.

    2015-01-01

    ZOSTAVAX® is a live attenuated varicella-zoster virus (VZV) vaccine that is licensed for the protection of individuals ≥50 years against shingles and its most common complication, postherpetic neuralgia. While IFNγ responses increase upon vaccination, the quality of the T cell response has not been elucidated. By using polychromatic flow cytometry, we characterized the breadth, magnitude, and quality of ex vivo CD4+ and CD8+ T cell responses induced 3–4 weeks after ZOSTAVAX vaccination of healthy adults. We show, for the first time that the highest frequencies of VZV-specific CD4+ T cells were poly-functional CD154+IFNγ+IL-2+TNFα+ cells, which were boosted upon vaccination. The CD4+ T cells were broadly reactive to several VZV proteins, with immediate early (IE) 63 ranking the highest among them in the fold rise of poly-functional cells, followed by IE62, gB, open reading frame (ORF) 9, and gE. We identified a novel poly-functional ORF9-specific CD8+ T cell population in 62% of the subjects, and these were boosted upon vaccination. Poly-functional CD4+ and CD8+ T cells produced significantly higher levels of IFNγ, IL-2, and TNFα compared to mono-functional cells. After vaccination, a boost in the expression of IFNγ by poly-functional IE63- and ORF9-specific CD4+ T cells and IFNγ, IL-2, and TNFα by ORF9-specific poly-functional CD8+ T cells was observed. Responding poly-functional T cells exhibited both effector (CCR7−CD45RA−CD45RO+), and central (CCR7+CD45RA−CD45RO+) memory phenotypes, which expressed comparable levels of cytokines. Altogether, our studies demonstrate that a boost in memory poly-functional CD4+ T cells and ORF9-specific CD8+ T cells may contribute toward ZOSTAVAX efficacy. PMID:26579128

  13. Platelet activation attracts a subpopulation of effector monocytes to sites of Leishmania major infection.

    PubMed

    Goncalves, Ricardo; Zhang, Xia; Cohen, Heather; Debrabant, Alain; Mosser, David M

    2011-06-01

    Leishmania species trigger a brisk inflammatory response and efficiently induce cell-mediated immunity. We examined the mechanisms whereby leukocytes were recruited into lesions after Leishmania major infection of mice. We found that a subpopulation of effector monocytes expressing the granulocyte marker GR1 (Ly6C) is rapidly recruited into lesions, and these monocytes efficiently kill L. major parasites. The recruitment of this subpopulation of monocytes depends on the chemokine receptor CCR2 and the activation of platelets. Activated platelets secrete platelet-derived growth factor, which induces the rapid release of CCL2 from leukocytes and mesenchymal cells. This work points to a new role for platelets in host defense involving the selective recruitment of a subpopulation of effector monocytes from the blood to efficiently kill this intracellular parasite.

  14. Expansion of dysfunctional Tim-3-expressing effector memory CD8+ T cells during simian immunodeficiency virus infection in rhesus macaques.

    PubMed

    Fujita, Tsuyoshi; Burwitz, Benjamin J; Chew, Glen M; Reed, Jason S; Pathak, Reesab; Seger, Elizabeth; Clayton, Kiera L; Rini, James M; Ostrowski, Mario A; Ishii, Naoto; Kuroda, Marcelo J; Hansen, Scott G; Sacha, Jonah B; Ndhlovu, Lishomwa C

    2014-12-01

    The T cell Ig- and mucin domain-containing molecule-3 (Tim-3) negative immune checkpoint receptor demarcates functionally exhausted CD8(+) T cells arising from chronic stimulation in viral infections like HIV. Tim-3 blockade leads to improved antiviral CD8(+) T cell responses in vitro and, therefore, represents a novel intervention strategy to restore T cell function in vivo and protect from disease progression. However, the Tim-3 pathway in the physiologically relevant rhesus macaque SIV model of AIDS remains uncharacterized. We report that Tim-3(+)CD8(+) T cell frequencies are significantly increased in lymph nodes, but not in peripheral blood, in SIV-infected animals. Tim-3(+)PD-1(+)CD8(+) T cells are similarly increased during SIV infection and positively correlate with SIV plasma viremia. Tim-3 expression was found primarily on effector memory CD8(+) T cells in all tissues examined. Tim-3(+)CD8(+) T cells have lower Ki-67 content and minimal cytokine responses to SIV compared with Tim-3(-)CD8(+) T cells. During acute-phase SIV replication, Tim-3 expression peaked on SIV-specific CD8(+) T cells by 2 wk postinfection and then rapidly diminished, irrespective of mutational escape of cognate Ag, suggesting non-TCR-driven mechanisms for Tim-3 expression. Thus, rhesus Tim-3 in SIV infection partially mimics human Tim-3 in HIV infection and may serve as a novel model for targeted studies focused on rejuvenating HIV-specific CD8(+) T cell responses. PMID:25348621

  15. TNFSF10/TRAIL regulates human T4 effector memory lymphocyte radiosensitivity and predicts radiation-induced acute and subacute dermatitis

    PubMed Central

    Baijer, Jan; Déchamps, Nathalie; Perdry, Hervé; Morales, Pablo; Kerns, Sarah; Vasilescu, Alexandre; Baulande, Sylvain; Azria, David; Roméo, Paul Henri; Schmitz, Annette

    2016-01-01

    Sensitivity of T4 effector-memory (T4EM) lymphocytes to radiation-induced apoptosis shows heritability compatible with a Mendelian mode of transmission. Using gene expression studies and flow cytometry, we show a higher TNF-Related Apoptosis Inducing Ligand (TRAIL/TNFSF10) mRNA level and a higher level of membrane bound TRAIL (mTRAIL) on radiosensitive compared to radioresistant T4EM lymphocytes. Functionally, we show that mTRAIL mediates a pro-apoptotic autocrine signaling after irradiation of T4EM lymphocytes linking mTRAIL expression to T4EM radiosensitivity. Using single marker and multimarker Family-Based Association Testing, we identified 3 SNPs in the TRAIL gene that are significantly associated with T4EM lymphocytes radiosensitivity. Among these 3 SNPs, two are also associated with acute and subacute dermatitis after radiotherapy in breast cancer indicating that T4EM lymphocytes radiosensitivity may be used to predict response to radiotherapy. Altogether, these results show that mTRAIL level regulates the response of T4EM lymphocytes to ionizing radiation and suggest that TRAIL/TNFSF10 genetic variants hold promise as markers of individual radiosensitivity. PMID:26982083

  16. Rapid parallel flow cytometry assays of active GTPases using effector beads

    PubMed Central

    Buranda, Tione; BasuRay, Soumik; Swanson, Scarlett; Agola, Jacob; Bondu, Virginie; Wandinger-Ness, Angela

    2013-01-01

    We describe a rapid assay for measuring the cellular activity of small GTPases in response to a specific stimulus. Effector functionalized beads are used to quantify in parallel multiple, GTP-bound GTPases in the same cell lysate by flow cytometry. In a biologically relevant example, five different Ras family GTPases are shown for the first time to be involved in a concerted signaling cascade downstream of receptor ligation by Sin Nombre hantavirus. PMID:23928044

  17. Kindlin-3 regulates integrin activation and adhesion reinforcement of effector T cells.

    PubMed

    Moretti, Federico A; Moser, Markus; Lyck, Ruth; Abadier, Michael; Ruppert, Raphael; Engelhardt, Britta; Fässler, Reinhard

    2013-10-15

    Activated T cells use very late antigen-4/α4β1 integrin for capture, rolling on, and firm adhesion to endothelial cells, and use leukocyte function-associated antigen-1/αLβ2 integrin for subsequent crawling and extravasation. Inhibition of α4β1 is sufficient to prevent extravasation of activated T cells and is successfully used to combat autoimmune diseases, such as multiple sclerosis. Here we show that effector T cells lacking the integrin activator Kindlin-3 extravasate and induce experimental autoimmune encephalomyelitis in mice immunized with autoantigen. In sharp contrast, adoptively transferred autoreactive T cells from Kindlin-3-deficient mice fail to extravasate into the naïve CNS. Mechanistically, autoreactive Kindlin-3-null T cells extravasate when the CNS is inflamed and the brain microvasculature expresses high levels of integrin ligands. Flow chamber assays under physiological shear conditions confirmed that Kindlin-3-null effector T cells adhere to high concentrations of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1, albeit less efficiently than WT T cells. Although these arrested T cells polarize and start crawling, only few remain firmly adherent over time. Our data demonstrate that the requirement of Kindlin-3 for effector T cells to induce α4β1 and αLβ2 integrin ligand binding and stabilization of integrin-ligand bonds is critical when integrin ligand levels are low, but of less importance when integrin ligand levels are high. PMID:24089451

  18. Memory and effector CD8 T-cell responses after nanoparticle vaccination of melanoma patients.

    PubMed

    Speiser, Daniel E; Schwarz, Katrin; Baumgaertner, Petra; Manolova, Vania; Devevre, Estelle; Sterry, Wolfram; Walden, Peter; Zippelius, Alfred; Conzett, Katrin Baumann; Senti, Gabriela; Voelter, Verena; Cerottini, Jean-Philippe; Guggisberg, David; Willers, Jörg; Geldhof, Christine; Romero, Pedro; Kündig, Thomas; Knuth, Alexander; Dummer, Reinhard; Trefzer, Uwe; Bachmann, Martin F

    2010-10-01

    Induction of cytotoxic CD8 T-cell responses is enhanced by the exclusive presentation of antigen through dendritic cells, and by innate stimuli, such as toll-like receptor ligands. On the basis of these 2 principles, we designed a vaccine against melanoma. Specifically, we linked the melanoma-specific Melan-A/Mart-1 peptide to virus-like nanoparticles loaded with A-type CpG, a ligand for toll-like receptor 9. Melan-A/Mart-1 peptide was cross-presented, as shown in vitro with human dendritic cells and in HLA-A2 transgenic mice. A phase I/II study in stage II-IV melanoma patients showed that the vaccine was well tolerated, and that 14/22 patients generated ex vivo detectable T-cell responses, with in part multifunctional T cells capable to degranulate and produce IFN-γ, TNF-α, and IL-2. No significant influence of the route of immunization (subcutaneous versus intradermal) nor dosing regimen (weekly versus daily clusters) could be observed. It is interesting to note that, relatively large fractions of responding specific T cells exhibited a central memory phenotype, more than what is achieved by other nonlive vaccines. We conclude that vaccination with CpG loaded virus-like nanoparticles is associated with a human CD8 T-cell response with properties of a potential long-term immune protection from the disease. PMID:20842051

  19. MARTX effector cross kingdom activation by Golgi-associated ADP-ribosylation factors.

    PubMed

    Kim, Byoung Sik; Satchell, Karla J F

    2016-08-01

    Vibrio vulnificus infects humans and causes lethal septicemia. The primary virulence factor is a multifunctional-autoprocessing repeats-in-toxin (MARTX) toxin consisting of conserved repeats-containing regions and various effector domains. Recent genomic analyses for the newly emerged V. vulnificus biotype 3 strain revealed that its MARTX toxin has two previously unknown effector domains. Herein, we characterized one of these domains, Domain X (DmXVv ). A structure-based homology search revealed that DmXVv belongs to the C58B cysteine peptidase subfamily. When ectopically expressed in cells, DmXVv was autoprocessed and induced cytopathicity including Golgi dispersion. When the catalytic cysteine or the region flanking the scissile bond was mutated, both autoprocessing and cytopathicity were significantly reduced indicating that DmXVv cytopathicity is activated by amino-terminal autoprocessing. Consistent with this, host cell protein export was affected by Vibrio cells producing a toxin with wild-type, but not catalytically inactive, DmXVv . DmXVv was found to localize to Golgi and to directly interact with Golgi-associated ADP-ribosylation factors ARF1, ARF3 and ARF4, although ARF binding was not necessary for the subcellular localization. Rather, this interaction was found to induce autoprocessing of DmXVv . These data demonstrate that the V. vulnificus hijacks the host ARF proteins to activate the cytopathic DmXVv effector domain of MARTX toxin. PMID:26780191

  20. Antigen-dependent proliferation and cytokine induction in respiratory syncytial virus-infected cotton rats reflect the presence of effector-memory T cells

    SciTech Connect

    Richter, Bettina W.M.; Onuska, Jaya M.; Niewiesk, Stefan; Prince, Gregory A.; Eichelberger, Maryna C. . E-mail: MarynaE@virionsystems.com

    2005-06-20

    Respiratory syncytial virus (RSV) is a major cause of lower airway disease in infants and children. Immunity to RSV is not long lasting, resulting in re-occurring infections throughout life. Effective long-lived immunity results when central-memory T cells that proliferate vigorously and secrete IL-2 are present. In contrast, effector-memory T cells that mainly produce IFN-{gamma}, facilitate virus clearance but are not long lived. To identify the type of memory response induced after RSV-A (Long) infection, we characterized the kinetics of the antigen-specific immune response and identified the types of cytokines induced. RSV-specific lymphocytic proliferation following primary and secondary infection was similar, and in both cases responses waned within a short period of time. In addition, mRNA for IFN-{gamma} but not IL-2 was induced in RSV-specific CD4{sup +} T cells. This supports the idea that the presence of effector-memory rather than central-memory T cells contributes to the ineffectiveness of the immune response to RSV.

  1. Constitutive expression of CCR7 directs effector CD8 T cells into the splenic white pulp and impairs functional activity.

    PubMed

    Unsoeld, Heike; Voehringer, David; Krautwald, Stefan; Pircher, Hanspeter

    2004-09-01

    Antigenic stimulation down-regulates CCR7 on effector T cells. To analyze the importance of CCR7 down-regulation, transgenic (tg) mice constitutively expressing CCR7 were generated. CD8 T cells with defined Ag specificity were obtained by breeding CCR7-tg mice with P14 TCR-tg mice specific for lymphocytic choriomeningitis virus. Transgenic CCR7 expression did not impair proliferation of P14.CCR7 T cells induced by lymphocytic choriomeningitis virus infection, but prevented CCR7 down-regulation. Compared with wild-type P14 effector cells, P14.CCR7 effector cells, expressing the CCR7 transgene, were increased in the spleen, but decreased in blood and peripheral tissues. Moreover, P14.CCR7 effector cells localized almost exclusively in the splenic white pulp, whereas P14 effector cells were excluded from splenic white pulp cords and were found preferentially in the red pulp. Functional experiments further revealed that P14.CCR7 effector cells were impaired in rapid viral clearance and in inducing Ag-specific delayed-type hypersensitivity reactions. Thus, the present study demonstrates that down-regulation of CCR7 during CD8 T cell activation is important to release effector cells from the white pulp of the spleen, and highlights the importance of effector cell localization in providing rapid immunity. PMID:15322160

  2. Bacterial Effector Activates Jasmonate Signaling by Directly Targeting JAZ Transcriptional Repressors

    PubMed Central

    Jiang, Shushu; Yao, Jian; Ma, Ka-Wai; Zhou, Huanbin; Song, Jikui; He, Sheng Yang; Ma, Wenbo

    2013-01-01

    Gram-negative bacterial pathogens deliver a variety of virulence proteins through the type III secretion system (T3SS) directly into the host cytoplasm. These type III secreted effectors (T3SEs) play an essential role in bacterial infection, mainly by targeting host immunity. However, the molecular basis of their functionalities remains largely enigmatic. Here, we show that the Pseudomonas syringae T3SE HopZ1a, a member of the widely distributed YopJ effector family, directly interacts with jasmonate ZIM-domain (JAZ) proteins through the conserved Jas domain in plant hosts. JAZs are transcription repressors of jasmonate (JA)-responsive genes and major components of the jasmonate receptor complex. Upon interaction, JAZs can be acetylated by HopZ1a through a putative acetyltransferase activity. Importantly, P. syringae producing the wild-type, but not a catalytic mutant of HopZ1a, promotes the degradation of HopZ1-interacting JAZs and activates JA signaling during bacterial infection. Furthermore, HopZ1a could partially rescue the virulence defect of a P. syringae mutant that lacks the production of coronatine, a JA-mimicking phytotoxin produced by a few P. syringae strains. These results highlight a novel example by which a bacterial effector directly manipulates the core regulators of phytohormone signaling to facilitate infection. The targeting of JAZ repressors by both coronatine toxin and HopZ1 effector suggests that the JA receptor complex is potentially a major hub of host targets for bacterial pathogens. PMID:24204266

  3. 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. PMID:27271569

  4. IgE epitope proximity determines immune complex shape and effector cell activation capacity

    PubMed Central

    Gieras, Anna; Linhart, Birgit; Roux, Kenneth H.; Dutta, Moumita; Khodoun, Marat; Zafred, Domen; Cabauatan, Clarissa R.; Lupinek, Christian; Weber, Milena; Focke-Tejkl, Margarete; Keller, Walter; Finkelman, Fred D.; Valenta, Rudolf

    2016-01-01

    Background IgE-allergen complexes induce mast cell and basophil activation and thus immediate allergic inflammation. They are also important for IgE-facilitated allergen presentation to T cells by antigen-presenting cells. Objective To investigate whether the proximity of IgE binding sites on an allergen affects immune complex shape and subsequent effector cell activation in vitro and in vivo. Methods We constructed artificial allergens by grafting IgE epitopes in different numbers and proximity onto a scaffold protein. The shape of immune complexes formed between artificial allergens and the corresponding IgE was studied by negative-stain electron microscopy. Allergenic activity was determined using basophil activation assays. Mice were primed with IgE, followed by injection of artificial allergens to evaluate their in vivo allergenic activity. Severity of systemic anaphylaxis was measured by changes in body temperature. Results We could demonstrate simultaneous binding of 4 IgE antibodies in close vicinity to each other. The proximity of IgE binding sites on allergens influenced the shape of the resulting immune complexes and the magnitude of effector cell activation and in vivo inflammation. Conclusions Our results demonstrate that the proximity of IgE epitopes on an allergen affects its allergenic activity. We thus identified a novel mechanism by which IgE-allergen complexes regulate allergic inflammation. This mechanism should be important for allergy and other immune complex–mediated diseases. PMID:26684291

  5. Cytotoxic Necrotizing Factor-Y Boosts Yersinia Effector Translocation by Activating Rac Protein*

    PubMed Central

    Wolters, Manuel; Boyle, Erin C.; Lardong, Kerstin; Trülzsch, Konrad; Steffen, Anika; Rottner, Klemens; Ruckdeschel, Klaus; Aepfelbacher, Martin

    2013-01-01

    Pathogenic Yersinia spp. translocate the effectors YopT, YopE, and YopO/YpkA into target cells to inactivate Rho family GTP-binding proteins and block immune responses. Some Yersinia spp. also secrete the Rho protein activator cytotoxic necrotizing factor-Y (CNF-Y), but it has been unclear how the bacteria may benefit from Rho protein activation. We show here that CNF-Y increases Yop translocation in Yersinia enterocolitica-infected cells up to 5-fold. CNF-Y strongly activated RhoA and also delayed in time Rac1 and Cdc42, but when individually expressed, constitutively active mutants of Rac1, but not of RhoA, increased Yop translocation. Consistently, knock-out or knockdown of Rac1 but not of RhoA, -B, or -C inhibited Yersinia effector translocation in CNF-Y-treated and control cells. Activation or knockdown of Cdc42 also affected Yop translocation but much less efficiently than Rac. The increase in Yop translocation induced by CNF-Y was essentially independent of the presence of YopE, YopT, or YopO in the infecting Yersinia strain, indicating that none of the Yops reported to inhibit translocation could reverse the CNF-Y effect. In summary, the CNF-Y activity of Yersinia strongly enhances Yop translocation through activation of Rac. PMID:23803609

  6. Identification and characterization of a 29-kilodalton protein from Mycobacterium tuberculosis culture filtrate recognized by mouse memory effector cells.

    PubMed

    Rosenkrands, I; Rasmussen, P B; Carnio, M; Jacobsen, S; Theisen, M; Andersen, P

    1998-06-01

    Culture filtrate proteins from Mycobacterium tuberculosis induce protective immunity in various animal models of tuberculosis. Two molecular mass regions (6 to 10 kDa and 24 to 36 kDa) of short-term culture filtrate are preferentially recognized by Th1 cells in animal models as well as by patients with minimal disease. In the present study, the 24- to 36-kDa region has been studied, and the T-cell reactivity has been mapped in detail. Monoclonal antibodies were generated, and one monoclonal antibody, HYB 71-2, with reactivity against a 29-kDa antigen located in the highly reactive region below the antigen 85 complex was selected. The 29-kDa antigen (CFP29) was purified from M. tuberculosis short-term culture filtrate by thiophilic adsorption chromatography, anion-exchange chromatography, and gel filtration. In its native form, CFP29 forms a polymer with a high molecular mass. CFP29 was mapped in two-dimensional electrophoresis gels as three distinct spots just below the antigen 85 complex component MPT59. CFP29 is present in both culture filtrate and the membrane fraction from M. tuberculosis, suggesting that this antigen is released from the envelope to culture filtrate during growth. Determination of the N-terminal amino acid sequence allowed cloning and sequencing of the cfp29 gene. The nucleotide sequence showed 62% identity to the bacteriocin Linocin from Brevibacterium linens. Purified recombinant histidine-tagged CFP29 and native CFP29 had similar T-cell stimulatory properties, and they both elicited the release of high levels of gamma interferon from mouse memory effector cells isolated during the recall of protective immunity to tuberculosis. Interspecies analysis by immunoblotting and PCR demonstrated that CFP29 is widely distributed in mycobacterial species.

  7. Activating Attachments Reduces Memories of Traumatic Images.

    PubMed

    Bryant, Richard A; Foord, Rachael

    2016-01-01

    Emotional memories, and especially intrusive memories, are a common feature of many psychological disorders, and are overconsolidated by stress. Attachment theory posits that activation of mental representations of attachment figures can reduce stress and boost coping. This study tested the proposition that attachment activation would reduce consolidation of emotional and intrusive memories. Sixty-seven undergraduate students viewed subliminal presentations of traumatic and neutral images, which were preceded by subliminal presentations of either attachment-related images or non-attachment-related images; free recall and intrusive memories were assessed two days later. Participants with low avoidant attachment tendencies who received the attachment primes recalled fewer memories and reported fewer intrusions than those who received the non-attachment primes. Unexpectedly, those with high anxious attachment tendencies reported fewer memories. These findings generally accord with attachment theory, and suggest that consolidation of emotional memories can be moderated by activation of attachment representations. PMID:27631498

  8. Activating Attachments Reduces Memories of Traumatic Images.

    PubMed

    Bryant, Richard A; Foord, Rachael

    2016-01-01

    Emotional memories, and especially intrusive memories, are a common feature of many psychological disorders, and are overconsolidated by stress. Attachment theory posits that activation of mental representations of attachment figures can reduce stress and boost coping. This study tested the proposition that attachment activation would reduce consolidation of emotional and intrusive memories. Sixty-seven undergraduate students viewed subliminal presentations of traumatic and neutral images, which were preceded by subliminal presentations of either attachment-related images or non-attachment-related images; free recall and intrusive memories were assessed two days later. Participants with low avoidant attachment tendencies who received the attachment primes recalled fewer memories and reported fewer intrusions than those who received the non-attachment primes. Unexpectedly, those with high anxious attachment tendencies reported fewer memories. These findings generally accord with attachment theory, and suggest that consolidation of emotional memories can be moderated by activation of attachment representations.

  9. Activating Attachments Reduces Memories of Traumatic Images

    PubMed Central

    Foord, Rachael

    2016-01-01

    Emotional memories, and especially intrusive memories, are a common feature of many psychological disorders, and are overconsolidated by stress. Attachment theory posits that activation of mental representations of attachment figures can reduce stress and boost coping. This study tested the proposition that attachment activation would reduce consolidation of emotional and intrusive memories. Sixty-seven undergraduate students viewed subliminal presentations of traumatic and neutral images, which were preceded by subliminal presentations of either attachment-related images or non-attachment-related images; free recall and intrusive memories were assessed two days later. Participants with low avoidant attachment tendencies who received the attachment primes recalled fewer memories and reported fewer intrusions than those who received the non-attachment primes. Unexpectedly, those with high anxious attachment tendencies reported fewer memories. These findings generally accord with attachment theory, and suggest that consolidation of emotional memories can be moderated by activation of attachment representations. PMID:27631498

  10. Real-time tracking of cell cycle progression during CD8+ effector and memory T-cell differentiation

    PubMed Central

    Kinjyo, Ichiko; Qin, Jim; Tan, Sioh-Yang; Wellard, Cameron J.; Mrass, Paulus; Ritchie, William; Doi, Atsushi; Cavanagh, Lois L.; Tomura, Michio; Sakaue-Sawano, Asako; Kanagawa, Osami; Miyawaki, Atsushi; Hodgkin, Philip D.; Weninger, Wolfgang

    2015-01-01

    The precise pathways of memory T-cell differentiation are incompletely understood. Here we exploit transgenic mice expressing fluorescent cell cycle indicators to longitudinally track the division dynamics of individual CD8+ T cells. During influenza virus infection in vivo, naive T cells enter a CD62Lintermediate state of fast proliferation, which continues for at least nine generations. At the peak of the anti-viral immune response, a subpopulation of these cells markedly reduces their cycling speed and acquires a CD62Lhi central memory cell phenotype. Construction of T-cell family division trees in vitro reveals two patterns of proliferation dynamics. While cells initially divide rapidly with moderate stochastic variations of cycling times after each generation, a slow-cycling subpopulation displaying a CD62Lhi memory phenotype appears after eight divisions. Phenotype and cell cycle duration are inherited by the progeny of slow cyclers. We propose that memory precursors cell-intrinsically modulate their proliferative activity to diversify differentiation pathways. PMID:25709008

  11. Nano-particle vaccination combined with TLR-7 and -9 ligands triggers memory and effector CD8⁺ T-cell responses in melanoma patients.

    PubMed

    Goldinger, Simone M; Dummer, Reinhard; Baumgaertner, Petra; Mihic-Probst, Daniela; Schwarz, Katrin; Hammann-Haenni, Anya; Willers, Joerg; Geldhof, Christine; Prior, John O; Kündig, Thomas M; Michielin, Olivier; Bachmann, Martin F; Speiser, Daniel E

    2012-11-01

    Optimal vaccine strategies must be identified for improving T-cell vaccination against infectious and malignant diseases. MelQbG10 is a virus-like nano-particle loaded with A-type CpG-oligonucleotides (CpG-ODN) and coupled to peptide(16-35) derived from Melan-A/MART-1. In this phase IIa clinical study, four groups of stage III-IV melanoma patients were vaccinated with MelQbG10, given (i) with IFA (Montanide) s.c.; (ii) with IFA s.c. and topical Imiquimod; (iii) i.d. with topical Imiquimod; or (iv) as intralymph node injection. In total, 16/21 (76%) patients generated ex vivo detectable Melan-A/MART-1-specific T-cell responses. T-cell frequencies were significantly higher when IFA was used as adjuvant, resulting in detectable T-cell responses in all (11/11) patients, with predominant generation of effector-memory-phenotype cells. In turn, Imiquimod induced higher proportions of central-memory-phenotype cells and increased percentages of CD127(+) (IL-7R) T cells. Direct injection of MelQbG10 into lymph nodes resulted in lower T-cell frequencies, associated with lower proportions of memory and effector-phenotype T cells. Swelling of vaccine site draining lymph nodes, and increased glucose uptake at PET/CT was observed in 13/15 (87%) of evaluable patients, reflecting vaccine triggered immune reactions in lymph nodes. We conclude that the simultaneous use of both Imiquimod and CpG-ODN induced combined memory and effector CD8(+) T-cell responses. PMID:22806397

  12. Sandia Cognitive Runtime Engine with Active Memory

    2005-12-01

    The SCREAM (Sandia Cognitive Runtime Engine with Active memory) software implements a subset of a Cognitive Famework developed at Sandia National Laboratories. The software is implemented in the Umbra simulation and modular software framework, which is C++-based. SCREAM components include a Concept Instance Driver, Semantic Activation Network, Concept Database, Context Recognizer, Context Database, Episodic Memory, Egocentric Spatial Memory, Allocentric Spatial Memory, Comparator, and a Context to Abstract Action converter. At initialization, modules load the datamore » files that together specify all the components of a particular cognitive model, such as concept declarations, context declarations, spreading activation weights, and context/situation-cue-patterns.« less

  13. Sandia Cognitive Runtime Engine with Active Memory

    SciTech Connect

    Xavier, Patrick; Chen, Michael C.; Hart, Brian; Hart, Derek; Lippitt, Carl; Wolfenbarger, Paul; Waymire, Russel

    2005-12-01

    The SCREAM (Sandia Cognitive Runtime Engine with Active memory) software implements a subset of a Cognitive Famework developed at Sandia National Laboratories. The software is implemented in the Umbra simulation and modular software framework, which is C++-based. SCREAM components include a Concept Instance Driver, Semantic Activation Network, Concept Database, Context Recognizer, Context Database, Episodic Memory, Egocentric Spatial Memory, Allocentric Spatial Memory, Comparator, and a Context to Abstract Action converter. At initialization, modules load the data files that together specify all the components of a particular cognitive model, such as concept declarations, context declarations, spreading activation weights, and context/situation-cue-patterns.

  14. CC Chemokine Receptor 7 Expression by Effector/Memory CD4+ T Cells Depends on Antigen Specificity and Tissue Localization during Influenza A Virus Infection

    PubMed Central

    Debes, Gudrun F.; Bonhagen, Kerstin; Wolff, Thorsten; Kretschmer, Ute; Krautwald, Stefan; Kamradt, Thomas; Hamann, Alf

    2004-01-01

    The lung is an important entry site for respiratory pathogens such as influenza A virus. In order to combat such invading infectious agents, effector/memory T cells home to the lung and other peripheral tissues as well as lymphoid organs. In this process, chemokines and their receptors fulfill important roles in the guidance of T cells into such organs and specialized microenvironments within tissues. In this study, we determined if CD4+ T cells residing in different lung compartments and draining lymph nodes of influenza A virus-infected and naïve mice express receptors allowing their recirculation into secondary lymphoid tissues. We found high levels of l-selectin and CC chemokine receptor 7 (CCR7) expression in lung-derived CD4+ T cells, similar to that detected on T cells in secondary lymphoid organs. Upon influenza A virus infection, the bulk of gamma interferon-positive (IFN-γ+) and IFN-γ− CD4+ T cells recovered from lung parenchyma retained functional CCR7, whereas virus-specific IFN-γ-producing T cells were CCR7−. In contrast, a majority of virus-specific IFN-γ+ T cells in the lung draining lymph node were CCR7+. Independent of infection, CD4+ T cells obtained from the lung airways exhibited the lowest expression level of l-selectin and CCR7, indicating that T cells at this anatomical site represent the most differentiated effector cell type, lacking the ability to recirculate. Our results suggest that effector/memory T cells that enter inflammatory sites retain functional CCR7 expression, which is lost only upon response to viral antigen and after localization to the final effector site. PMID:15220427

  15. CC chemokine receptor 7 expression by effector/memory CD4+ T cells depends on antigen specificity and tissue localization during influenza A virus infection.

    PubMed

    Debes, Gudrun F; Bonhagen, Kerstin; Wolff, Thorsten; Kretschmer, Ute; Krautwald, Stefan; Kamradt, Thomas; Hamann, Alf

    2004-07-01

    The lung is an important entry site for respiratory pathogens such as influenza A virus. In order to combat such invading infectious agents, effector/memory T cells home to the lung and other peripheral tissues as well as lymphoid organs. In this process, chemokines and their receptors fulfill important roles in the guidance of T cells into such organs and specialized microenvironments within tissues. In this study, we determined if CD4(+) T cells residing in different lung compartments and draining lymph nodes of influenza A virus-infected and naïve mice express receptors allowing their recirculation into secondary lymphoid tissues. We found high levels of l-selectin and CC chemokine receptor 7 (CCR7) expression in lung-derived CD4(+) T cells, similar to that detected on T cells in secondary lymphoid organs. Upon influenza A virus infection, the bulk of gamma interferon-positive (IFN-gamma(+)) and IFN-gamma(-) CD4(+) T cells recovered from lung parenchyma retained functional CCR7, whereas virus-specific IFN-gamma-producing T cells were CCR7(-). In contrast, a majority of virus-specific IFN-gamma(+) T cells in the lung draining lymph node were CCR7(+). Independent of infection, CD4(+) T cells obtained from the lung airways exhibited the lowest expression level of l-selectin and CCR7, indicating that T cells at this anatomical site represent the most differentiated effector cell type, lacking the ability to recirculate. Our results suggest that effector/memory T cells that enter inflammatory sites retain functional CCR7 expression, which is lost only upon response to viral antigen and after localization to the final effector site. PMID:15220427

  16. Effector and suppressor circuits of the immune response are activated in vivo by different mechanisms.

    PubMed

    Okamoto, H; Kripke, M L

    1987-06-01

    The application of fluorescein isothiocyanate (FITC) onto the skin of mice induces a contact hypersensitivity immune response. Lymph nodes draining the skin painted with FITC contain fluorescent cells that induce contact hypersensitivity to FITC when injected into normal mice. The antigen-presenting cells responsible for activating the effector pathway of the contact hypersensitivity response express Ia histocompatibility determinants and are resistant to inactivation with gamma-radiation. Exposing the skin to low doses of UV radiation (280-320 nm) before the application of FITC suppresses the contact hypersensitivity response to FITC. Cells present in the draining lymph nodes of these mice induce suppressor T lymphocytes when injected into normal recipients. The inducer cells in the draining lymph nodes are Thy 1+, Ia- and are inactivated by gamma-radiation. These studies demonstrate that different mechanisms are involved in the in vivo activation of effector and suppressor immune responses, and they suggest that the mode of initial antigen presentation determines which immunologic circuit will be activated in response to a contact-sensitizing antigen. PMID:2884661

  17. Effector and suppressor circuits of the immune response are activated in vivo by different mechanisms

    SciTech Connect

    Okamoto, H.; Kripke, M.L.

    1987-06-01

    The application of fluorescein isothiocyanate (FITC) onto the skin of mice induces a contact hypersensitivity immune response. Lymph nodes draining the skin painted with FITC contain fluorescent cells that induce contact hypersensitivity to FITC when injected into normal mice. The antigen-presenting cells responsible for activating the effector pathway of the contact hypersensitivity response express Ia histocompatibility determinants and are resistant to inactivation with gamma-radiation. Exposing the skin to low doses of UV radiation (280-320 nm) before the application of FITC suppresses the contact hypersensitivity response to FITC. Cells present in the draining lymph nodes of these mice induce suppressor T lymphocytes when injected into normal recipients. The inducer cells in the draining lymph nodes are Thy 1+, Ia- and are inactivated by gamma-radiation. These studies demonstrate that different mechanisms are involved in the in vivo activation of effector and suppressor immune responses, and they suggest that the mode of initial antigen presentation determines which immunologic circuit will be activated in response to a contact-sensitizing antigen.

  18. Comparison of gene activation by two TAL effectors from Xanthomonas axonopodis pv. manihotis reveals candidate host susceptibility genes in cassava.

    PubMed

    Cohn, Megan; Morbitzer, Robert; Lahaye, Thomas; Staskawicz, Brian J

    2016-08-01

    Xanthomonas axonopodis pv. manihotis (Xam) employs transcription activator-like (TAL) effectors to promote bacterial growth and symptom formation during infection of cassava. TAL effectors are secreted via the bacterial type III secretion system into plant cells, where they are directed to the nucleus, bind DNA in plant promoters and activate the expression of downstream genes. The DNA-binding activity of TAL effectors is carried out by a central domain which contains a series of repeat variable diresidues (RVDs) that dictate the sequence of bound nucleotides. TAL14Xam668 promotes virulence in Xam strain Xam668 and has been shown to activate multiple cassava genes. In this study, we used RNA sequencing to identify the full target repertoire of TAL14Xam668 in cassava, which includes over 50 genes. A subset of highly up-regulated genes was tested for activation by TAL14CIO151 from Xam strain CIO151. Although TAL14CIO151 and TAL14Xam668 differ by only a single RVD, they display differential activation of gene targets. TAL14CIO151 complements the TAL14Xam668 mutant defect, implying that shared target genes are important for TAL14Xam668 -mediated disease susceptibility. Complementation with closely related TAL effectors is a novel approach to the narrowing down of biologically relevant susceptibility genes of TAL effectors with multiple targets. This study provides an example of how TAL effector target activation by two strains within a single species of Xanthomonas can be dramatically affected by a small change in RVD-nucleotide affinity at a single site, and reflects the parameters of RVD-nucleotide interaction determined using designer TAL effectors in transient systems. PMID:26575863

  19. Comparison of gene activation by two TAL effectors from Xanthomonas axonopodis pv. manihotis reveals candidate host susceptibility genes in cassava.

    PubMed

    Cohn, Megan; Morbitzer, Robert; Lahaye, Thomas; Staskawicz, Brian J

    2016-08-01

    Xanthomonas axonopodis pv. manihotis (Xam) employs transcription activator-like (TAL) effectors to promote bacterial growth and symptom formation during infection of cassava. TAL effectors are secreted via the bacterial type III secretion system into plant cells, where they are directed to the nucleus, bind DNA in plant promoters and activate the expression of downstream genes. The DNA-binding activity of TAL effectors is carried out by a central domain which contains a series of repeat variable diresidues (RVDs) that dictate the sequence of bound nucleotides. TAL14Xam668 promotes virulence in Xam strain Xam668 and has been shown to activate multiple cassava genes. In this study, we used RNA sequencing to identify the full target repertoire of TAL14Xam668 in cassava, which includes over 50 genes. A subset of highly up-regulated genes was tested for activation by TAL14CIO151 from Xam strain CIO151. Although TAL14CIO151 and TAL14Xam668 differ by only a single RVD, they display differential activation of gene targets. TAL14CIO151 complements the TAL14Xam668 mutant defect, implying that shared target genes are important for TAL14Xam668 -mediated disease susceptibility. Complementation with closely related TAL effectors is a novel approach to the narrowing down of biologically relevant susceptibility genes of TAL effectors with multiple targets. This study provides an example of how TAL effector target activation by two strains within a single species of Xanthomonas can be dramatically affected by a small change in RVD-nucleotide affinity at a single site, and reflects the parameters of RVD-nucleotide interaction determined using designer TAL effectors in transient systems.

  20. Dissecting signaling through activation of specific Src-effector complexes in vivo

    PubMed Central

    Karginov, A.V.; Tsygankov, D.; Berginski, M.; Chu, P.-H.; Trudeau, E.D.; Yi, J.J.; Gomez, S.; Elston, T.C.; Hahn, K.M.

    2014-01-01

    We describe an approach to selectively activate a kinase in a specific protein complex or at a specific subcellular location within living cells, and within minutes. This reveals the effects of specific kinase pathways without time for genetic compensation. The new technique, dubbed RapRTAP (rapamycin regulated targeted activation of pathways) was used to dissect the role of Src kinase interactions with FAK and p130Cas in cell motility and morphodynamics. The overall effects of Src activation on cell morphology and adhesion dynamics were first quantified, without restricting effector access. Subsets of Src induced behaviors were then attributed to specific interactions between Src and the two downstream proteins. Activation of Src in the cytoplasm versus at the cell membrane also produced distinct phenotypes. The conserved nature of the kinase site modified for RapRTAP indicates that the technique can be applied to many kinases. PMID:24609359

  1. CD20+ T cells have a predominantly Tc1 effector memory phenotype and are expanded in the ascites of patients with ovarian cancer

    PubMed Central

    de Bruyn, Marco; Wiersma, Valerie R; Wouters, Maartje C A; Samplonius, Douwe F; Klip, Harry G; Helfrich, Wijnand; Nijman, Hans W; Eggleton, Paul; Bremer, Edwin

    2015-01-01

    Recently, a small subset of T cells that expresses the B cell marker CD20 has been identified in healthy volunteers and in patients with rheumatoid arthritis and multiple sclerosis. The origin of these CD20-positive T cells as well as their relevance in human disease remains unclear. Here, we identified that after functional B cell/T cell interaction CD20 molecules are transferred to the cell surface of T cells by trogocytosis together with the established trogocytosis marker HLA-DR. Further, the presence of CD20 on isolated CD20+ T cells remained stable for up to 48h of ex vivo culture. These CD20+ T cells almost exclusively produced IFNγ (∼70% vs. ∼20% in the CD20− T cell population) and were predominantly (CD8+) effector memory T cells (∼60–70%). This IFNγ producing and effector memory phenotype was also determined for CD20+ T cells as detected in the peripheral blood and ascitic fluids of ovarian cancer (OC) patients. In the latter, the percentage of CD20+ T cells was further strongly increased (from ∼6% in peripheral blood to 23% in ascitic fluid). Taken together, the data presented here indicate that CD20 is transferred to T cells upon intimate T cell/B cell interaction. Further, CD20+ T cells are of memory and IFNγ producing phenotype and are present in increased amounts in ascitic fluid of OC patients. PMID:26137418

  2. Distinct effector-binding sites enable synergistic transcriptional activation by BenM, a LysR-type regulator.

    PubMed

    Ezezika, Obidimma C; Haddad, Sandra; Clark, Todd J; Neidle, Ellen L; Momany, Cory

    2007-03-30

    BenM, a bacterial transcriptional regulator, responds synergistically to two effectors, benzoate and cis,cis-muconate. CatM, a paralog with overlapping function, responds only to muconate. Structures of their effector-binding domains revealed two effector-binding sites in BenM. BenM and CatM are the first LysR-type regulators to be structurally characterized while bound with physiologically relevant exogenous inducers. The effector complexes were obtained by soaking crystals with stabilizing solutions containing high effector concentrations and minimal amounts of competing ions. This strategy, including data collection with fragments of fractured crystals, may be generally applicable to related proteins. In BenM and CatM, the binding of muconate to an interdomain pocket was facilitated by helix dipoles that provide charge stabilization. In BenM, benzoate also bound in an adjacent hydrophobic region where it alters the effect of muconate bound in the primary site. A charge relay system within the BenM protein appears to underlie synergistic transcriptional activation. According to this model, Glu162 is a pivotal residue that forms salt-bridges with different arginine residues depending on the occupancy of the secondary effector-binding site. Glu162 interacts with Arg160 in the absence of benzoate and with Arg146 when benzoate is bound. This latter interaction enhances the negative charge of muconate bound to the adjacent primary effector-binding site. The redistribution of the electrostatic potential draws two domains of the protein more closely towards muconate, with the movement mediated by the dipole moments of four alpha helices. Therefore, with both effectors, BenM achieves a unique conformation capable of high level transcriptional activation.

  3. Piperine from black pepper inhibits activation-induced proliferation and effector function of T lymphocytes.

    PubMed

    Doucette, Carolyn D; Rodgers, Gemma; Liwski, Robert S; Hoskin, David W

    2015-11-01

    Piperine is a major alkaloid component of black pepper (Piper nigrum Linn), which is a widely consumed spice. Here, we investigated the effect of piperine on mouse T lymphocyte activation. Piperine inhibited polyclonal and antigen-specific T lymphocyte proliferation without affecting cell viability. Piperine also suppressed T lymphocyte entry into the S and G2 /M phases of the cell cycle, and decreased expression of G1 -associated cyclin D3, CDK4, and CDK6. In addition, piperine inhibited CD25 expression, synthesis of interferon-γ, interleukin (IL)-2, IL-4, and IL-17A, and the generation of cytotoxic effector cells. The inhibitory effect of piperine on T lymphocytes was associated with hypophosphorylation of Akt, extracellular signal-regulated kinase, and inhibitor of κBα, but not ZAP-70. The ability of piperine to inhibit several key signaling pathways involved in T lymphocyte activation and the acquisition of effector function suggests that piperine might be useful in the management of T lymphocyte-mediated autoimmune and chronic inflammatory disorders. PMID:25900378

  4. Piperine from black pepper inhibits activation-induced proliferation and effector function of T lymphocytes.

    PubMed

    Doucette, Carolyn D; Rodgers, Gemma; Liwski, Robert S; Hoskin, David W

    2015-11-01

    Piperine is a major alkaloid component of black pepper (Piper nigrum Linn), which is a widely consumed spice. Here, we investigated the effect of piperine on mouse T lymphocyte activation. Piperine inhibited polyclonal and antigen-specific T lymphocyte proliferation without affecting cell viability. Piperine also suppressed T lymphocyte entry into the S and G2 /M phases of the cell cycle, and decreased expression of G1 -associated cyclin D3, CDK4, and CDK6. In addition, piperine inhibited CD25 expression, synthesis of interferon-γ, interleukin (IL)-2, IL-4, and IL-17A, and the generation of cytotoxic effector cells. The inhibitory effect of piperine on T lymphocytes was associated with hypophosphorylation of Akt, extracellular signal-regulated kinase, and inhibitor of κBα, but not ZAP-70. The ability of piperine to inhibit several key signaling pathways involved in T lymphocyte activation and the acquisition of effector function suggests that piperine might be useful in the management of T lymphocyte-mediated autoimmune and chronic inflammatory disorders.

  5. Vaccines expressing the innate immune modulator EAT-2 elicit potent effector memory T lymphocyte responses despite pre-existing vaccine immunity.

    PubMed

    Aldhamen, Yasser Ali; Seregin, Sergey S; Schuldt, Nathaniel J; Rastall, David P W; Liu, Chyong-Jy J; Godbehere, Sarah; Amalfitano, Andrea

    2012-08-01

    The mixed results from recent vaccine clinical trials targeting HIV-1 justify the need to enhance the potency of HIV-1 vaccine platforms in general. Use of first-generation recombinant adenovirus serotype 5 (rAd5) platforms failed to protect vaccinees from HIV-1 infection. One hypothesis is that the rAd5-based vaccine failed due to the presence of pre-existing Ad5 immunity in many vaccines. We recently confirmed that EAT-2-expressing rAd5 vectors uniquely activate the innate immune system and improve cellular immune responses against rAd5-expressed Ags, inclusive of HIV/Gag. In this study, we report that use of the rAd5-EAT-2 vaccine can also induce potent cellular immune responses to HIV-1 Ags despite the presence of Ad5-specific immunity. Compared to controls expressing a mutant SH2 domain form of EAT-2, Ad5 immune mice vaccinated with an rAd5-wild-type EAT-2 HIV/Gag-specific vaccine formulation significantly facilitated the induction of several arms of the innate immune system. These responses positively correlated with an improved ability of the vaccine to induce stronger effector memory T cell-biased, cellular immune responses to a coexpressed Ag despite pre-existing anti-Ad5 immunity. Moreover, inclusion of EAT-2 in the vaccine mixture improves the generation of polyfunctional cytolytic CD8(+) T cell responses as characterized by enhanced production of IFN-γ, TNF-α, cytotoxic degranulation, and increased in vivo cytolytic activity. These data suggest a new approach whereby inclusion of EAT-2 expression in stringent human vaccination applications can provide a more effective vaccine against HIV-1 specifically in Ad5 immune subjects.

  6. TAL effectors and activation of predicted host targets distinguish Asian from African strains of the rice pathogen Xanthomonas oryzae pv. oryzicola while strict conservation suggests universal importance of five TAL effectors

    PubMed Central

    Wilkins, Katherine E.; Booher, Nicholas J.; Wang, Li; Bogdanove, Adam J.

    2015-01-01

    Xanthomonas oryzae pv. oryzicola (Xoc) causes the increasingly important disease bacterial leaf streak of rice (BLS) in part by type III delivery of repeat-rich transcription activator-like (TAL) effectors to upregulate host susceptibility genes. By pathogen whole genome, single molecule, real-time sequencing and host RNA sequencing, we compared TAL effector content and rice transcriptional responses across 10 geographically diverse Xoc strains. TAL effector content is surprisingly conserved overall, yet distinguishes Asian from African isolates. Five TAL effectors are conserved across all strains. In a prior laboratory assay in rice cv. Nipponbare, only two contributed to virulence in strain BLS256 but the strict conservation indicates all five may be important, in different rice genotypes or in the field. Concatenated and aligned, TAL effector content across strains largely reflects relationships based on housekeeping genes, suggesting predominantly vertical transmission. Rice transcriptional responses did not reflect these relationships, and on average, only 28% of genes upregulated and 22% of genes downregulated by a strain are up- and down- regulated (respectively) by all strains. However, when only known TAL effector targets were considered, the relationships resembled those of the TAL effectors. Toward identifying new targets, we used the TAL effector-DNA recognition code to predict effector binding elements in promoters of genes upregulated by each strain, but found that for every strain, all upregulated genes had at least one. Filtering with a classifier we developed previously decreases the number of predicted binding elements across the genome, suggesting that it may reduce false positives among upregulated genes. Applying this filter and eliminating genes for which upregulation did not strictly correlate with presence of the corresponding TAL effector, we generated testable numbers of candidate targets for four of the five strictly conserved TAL

  7. High quality long-term CD4+ and CD8+ effector memory populations stimulated by DNA-LACK/MVA-LACK regimen in Leishmania major BALB/c model of infection.

    PubMed

    Sánchez-Sampedro, Lucas; Gómez, Carmen Elena; Mejías-Pérez, Ernesto; Sorzano, Carlos Oscar S; Esteban, Mariano

    2012-01-01

    Heterologous vaccination based on priming with a plasmid DNA vector and boosting with an attenuated vaccinia virus MVA recombinant, with both vectors expressing the Leishmania infantum LACK antigen (DNA-LACK and MVA-LACK), has shown efficacy conferring protection in murine and canine models against cutaneus and visceral leishmaniasis, but the immune parameters of protection remain ill defined. Here we performed by flow cytometry an in depth analysis of the T cell populations induced in BALB/c mice during the vaccination protocol DNA-LACK/MVA-LACK, as well as after challenge with L. major parasites. In the adaptive response, there is a polyfunctional CD4(+) and CD8(+) T cell activation against LACK antigen. At the memory phase the heterologous vaccination induces high quality LACK-specific long-term CD4(+) and CD8(+) effector memory cells. After parasite challenge, there is a moderate boosting of LACK-specific CD4(+) and CD8(+) T cells. Anti-vector responses were largely CD8(+)-mediated. The immune parameters induced against LACK and triggered by the combined vaccination DNA/MVA protocol, like polyfunctionality of CD4(+) and CD8(+) T cells with an effector phenotype, could be relevant in protection against leishmaniasis. PMID:22715418

  8. Histological improvement in salivary gland along with effector memory Th17-1 cell reduction in a primary Sjogren's syndrome patient with dermatomyositis and diffuse large B-cell lymphoma by R-CHOP therapy.

    PubMed

    Koga, Tomohiro; Mizokami, Akinari; Nakashima, Masahiro; Shimizu, Toshimasa; Nakashima, Yoshikazu; Nakamura, Hideki; Chiwata, Masahiko; Daisuke, Niino; Kawakami, Atsushi

    2016-04-01

    We treated a 45-year-old Japanese woman with primary Sjogren's syndrome (SS) complicated with dermatomyositis (DM) followed by diffuse large B-cell lymphoma. She was admitted to our hospital for further evaluation of fever, weight loss and peritoneal lymphadenopathy. The histological examination of her lymph node revealed diffuse large B-cell lymphoma. The patient was then treated with 8 cycles of R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) therapy, which resulted in histological and functional improvement of her salivary glands. Of note, the percentage of effector memory Th17-1 (CD3+CD4+CD45RA-CCR7-CXCR3+CCR6+) cells in the peripheral blood was decreased after the R-CHOP treatment. This case suggests that an altered Th17-1 cell subset by B-cell depletion therapy is critical for the improvement of tissue damage in patients with SS, and the case suggests that clinicians should consider measuring the effector memory Th-subsets to predict the disease activity in SS patients. PMID:26960952

  9. Ex vivo culture of chimeric antigen receptor T cells generates functional CD8+ T cells with effector and central memory-like phenotype.

    PubMed

    Neeson, P; Shin, A; Tainton, K M; Guru, P; Prince, H M; Harrison, S J; Peinert, S; Smyth, M J; Trapani, J A; Kershaw, M H; Darcy, P K; Ritchie, D S

    2010-09-01

    The anti-tumor efficacy of adoptively transferred T cells requires their in vivo persistence and memory polarization. It is unknown if human chimeric antigen receptor (CAR)-expressing T cells can also undergo memory polarization. We examined the functional status of CAR CD8(+) T cells, re-directed to Lewis Y antigen (LeY-T), throughout a period of ex vivo expansion. Immediately before culture CD8(+) T cells comprised a mixture of phenotypes including naive (CD45RA(+)/CCR7(+)/CD27(+)/CD28(+)/perforin-), central memory (CM, CD45RA(-)/CCR7(lo)/CD27(+)/CD28(+)/perforin(lo)), effector memory (EM, CD45RA(-)/CCR7(-)/CD27(+)/CD28(+)/perforin(mod)) and effector (Eff, CD45RA(+)/CCR7(-)/CD27(-)/CD28(-)/perforin(hi)) cells. After transduction and expansion culture of peripheral blood mononuclear cells from normal donors or multiple myeloma patients, CD8(+) LeY-T cells polarized to EM- and CM-like phenotype. CD8(+) LeY-T cells differed from starting CD8(+) CM and EM T cells in that CD27, but not CD28, was downregulated. In addition, CD8(+) LeY-T cells expressed high levels of perforin, similar to starting CD8(+) Eff. CD8(+) LeY-T cells also showed hallmarks of both memory and Eff function, underwent homeostatic proliferation in response to interleukin (IL)-15, and showed interferon (IFN)-γ production and cytotoxicity in response to Le-Y antigen on OVCAR-3 (human ovarian adenocarcinoma) cells. This study confirms CD8(+) LeY-T cells have a CM- and EM-like phenotype and heterogeneous function consistent with potential to persist in vivo after adoptive transfer.

  10. GITR ligand-costimulation activates effector and regulatory functions of CD4{sup +} T cells

    SciTech Connect

    Igarashi, Hanna; Cao, Yujia; Iwai, Hideyuki; Piao, Jinhua; Kamimura, Yosuke; Hashiguchi, Masaaki; Amagasa, Teruo; Azuma, Miyuki

    2008-05-16

    Engagement of glucocorticoid-induced TNFR-related protein (GITR) enables the costimulation of both CD25{sup -}CD4{sup +} effector (Teff) and CD25{sup +}CD4{sup +} regulatory (Treg) cells; however, the effects of GITR-costimulation on Treg function remain controversial. In this study, we examined the effects of GITR ligand (GITRL) binding on the respective functions of CD4{sup +} T cells. GITRL-P815 transfectants efficiently augmented anti-CD3-induced proliferation and cytokine production by Teff cells. Proliferation and IL-10 production in Treg were also enhanced by GITRL transfectants when exogenous IL-2 and stronger CD3 stimulation was provided. Concomitant GITRL-costimulation of Teff and Treg converted the anergic state of Treg into a proliferating state, maintaining and augmenting their function. Thus, GITRL-costimulation augments both effector and regulatory functions of CD4{sup +} T cells. Our results suggest that highly activated and increased ratios of Treg reverse the immune-enhancing effects of GITRL-costimulation in Teff, which may be problematic for therapeutic applications using strong GITR agonists.

  11. The Ustilago maydis effector Pep1 suppresses plant immunity by inhibition of host peroxidase activity.

    PubMed

    Hemetsberger, Christoph; Herrberger, Christian; Zechmann, Bernd; Hillmer, Morten; Doehlemann, Gunther

    2012-01-01

    The corn smut Ustilago maydis establishes a biotrophic interaction with its host plant maize. This interaction requires efficient suppression of plant immune responses, which is attributed to secreted effector proteins. Previously we identified Pep1 (Protein essential during penetration-1) as a secreted effector with an essential role for U. maydis virulence. pep1 deletion mutants induce strong defense responses leading to an early block in pathogenic development of the fungus. Using cytological and functional assays we show that Pep1 functions as an inhibitor of plant peroxidases. At sites of Δpep1 mutant penetrations, H₂O₂ strongly accumulated in the cell walls, coinciding with a transcriptional induction of the secreted maize peroxidase POX12. Pep1 protein effectively inhibited the peroxidase driven oxidative burst and thereby suppresses the early immune responses of maize. Moreover, Pep1 directly inhibits peroxidases in vitro in a concentration-dependent manner. Using fluorescence complementation assays, we observed a direct interaction of Pep1 and the maize peroxidase POX12 in vivo. Functional relevance of this interaction was demonstrated by partial complementation of the Δpep1 mutant defect by virus induced gene silencing of maize POX12. We conclude that Pep1 acts as a potent suppressor of early plant defenses by inhibition of peroxidase activity. Thus, it represents a novel strategy for establishing a biotrophic interaction.

  12. Cytotoxic minor histocompatibility antigen HA-1-specific CD8+ effector memory T cells: artificial APCs pave the way for clinical application by potent primary in vitro induction.

    PubMed

    Schilbach, Karin; Kerst, Gunter; Walter, Steffen; Eyrich, Matthias; Wernet, Dorothee; Handgretinger, Rupert; Xie, Weidong; Rammensee, Hans-Georg; Müller, Ingo; Bühring, Hans-Jörg; Niethammer, Dietrich

    2005-07-01

    Induction of cytotoxic T lymphocytes (CTLs) for treatment of relapsed leukemia after allogeneic stem-cell transplantation is hindered by the laborious and time-consuming procedure of generating dendritic cells for antigen presentation. Artificial antigen-presenting cells (aAPCs) offer the advantage of being readily available in sufficient numbers, thus allowing for a highly standardized in vitro induction of CTLs. We generated aAPCs coated with anti-CD28 antibody (Ab) and either high-density (HD) or low-density (LD) major histocompatibility complex (MHC) class I molecules loaded with HA-1(H), a nonapeptide derived from the hematopoiesis-restricted minor histocompatibility antigen HA-1. HD- and LD-aAPCs potently induced HA-1(H)-specific CD8+ CTLs from untouched CD8+ T cells of healthy donors. CTLs were subsequently purified by magnetic-activated cell sorting. HD- as well as LD-aAPC-induced CTLs exerted high HA-1H-specific cytotoxicity, resembled T(c)1 effector memory cells, survived a long time in vitro, and were expanded by a factor varying between 8.2 x 10(4) and 51 x 10(4). The T-cell receptor (TCR) repertoire of HA-1H tetramer-positive CTLs was oligoclonal with a prominent usage of Vbeta6. The TCR repertoire of tetramer-positive CTLs was distinct from and more restricted than that of tetramer-negative cells. These findings indicate that aAPCs are attractive tools for the ex vivo generation of HA-1H-specific CTLs suitable for immunotherapy of relapsed leukemia. PMID:15731181

  13. Relevance of long-lived CD8+ T effector memory cells for protective immunity elicited by heterologous prime-boost vaccination

    PubMed Central

    Vasconcelos, José R.; Dominguez, Mariana R.; Araújo, Adriano F.; Ersching, Jonatan; Tararam, Cibele A.; Bruna-Romero, Oscar; Rodrigues, Mauricio M.

    2012-01-01

    Owing to the importance of major histocompatibility complex class Ia-restricted CD8+ T cells for host survival following viral, bacterial, fungal, or parasitic infection, it has become largely accepted that these cells should be considered in the design of a new generation of vaccines. For the past 20 years, solid evidence has been provided that the heterologous prime-boost regimen achieves the best results in terms of induction of long-lived protective CD8+ T cells against a variety of experimental infections. Although this regimen has often been used experimentally, as is the case for many vaccines, the mechanism behind the efficacy of this vaccination regimen is still largely unknown. The main purpose of this review is to examine the characteristics of the protective CD8+ T cells generated by this vaccination regimen. Part of its efficacy certainly relies on the generation and maintenance of large numbers of specific lymphocytes. Other specific characteristics may also be important, and studies on this direction have only recently been initiated. So far, the characterization of these protective, long-lived T cell populations suggests that there is a high frequency of polyfunctional T cells; these cells cover a large breadth and display a T effector memory (TEM) phenotype. These TEM cells are capable of proliferating after an infectious challenge and are highly refractory to apoptosis due to a control of the expression of pro-apoptotic receptors such as CD95. Also, they do not undergo significant long-term immunological erosion. Understanding the mechanisms that control the generation and maintenance of the protective activity of these long-lived TEM cells will certainly provide important insights into the physiology of CD8+ T cells and pave the way for the design of new or improved vaccines. PMID:23264773

  14. A transcription activator-like effector (TALE) induction system mediated by proteolysis.

    PubMed

    Copeland, Matthew F; Politz, Mark C; Johnson, Charles B; Markley, Andrew L; Pfleger, Brian F

    2016-04-01

    Simple and predictable trans-acting regulatory tools are needed in the fields of synthetic biology and metabolic engineering to build complex genetic circuits and optimize the levels of native and heterologous gene products. Transcription activator-like effectors (TALEs) are bacterial virulence factors that have recently gained traction in biotechnology applications owing to their customizable DNA-binding specificity. In this work we expanded the versatility of these transcription factors to create an inducible TALE system by inserting tobacco-etch virus (TEV) protease recognition sites into the TALE backbone. The resulting engineered TALEs maintain transcriptional repression of their target genes in Escherichia coli, but are degraded after induction of the TEV protease, thereby promoting expression of the previously repressed target gene of interest. This TALE-TEV technology enables both repression and induction of plasmid or chromosomal target genes in a manner analogous to traditional repressor proteins but with the added flexibility of being operator-agnostic.

  15. A multiplexed transcription activator-like effector system for detecting specific DNA sequences.

    PubMed

    Honarmand, Ali; Mayall, Robert; George, Iain; Oberding, Lisa; Dastidar, Himika; Fegan, Jamie; Chaudhuri, Somshukla; Dole, Justin; Feng, Sharon; Hoang, Denny; Moges, Ruth; Osgood, Julie; Remondini, Taylor; van der Meulen, Wm Keith; Wang, Su; Wintersinger, Chris; Zaparoli Zucoloto, Amanda; Chatfield-Reed, Kate; Arcellana-Panlilio, Mayi; Nygren, Anders

    2014-12-19

    Transcription activator-like effectors (TALEs), originating from the Xanthomonas genus of bacteria, bind to specific DNA sequences based on amino acid sequence in the repeat-variable diresidue (RVD) positions of the protein. By altering these RVDs, it has been shown that a TALE protein can be engineered to bind virtually any DNA sequence of interest. The possibility of multiplexing TALEs for the purposes of identifying specific DNA sequences has yet to be explored. Here, we demonstrate a system in which a TALE protein bound to a nitrocellulose strip has been utilized to capture purified DNA, which is then detected using the binding of a second distinct TALE protein conjugated to a protein tag that is then detected by a dot blot. This system provides a signal only when both TALEs bind to their respective sequences, further demonstrating the specificity of the TALE binding.

  16. A transcription activator-like effector (TALE) induction system mediated by proteolysis.

    PubMed

    Copeland, Matthew F; Politz, Mark C; Johnson, Charles B; Markley, Andrew L; Pfleger, Brian F

    2016-04-01

    Simple and predictable trans-acting regulatory tools are needed in the fields of synthetic biology and metabolic engineering to build complex genetic circuits and optimize the levels of native and heterologous gene products. Transcription activator-like effectors (TALEs) are bacterial virulence factors that have recently gained traction in biotechnology applications owing to their customizable DNA-binding specificity. In this work we expanded the versatility of these transcription factors to create an inducible TALE system by inserting tobacco-etch virus (TEV) protease recognition sites into the TALE backbone. The resulting engineered TALEs maintain transcriptional repression of their target genes in Escherichia coli, but are degraded after induction of the TEV protease, thereby promoting expression of the previously repressed target gene of interest. This TALE-TEV technology enables both repression and induction of plasmid or chromosomal target genes in a manner analogous to traditional repressor proteins but with the added flexibility of being operator-agnostic. PMID:26854666

  17. A multiplexed transcription activator-like effector system for detecting specific DNA sequences.

    PubMed

    Honarmand, Ali; Mayall, Robert; George, Iain; Oberding, Lisa; Dastidar, Himika; Fegan, Jamie; Chaudhuri, Somshukla; Dole, Justin; Feng, Sharon; Hoang, Denny; Moges, Ruth; Osgood, Julie; Remondini, Taylor; van der Meulen, Wm Keith; Wang, Su; Wintersinger, Chris; Zaparoli Zucoloto, Amanda; Chatfield-Reed, Kate; Arcellana-Panlilio, Mayi; Nygren, Anders

    2014-12-19

    Transcription activator-like effectors (TALEs), originating from the Xanthomonas genus of bacteria, bind to specific DNA sequences based on amino acid sequence in the repeat-variable diresidue (RVD) positions of the protein. By altering these RVDs, it has been shown that a TALE protein can be engineered to bind virtually any DNA sequence of interest. The possibility of multiplexing TALEs for the purposes of identifying specific DNA sequences has yet to be explored. Here, we demonstrate a system in which a TALE protein bound to a nitrocellulose strip has been utilized to capture purified DNA, which is then detected using the binding of a second distinct TALE protein conjugated to a protein tag that is then detected by a dot blot. This system provides a signal only when both TALEs bind to their respective sequences, further demonstrating the specificity of the TALE binding. PMID:25524096

  18. A transcription activator-like effector induction system mediated by proteolysis

    PubMed Central

    Copeland, Matthew F.; Politz, Mark C.; Johnson, Charles B.; Markley, Andrew L.; Pfleger, Brian F.

    2016-01-01

    Simple and predictable trans-acting regulatory tools are needed in the fields of synthetic biology and metabolic engineering to build complex genetic circuits and optimize the levels of native and heterologous gene products. Transcription activator-like effectors (TALEs) are bacterial virulence factors that have recently gained traction in biotechnology applications due to their customizable DNA binding specificity. In this work we expand the versatility of these transcription factors to create an inducible TALE system by inserting tobacco-etch virus (TEV) protease recognition sites into the TALE backbone. The resulting engineered TALEs maintain transcriptional repression of their target genes in Escherichia coli, but are degraded following the induction of the TEV protease, thereby promoting expression of the previously repressed target gene of interest. This TALE-TEV technology enables both repression and induction of plasmid or chromosomal target genes in a manner analogous to traditional repressor proteins but with the added flexibility of being operator agnostic. PMID:26854666

  19. STAT3 signaling contributes to the high effector activities of interleukin-15-derived dendritic cells

    PubMed Central

    Okada, Starlyn; Han, Shuhong; Patel, Ekta S; Yang, Li-Jun; Chang, Lung-Ji

    2015-01-01

    Dendritic cells (DCs) are important innate and adaptive immune effectors, and have a key role in antigen presentation and T-cell activation. Different lineages of DCs can be developed from hematopoietic progenitors following cytokine signaling, and the various lineages of DCs display distinct morphology, phenotype and functions. There has been limited information on differential cytokine-mediated molecular signaling in DCs. Analyses of surface molecules by flow cytometry and quantitative RNA profiling revealed differences between DCs derived from interleukin-4 (IL-4) versus IL-15 signaling, yet both lineages of DCs exhibited similar levels of surface molecules key to immune activation. Functional assays confirmed that IL-15-derived DCs elicited greater antigen-specific, primary and secondary CD8 and CD4 T-cell responses than did IL-4-derived DCs. Importantly, IL-15 DCs secreted substantial amounts of proinflammatory cytokines, including IL-6, interferon-γ (IFN-γ) and tumor necrosis factor-α (TNFα), which helped polarize a strong T-cell response. Assessment of signaling pathways revealed that IL-15 DCs exhibited a lower levels of activated signal transducer and activator of transcription 5 (STAT5), STAT6 and extracellular signal-regulated kinase 1/2 than IL-4 DCs, but after lipopolysaccharide (LPS)/TNFα treatment, the STAT3 and p38 mitogen-activated protein kinase (MAPK) activities were significantly enhanced in the IL-15 DCs. Surprisingly, contrary to the canonical IL-15-mediated STAT5 signaling pathway in lymphoid cells, IL-15 did not mediate a strong STAT5 or STAT3 activation in DCs. Further analysis using specific inhibitors to STAT3 and p38 MAPK pathways revealed that the STAT3 signaling, but not p38 MAPK signaling, contributed to IFN-γ production in DCs. Therefore, while IL-15 does not promote the STAT signaling in DCs, the increased STAT3 activity after LPS/TNFα treatment of the IL-15 DCs has a key role in their high IFN-γ effector activities. PMID

  20. Malaria-associated atypical memory B cells exhibit markedly reduced B cell receptor signaling and effector function.

    PubMed

    Portugal, Silvia; Tipton, Christopher M; Sohn, Haewon; Kone, Younoussou; Wang, Jing; Li, Shanping; Skinner, Jeff; Virtaneva, Kimmo; Sturdevant, Daniel E; Porcella, Stephen F; Doumbo, Ogobara K; Doumbo, Safiatou; Kayentao, Kassoum; Ongoiba, Aissata; Traore, Boubacar; Sanz, Inaki; Pierce, Susan K; Crompton, Peter D

    2015-05-08

    Protective antibodies in Plasmodium falciparum malaria are only acquired after years of repeated infections. Chronic malaria exposure is associated with a large increase in atypical memory B cells (MBCs) that resemble B cells expanded in a variety of persistent viral infections. Understanding the function of atypical MBCs and their relationship to classical MBCs will be critical to developing effective vaccines for malaria and other chronic infections. We show that VH gene repertoires and somatic hypermutation rates of atypical and classical MBCs are indistinguishable indicating a common developmental history. Atypical MBCs express an array of inhibitory receptors and B cell receptor (BCR) signaling is stunted in atypical MBCs resulting in impaired B cell responses including proliferation, cytokine production and antibody secretion. Thus, in response to chronic malaria exposure, atypical MBCs appear to differentiate from classical MBCs becoming refractory to BCR-mediated activation and potentially interfering with the acquisition of malaria immunity.

  1. Locked and proteolysis-based transcription activator-like effector (TALE) regulation.

    PubMed

    Lonzarić, Jan; Lebar, Tina; Majerle, Andreja; Manček-Keber, Mateja; Jerala, Roman

    2016-02-18

    Development of orthogonal, designable and adjustable transcriptional regulators is an important goal of synthetic biology. Their activity has been typically modulated through stimulus-induced oligomerization or interaction between the DNA-binding and activation/repression domain. We exploited a feature of the designable Transcription activator-like effector (TALE) DNA-binding domain that it winds around the DNA which allows to topologically prevent it from binding by intramolecular cyclization. This new approach was investigated through noncovalent ligand-induced cyclization or through a covalent split intein cyclization strategy, where the topological inhibition of DNA binding by cyclization and its restoration by a proteolytic release of the topologic constraint was expected. We show that locked TALEs indeed have diminished DNA binding and regain full transcriptional activity by stimulation with the rapamycin ligand or site-specific proteolysis of the peptide linker, with much higher level of activation than rapamycin-induced heterodimerization. Additionally, we demonstrated reversibility, activation of genomic targets and implemented logic gates based on combinations of protein cyclization, proteolytic cleavage and ligand-induced dimerization, where the strongest fold induction was achieved by the proteolytic cleavage of a repression domain from a linear TALE.

  2. Inhibition of inflammasome activation by Coxiella burnetii type IV secretion system effector IcaA

    PubMed Central

    Cunha, Larissa D.; Ribeiro, Juliana M.; Fernandes, Talita D.; Massis, Liliana M.; Khoo, Chen Ai; Moffatt, Jennifer H.; Newton, Hayley J.; Roy, Craig R.; Zamboni, Dario S.

    2015-01-01

    Coxiella burnetii is a highly infectious bacterium that promotes its own replication in macrophages by inhibiting several host cell responses. Here, we show that C. burnetii inhibits caspase-1 activation in primary mouse macrophages. By using co-infection experiments, we determine that the infection of macrophages with C. burnetii inhibits the caspase-11-mediated non-canonical activation of the NLRP3 inflammasome induced by subsequent infection with Escherichia coli or Legionella pneumophila. Genetic screening using flagellin mutants of L. pneumophila as a surrogate host, reveals a novel C. burnetii gene (IcaA) involved in the inhibition of caspase activation. Expression of IcaA in L. pneumophila inhibited the caspase-11 activation in macrophages. Moreover, icaA- mutants of C. burnetii failed to suppress the caspase-11-mediated inflammasome activation induced by L. pneumophila. Our data reveal IcaA as a novel C. burnetii effector protein that is secreted by the Dot/Icm type IV secretion system and interferes with the caspase-11-induced, non-canonical activation of the inflammasome. PMID:26687278

  3. Locked and proteolysis-based transcription activator-like effector (TALE) regulation.

    PubMed

    Lonzarić, Jan; Lebar, Tina; Majerle, Andreja; Manček-Keber, Mateja; Jerala, Roman

    2016-02-18

    Development of orthogonal, designable and adjustable transcriptional regulators is an important goal of synthetic biology. Their activity has been typically modulated through stimulus-induced oligomerization or interaction between the DNA-binding and activation/repression domain. We exploited a feature of the designable Transcription activator-like effector (TALE) DNA-binding domain that it winds around the DNA which allows to topologically prevent it from binding by intramolecular cyclization. This new approach was investigated through noncovalent ligand-induced cyclization or through a covalent split intein cyclization strategy, where the topological inhibition of DNA binding by cyclization and its restoration by a proteolytic release of the topologic constraint was expected. We show that locked TALEs indeed have diminished DNA binding and regain full transcriptional activity by stimulation with the rapamycin ligand or site-specific proteolysis of the peptide linker, with much higher level of activation than rapamycin-induced heterodimerization. Additionally, we demonstrated reversibility, activation of genomic targets and implemented logic gates based on combinations of protein cyclization, proteolytic cleavage and ligand-induced dimerization, where the strongest fold induction was achieved by the proteolytic cleavage of a repression domain from a linear TALE. PMID:26748097

  4. Ultrahigh Enzyme Activity Assembled in Layered Double Hydroxides via Mg(2+)-Allosteric Effector.

    PubMed

    Wang, Min; Huang, Shu-Wan; Xu, Dan; Bao, Wen-Jing; Xia, Xing-Hua

    2015-06-01

    It is well-known that some metal ions could be allosteric effectors of allosteric enzymes to activate/inhibit the catalytic activities of enzymes. In nanobiocatalytic systems constructed based on the positive metal ion-induced allosteric effect, the incorporated enzymes will be activated and thus exhibit excellent catalytic performance. Herein, we present an environmentally friendly strategy to construct a novel allosteric effect-based β-galactosidase/Mg-Al layered double hydroxide (β-gal/Mg-Al-LDH) nanobiocatalytic system via the delamination-reconstruction method. The intercalated β-gal in the LDH galleries changes its conformation significantly due to the Mg(2+)-induced allosteric interactions and other weak interactions, which causes the activation of enzymatic activity. The β-gal/Mg-Al-LDH nanobiocatalytic system shows much higher catalytic activity and affinity toward its substrate and about 30 times higher catalytic reaction velocity than the free β-gal, which suggests that Mg(2+)-induced allosteric effect plays a vital role in the improvement of enzymatic performance.

  5. ICOS is required for the generation of both central and effector CD4(+) memory T-cell populations following acute bacterial infection.

    PubMed

    Marriott, Clare L; Carlesso, Gianluca; Herbst, Ronald; Withers, David R

    2015-06-01

    Interactions between ICOS and ICOS ligand (ICOSL) are essential for the development of T follicular helper (Tfh) cells and thus the formation and maintenance of GC reactions. Given the conflicting reports on the requirement of other CD4(+) T-cell populations for ICOS signals, we have employed a range of in vivo approaches to dissect requirements for ICOS signals in mice during an endogenous CD4(+) T-cell response and contrasted this with CD28 signals. Genetic absence of ICOSL only modestly reduced the total number of antigen-specific CD4(+) T cells at the peak of the primary response, but resulted in a severely diminished number of both T central memory and T effector memory cells. Treatment with blocking anti-ICOS mAb during the primary response recapitulated these effects and caused a more substantial reduction than blocking CD28 signals with CTLA4Ig. During the memory phase of the response further signals through ICOS or CD28 were not required for survival. However, upon secondary challenge only Tfh cell expansion remained heavily ICOS-dependent, while CD28 signals were required for optimal expansion of all subsets. These data demonstrate the importance of ICOS signals specifically for memory CD4(+) T-cell formation, while highlighting the potential of therapeutically targeting this pathway.

  6. ICOS is required for the generation of both central and effector CD4+ memory T‐cell populations following acute bacterial infection

    PubMed Central

    Marriott, Clare L.; Carlesso, Gianluca; Herbst, Ronald

    2015-01-01

    Interactions between ICOS and ICOS ligand (ICOSL) are essential for the development of T follicular helper (Tfh) cells and thus the formation and maintenance of GC reactions. Given the conflicting reports on the requirement of other CD4+ T‐cell populations for ICOS signals, we have employed a range of in vivo approaches to dissect requirements for ICOS signals in mice during an endogenous CD4+ T‐cell response and contrasted this with CD28 signals. Genetic absence of ICOSL only modestly reduced the total number of antigen‐specific CD4+ T cells at the peak of the primary response, but resulted in a severely diminished number of both T central memory and T effector memory cells. Treatment with blocking anti‐ICOS mAb during the primary response recapitulated these effects and caused a more substantial reduction than blocking CD28 signals with CTLA4Ig. During the memory phase of the response further signals through ICOS or CD28 were not required for survival. However, upon secondary challenge only Tfh cell expansion remained heavily ICOS‐dependent, while CD28 signals were required for optimal expansion of all subsets. These data demonstrate the importance of ICOS signals specifically for memory CD4+ T‐cell formation, while highlighting the potential of therapeutically targeting this pathway. PMID:25754933

  7. RipAY, a Plant Pathogen Effector Protein, Exhibits Robust γ-Glutamyl Cyclotransferase Activity When Stimulated by Eukaryotic Thioredoxins.

    PubMed

    Fujiwara, Shoko; Kawazoe, Tomoki; Ohnishi, Kouhei; Kitagawa, Takao; Popa, Crina; Valls, Marc; Genin, Stéphane; Nakamura, Kazuyuki; Kuramitsu, Yasuhiro; Tanaka, Naotaka; Tabuchi, Mitsuaki

    2016-03-25

    The plant pathogenic bacterium Ralstonia solanacearum injects more than 70 effector proteins (virulence factors) into the host plant cells via the needle-like structure of a type III secretion system. The type III secretion system effector proteins manipulate host regulatory networks to suppress defense responses with diverse molecular activities. Uncovering the molecular function of these effectors is essential for a mechanistic understanding of R. solanacearum pathogenicity. However, few of the effectors from R. solanacearum have been functionally characterized, and their plant targets remain largely unknown. Here, we show that the ChaC domain-containing effector RipAY/RSp1022 from R. solanacearum exhibits γ-glutamyl cyclotransferase (GGCT) activity to degrade the major intracellular redox buffer, glutathione. Heterologous expression of RipAY, but not other ChaC family proteins conserved in various organisms, caused growth inhibition of yeast Saccharomyces cerevisiae, and the intracellular glutathione level was decreased to ∼30% of the normal level following expression of RipAY in yeast. Although active site mutants of GGCT activity were non-toxic, the addition of glutathione did not reverse the toxicity, suggesting that the toxicity might be a consequence of activity against other γ-glutamyl compounds. Intriguingly, RipAY protein purified from a bacterial expression system did not exhibit any GGCT activity, whereas it exhibited robust GGCT activity upon its interaction with eukaryotic thioredoxins, which are important for intracellular redox homeostasis during bacterial infection in plants. Our results suggest that RipAY has evolved to sense the host intracellular redox environment, which triggers its enzymatic activity to create a favorable environment for R. solanacearum infection. PMID:26823466

  8. Draft genome sequence of Xanthomonas axonopodis pv. glycines 8ra possessing transcription activator-like effectors used for genetic engineering.

    PubMed

    Lee, Ju-Hoon; Shin, Hakdong; Park, Hye-Jee; Ryu, Sangryeol; Han, Sang-Wook

    2014-06-10

    Xanthomonas axonopodis pv. glycines 8ra is a causal agent of bacterial pustule disease in soybean. This bacterium possesses transcription activator-like (TAL) effectors which are useful for genetic/protein engineering applications in higher organisms including plants and humans. Here, we report that the draft genome sequence consists of 5,337,885-bp double-stranded DNA encoding 4674 open reading frames (ORFs) in 13 different contigs. This genome sequence would be useful in applications of TAL effectors in genetic engineering and in elucidating virulence factors against plants. PMID:24657734

  9. Structural and Functional Studies Indicate That the EPEC Effector, EspG, Directly Binds p21-Activated Kinase

    SciTech Connect

    Germane, Katherine L.; Spiller, Benjamin W.

    2011-09-20

    Bacterial pathogens secrete effectors into their hosts that subvert host defenses and redirect host processes. EspG is a type three secretion effector with a disputed function that is found in enteropathogenic Escherichia coli. Here we show that EspG is structurally similar to VirA, a Shigella virulence factor; EspG has a large, conserved pocket on its surface; EspG binds directly to the amino-terminal inhibitory domain of human p21-activated kinase (PAK); and mutations to conserved residues in the surface pocket disrupt the interaction with PAK.

  10. SfDredd, a Novel Initiator Caspase Possessing Activity on Effector Caspase Substrates in Spodoptera frugiperda

    PubMed Central

    Liu, Hao; Wu, Andong; Mei, Long; Liu, Qingzhen

    2016-01-01

    Sf9, a cell line derived from Spodoptera frugiperda, is an ideal model organism for studying insect apoptosis. The first notable study that attempted to identify the apoptotic pathway in Sf9 was performed in 1997 and included the discovery of Sf-caspase-1, an effector caspase of Sf9. However, it was not until 2013 that the first initiator caspase in Sf9, SfDronc, was discovered, and the apoptotic pathway in Sf9 became clearer. In this study, we report another caspase of Sf9, SfDredd. SfDredd is highly similar to insect initiator caspase Dredd homologs. Experimentally, recombinant SfDredd underwent autocleavage and exhibited different efficiencies in cleavage of synthetic caspase substrates. This was attributed to its caspase activity for the predicted active site mutation blocked the above autocleavage and synthetic caspase substrates cleavage activity. SfDredd was capable of not only cleaving Sf-caspase-1 in vitro but also cleaving Sf-caspase-1 and inducing apoptosis when it was co-expressed with Sf-caspase-1 in Sf9 cells. The protein level of SfDredd was increased when Sf9 cells were treated by Actinomycin D, whereas silencing of SfDredd reduced apoptosis and Sf-caspase-1 cleavage induced by Actinomycin D treatment. These results clearly indicate that SfDredd functioned as an apoptotic initiator caspase. Apoptosis induced in Sf9 cells by overexpression of SfDredd alone was not as obvious as that induced by SfDronc alone, and the cleavage sites of Sf-caspase-1 for SfDredd and SfDronc are different. In addition, despite sharing a sequence homology with initiator caspases and possessing weak activity on initiator caspase substrates, SfDredd showed strong activity on effector caspase substrates, making it the only insect caspase reported so far functioning similar to human caspase-2 in this aspect. We believe that the discovery of SfDredd, and its different properties from SfDronc, will improve the understanding of apoptosis pathway in Sf9 cells. PMID:26977926

  11. Activated mast cells promote differentiation of B cells into effector cells

    PubMed Central

    Palm, Anna-Karin E.; Garcia-Faroldi, Gianni; Lundberg, Marcus; Pejler, Gunnar; Kleinau, Sandra

    2016-01-01

    Based on the known accumulation of mast cells (MCs) in B cell-dependent inflammatory diseases, including rheumatoid arthritis, we hypothesized that MCs directly modulate B cells. We show here that degranulated, and to a lesser extent naïve or IgE-sensitized, MCs activate both naïve and B cell receptor-activated B cells. This was shown by increased proliferation, blast formation, and expression of CD19, MHC class II and CD86 in the B cells. Further, MCs stimulated the secretion of IgM and IgG in IgM+ B cells, indicating that MCs can induce class-switch recombination in B cells. We also show that coculture of MCs with B cells promotes surface expression of L-selectin, a homing receptor, on the B cells. The effects of MCs on B cells were partly dependent on cell-cell contact and both follicular and marginal zone B cells could be activated by MCs. Our findings suggest that degranulated MCs support optimal activation of B cells, a finding that is in line with in vivo studies showing that MCs frequently degranulate in the context of B-cell driven pathologies such as arthritis. Together, our findings show that MCs have the capacity to differentiate B cells to effector cells. PMID:26847186

  12. Urban Particulate Matter-Activated Human Dendritic Cells Induce the Expansion of Potent Inflammatory Th1, Th2, and Th17 Effector Cells.

    PubMed

    Matthews, Nick C; Pfeffer, Paul E; Mann, Elizabeth H; Kelly, Frank J; Corrigan, Christopher J; Hawrylowicz, Catherine M; Lee, Tak H

    2016-02-01

    Exposure to urban particulate matter (UPM) exacerbates asthmatic lung inflammation. Lung dendritic cells (DCs) are critical for stimulating T cell immunity and in maintaining airway tolerance, but they also react to airway UPM. The adjuvant role of UPM in enhancing primary immune responses by naive cells to allergen has been reported, but the direct effects of UPM-activated DCs on the functionality of human memory CD4 T cells (Tms), which constitute the majority of T cells in the lung, has not been investigated. Blood CD1c(+) DCs were purified and activated with UPM in the presence or absence of house dust mite or tetanus toxoid control antigen. 5-(and -6)-Carboxyfluorescein diacetate succinimidyl ester-labeled blood Tms were cocultured with autologous DCs, T cell proliferation and effector function were assessed using flow cytometry, and secreted cytokines were measured by combined bead array. UPM-DCs elicited IFN-γ and IL-13 secretion and induced proliferation in Tms isolated from both allergic patients with asthma and healthy control subjects, whereas only IL-13 was produced by Tms from patients with atopic asthma stimulated by house dust mite-loaded DCs. UPM-DCs drove the expansion and differentiation of a mixed population of Th1, Th2, and Th17 cell effectors through a mechanism that was dependent on major histocompatibility class II but not on cytokine-driven expansion. The data suggest that UPM not only has adjuvant properties but is also a source of antigen that stimulates the generation of Th2, Th1, and Th17 effector phenotypes, which have been implicated in both exacerbations of asthma and chronic inflammatory diseases. PMID:26196219

  13. Urban Particulate Matter-Activated Human Dendritic Cells Induce the Expansion of Potent Inflammatory Th1, Th2, and Th17 Effector Cells.

    PubMed

    Matthews, Nick C; Pfeffer, Paul E; Mann, Elizabeth H; Kelly, Frank J; Corrigan, Christopher J; Hawrylowicz, Catherine M; Lee, Tak H

    2016-02-01

    Exposure to urban particulate matter (UPM) exacerbates asthmatic lung inflammation. Lung dendritic cells (DCs) are critical for stimulating T cell immunity and in maintaining airway tolerance, but they also react to airway UPM. The adjuvant role of UPM in enhancing primary immune responses by naive cells to allergen has been reported, but the direct effects of UPM-activated DCs on the functionality of human memory CD4 T cells (Tms), which constitute the majority of T cells in the lung, has not been investigated. Blood CD1c(+) DCs were purified and activated with UPM in the presence or absence of house dust mite or tetanus toxoid control antigen. 5-(and -6)-Carboxyfluorescein diacetate succinimidyl ester-labeled blood Tms were cocultured with autologous DCs, T cell proliferation and effector function were assessed using flow cytometry, and secreted cytokines were measured by combined bead array. UPM-DCs elicited IFN-γ and IL-13 secretion and induced proliferation in Tms isolated from both allergic patients with asthma and healthy control subjects, whereas only IL-13 was produced by Tms from patients with atopic asthma stimulated by house dust mite-loaded DCs. UPM-DCs drove the expansion and differentiation of a mixed population of Th1, Th2, and Th17 cell effectors through a mechanism that was dependent on major histocompatibility class II but not on cytokine-driven expansion. The data suggest that UPM not only has adjuvant properties but is also a source of antigen that stimulates the generation of Th2, Th1, and Th17 effector phenotypes, which have been implicated in both exacerbations of asthma and chronic inflammatory diseases.

  14. Malaria-associated atypical memory B cells exhibit markedly reduced B cell receptor signaling and effector function

    PubMed Central

    Portugal, Silvia; Tipton, Christopher M; Sohn, Haewon; Kone, Younoussou; Wang, Jing; Li, Shanping; Skinner, Jeff; Virtaneva, Kimmo; Sturdevant, Daniel E; Porcella, Stephen F; Doumbo, Ogobara K; Doumbo, Safiatou; Kayentao, Kassoum; Ongoiba, Aissata; Traore, Boubacar; Sanz, Inaki; Pierce, Susan K; Crompton, Peter D

    2015-01-01

    Protective antibodies in Plasmodium falciparum malaria are only acquired after years of repeated infections. Chronic malaria exposure is associated with a large increase in atypical memory B cells (MBCs) that resemble B cells expanded in a variety of persistent viral infections. Understanding the function of atypical MBCs and their relationship to classical MBCs will be critical to developing effective vaccines for malaria and other chronic infections. We show that VH gene repertoires and somatic hypermutation rates of atypical and classical MBCs are indistinguishable indicating a common developmental history. Atypical MBCs express an array of inhibitory receptors and B cell receptor (BCR) signaling is stunted in atypical MBCs resulting in impaired B cell responses including proliferation, cytokine production and antibody secretion. Thus, in response to chronic malaria exposure, atypical MBCs appear to differentiate from classical MBCs becoming refractory to BCR-mediated activation and potentially interfering with the acquisition of malaria immunity. DOI: http://dx.doi.org/10.7554/eLife.07218.001 PMID:25955968

  15. Comprehensive analysis of the specificity of transcription activator-like effector nucleases

    PubMed Central

    Juillerat, Alexandre; Dubois, Gwendoline; Valton, Julien; Thomas, Séverine; Stella, Stefano; Maréchal, Alan; Langevin, Stéphanie; Benomari, Nassima; Bertonati, Claudia; Silva, George H.; Daboussi, Fayza; Epinat, Jean-Charles; Montoya, Guillermo; Duclert, Aymeric; Duchateau, Philippe

    2014-01-01

    A key issue when designing and using DNA-targeting nucleases is specificity. Ideally, an optimal DNA-targeting tool has only one recognition site within a genomic sequence. In practice, however, almost all designer nucleases available today can accommodate one to several mutations within their target site. The ability to predict the specificity of targeting is thus highly desirable. Here, we describe the first comprehensive experimental study focused on the specificity of the four commonly used repeat variable diresidues (RVDs; NI:A, HD:C, NN:G and NG:T) incorporated in transcription activator-like effector nucleases (TALEN). The analysis of >15 500 unique TALEN/DNA cleavage profiles allowed us to monitor the specificity gradient of the RVDs along a TALEN/DNA binding array and to present a specificity scoring matrix for RVD/nucleotide association. Furthermore, we report that TALEN can only accommodate a relatively small number of position-dependent mismatches while maintaining a detectable activity at endogenous loci in vivo, demonstrating the high specificity of these molecular tools. We thus envision that the results we provide will allow for more deliberate choices of DNA binding arrays and/or DNA targets, extending our engineering capabilities. PMID:24569350

  16. End effectors and grapple fixtures

    NASA Astrophysics Data System (ADS)

    Vandersluis, Ron; Quittner, Erik

    1992-01-01

    An end effector has been developed for use with a space station remote manipulator system where capture and release capabilities are required, and which will provide for the transfer of substantial loads together with electrical power and signals across the end effector grapple fixture interface. The end effector has a latching mechanism for the transfer of substantial loads across the end effector grapple fixture interface. The functions associated with known nonlatching end effectors, namely their snaring and rigidizing capabilities, are maintained and can be operated independently of the new latching mechanisms and umbilical connectors of the end effector. The end effector is capable of functioning equally as a wrist (manipulator) and shoulder (arm base) unit. Applications of the new end effector include space station assembly, payload handling, capture of free-flyers, payload servicing, and providing stable bases for extravehicular activity work stations or robotic devices.

  17. Phenotypic and Functional Characterization of Herpes Simplex Virus Glycoprotein B Epitope-Specific Effector and Memory CD8+ T Cells from Symptomatic and Asymptomatic Individuals with Ocular Herpes

    PubMed Central

    Khan, Arif A.; Srivastava, Ruchi; Spencer, Doran; Garg, Sumit; Fremgen, Daniel; Vahed, Hawa; Lopes, Patricia P.; Pham, Thanh T.; Hewett, Charlie; Kuang, Jasmine; Ong, Nicolas; Huang, Lei; Scarfone, Vanessa M.; Nesburn, Anthony B.

    2015-01-01

    ABSTRACT Herpes simplex virus 1 (HSV-1) glycoprotein B (gB)-specific CD8+ T cells protect mice from herpes infection and disease. However, whether and which HSV-1 gB-specific CD8+ T cells play a key role in the “natural” protection seen in HSV-1-seropositive healthy asymptomatic (ASYMP) individuals (who have never had clinical herpes disease) remain to be determined. In this study, we have dissected the phenotypes and the functions of HSV-1 gB-specific CD8+ T cells from HLA-A*02:01 positive, HSV-1 seropositive ASYMP and symptomatic (SYMP) individuals (with a history of numerous episodes of recurrent ocular herpes disease). We found the following. (i) Healthy ASYMP individuals maintained a significantly higher proportion of differentiated HSV-1 gB-specific effector memory CD8+ T cells (TEM cells) (CD45RAlow CCR7low CD44high CD62Llow). In contrast, SYMP patients had frequent less-differentiated central memory CD8+ T cells (TCM cells) (CD45RAlow CCR7high CD44low CD62Lhigh). (ii) ASYMP individuals had significantly higher proportions of multifunctional effector CD8+ T cells which responded mainly to gB342–350 and gB561–569 “ASYMP” epitopes, and simultaneously produced IFN-γ, CD107a/b, granzyme B, and perforin. In contrast, effector CD8+ T cells from SYMP individuals were mostly monofunctional and were directed mainly against nonoverlapping gB17–25 and gB183–191 “SYMP” epitopes. (iii) Immunization of an HLA-A*02:01 transgenic mouse model of ocular herpes with “ASYMP” CD8+ TEM cell epitopes, but not with “SYMP” CD8+ TCM cell epitopes, induced a strong CD8+ T cell-dependent protective immunity against ocular herpes infection and disease. Our findings provide insights into the role of HSV-specific CD8+ TEM cells in protection against herpes and should be considered in the development of an effective vaccine. IMPORTANCE A significantly higher proportion of differentiated and multifunctional HSV-1 gB-specific effector memory CD8+ T cells (TEM

  18. Prefrontal Activity Links Nonoverlapping Events in Memory

    PubMed Central

    Miyawaki, Hiroyuki; Helmstetter, Fred J.; Diba, Kamran

    2013-01-01

    The medial prefrontal cortex (mPFC) plays an important role in memory. By maintaining a working memory buffer, neurons in prelimbic (PL) mPFC may selectively contribute to learning associations between stimuli that are separated in time, as in trace fear conditioning (TFC). Until now, evidence for this bridging role was largely descriptive. Here we used optogenetics to silence neurons in the PL mPFC of rats during learning in TFC. Memory formation was prevented when mPFC was silenced specifically during the interval separating the cue and shock. Our results provide support for a working memory function for these cells and indicate that associating two noncontiguous stimuli requires bridging activity in PL mPFC. PMID:23804110

  19. Regulation of gene expression in autoimmune disease loci and the genetic basis of proliferation in CD4+ effector memory T cells.

    PubMed

    Hu, Xinli; Kim, Hyun; Raj, Towfique; Brennan, Patrick J; Trynka, Gosia; Teslovich, Nikola; Slowikowski, Kamil; Chen, Wei-Min; Onengut, Suna; Baecher-Allan, Clare; De Jager, Philip L; Rich, Stephen S; Stranger, Barbara E; Brenner, Michael B; Raychaudhuri, Soumya

    2014-06-01

    Genome-wide association studies (GWAS) and subsequent dense-genotyping of associated loci identified over a hundred single-nucleotide polymorphism (SNP) variants associated with the risk of rheumatoid arthritis (RA), type 1 diabetes (T1D), and celiac disease (CeD). Immunological and genetic studies suggest a role for CD4-positive effector memory T (CD+ TEM) cells in the pathogenesis of these diseases. To elucidate mechanisms of autoimmune disease alleles, we investigated molecular phenotypes in CD4+ effector memory T cells potentially affected by these variants. In a cohort of genotyped healthy individuals, we isolated high purity CD4+ TEM cells from peripheral blood, then assayed relative abundance, proliferation upon T cell receptor (TCR) stimulation, and the transcription of 215 genes within disease loci before and after stimulation. We identified 46 genes regulated by cis-acting expression quantitative trait loci (eQTL), the majority of which we detected in stimulated cells. Eleven of the 46 genes with eQTLs were previously undetected in peripheral blood mononuclear cells. Of 96 risk alleles of RA, T1D, and/or CeD in densely genotyped loci, eleven overlapped cis-eQTLs, of which five alleles completely explained the respective signals. A non-coding variant, rs389862A, increased proliferative response (p=4.75 × 10-8). In addition, baseline expression of seventeen genes in resting cells reliably predicted proliferative response after TCR stimulation. Strikingly, however, there was no evidence that risk alleles modulated CD4+ TEM abundance or proliferation. Our study underscores the power of examining molecular phenotypes in relevant cells and conditions for understanding pathogenic mechanisms of disease variants. PMID:24968232

  20. Rh D blood group conversion using transcription activator-like effector nucleases.

    PubMed

    Kim, Young-Hoon; Kim, Hyun O; Baek, Eun J; Kurita, Ryo; Cha, Hyuk-Jin; Nakamura, Yukio; Kim, Hyongbum

    2015-06-16

    Group O D-negative blood cells are universal donors in transfusion medicine and methods for converting other blood groups into this universal donor group have been researched. However, conversion of D-positive cells into D-negative is yet to be achieved, although conversion of group A or B cells into O cells has been reported. The Rh D blood group is determined by the RHD gene, which encodes a 12-transmembrane domain protein. Here we convert Rh D-positive erythroid progenitor cells into D-negative cells using RHD-targeting transcription activator-like effector nucleases (TALENs). After transfection of TALEN-encoding plasmids, RHD-knockout clones are obtained. Erythroid-lineage cells differentiated from these knockout erythroid progenitor cells do not agglutinate in the presence of anti-D reagents and do not express D antigen, as assessed using flow cytometry. Our programmable nuclease-induced blood group conversion opens new avenues for compatible donor cell generation in transfusion medicine.

  1. Direct observation of transcription activator-like effector (TALE) protein dynamics

    NASA Astrophysics Data System (ADS)

    Cuculis, Luke; Abil, Zhanar; Zhao, Huimin; Schroeder, Charles M.

    2014-03-01

    In this work, we describe a single molecule assay to probe the site-search dynamics of transcription activator-like effector (TALE) proteins along DNA. In modern genetics, the ability to selectively edit the human genome is an unprecedented development, driven by recent advances in targeted nuclease proteins. Specific gene editing can be accomplished using TALE proteins, which are programmable DNA-binding proteins that can be fused to a nuclease domain. In this way, TALENs are a leading technology that has shown great success in the genomic editing of pluripotent stem cells. A major hurdle facing clinical implementation, however, is the potential for deleterious off-target binding events. For these reasons, a molecular-level understanding of TALE binding and target sequence search on DNA is essential. To this end, we developed a single-molecule fluorescence imaging assay that provides a first-of-its-kind view of the 1-D diffusion of TALE proteins along stretched DNA. Taken together with co-crystal structures of DNA-bound TALEs, our results suggest a rotationally-coupled, major groove tracking model for diffusion. We further report diffusion constants for TALE proteins as a function of salt concentration, consistent with previously described models of 1-D protein diffusion.

  2. Genome Editing in Astyanax mexicanus Using Transcription Activator-like Effector Nucleases (TALENs).

    PubMed

    Kowalko, Johanna E; Ma, Li; Jeffery, William R

    2016-01-01

    Identifying alleles of genes underlying evolutionary change is essential to understanding how and why evolution occurs. Towards this end, much recent work has focused on identifying candidate genes for the evolution of traits in a variety of species. However, until recently it has been challenging to functionally validate interesting candidate genes. Recently developed tools for genetic engineering make it possible to manipulate specific genes in a wide range of organisms. Application of this technology in evolutionarily relevant organisms will allow for unprecedented insight into the role of candidate genes in evolution. Astyanax mexicanus (A. mexicanus) is a species of fish with both surface-dwelling and cave-dwelling forms. Multiple independent lines of cave-dwelling forms have evolved from ancestral surface fish, which are interfertile with one another and with surface fish, allowing elucidation of the genetic basis of cave traits. A. mexicanus has been used for a number of evolutionary studies, including linkage analysis to identify candidate genes responsible for a number of traits. Thus, A. mexicanus is an ideal system for the application of genome editing to test the role of candidate genes. Here we report a method for using transcription activator-like effector nucleases (TALENs) to mutate genes in surface A. mexicanus. Genome editing using TALENs in A. mexicanus has been utilized to generate mutations in pigmentation genes. This technique can also be utilized to evaluate the role of candidate genes for a number of other traits that have evolved in cave forms of A. mexicanus. PMID:27404092

  3. Targeted genome regulation and modification using transcription activator-like effectors.

    PubMed

    Scott, James N F; Kupinski, Adam P; Boyes, Joan

    2014-10-01

    Transcription activator-like effectors (TALEs) are immensely powerful new tools for genome engineering that can be directed to bind to almost any DNA sequence of choice. They originate from the Xanthomonas species of plant pathogenic bacteria and, in nature, these proteins increase the virulence of Xanthomonas. However, in 2009, the DNA binding code of TALEs was deciphered and, subsequently, TALE proteins have been exploited for many diverse applications. Custom TALEs that target almost any required DNA sequence can be readily constructed in < 1 week. One major application is gene editing: TALEs fused with the Fok I endonuclease catalytic domain can induce double-stranded breaks at a chosen genomic location, similar to zinc finger nucleases. Designer TALE transcription factors have also been developed by linking TALEs to a transcription AD, such as VP64. More recently, TALEs have been developed that can repress transcription, bind methylated DNA or act as fluorescent chromatin probes. In the present review, we describe the assembly of designer TALEs, their expanding range of current and potential future applications, and briefly discuss alternatives, namely, zinc finger nucleases and clustered regularly interspaced short palindromic repeat/clustered regularly interspaced short palindromic repeat associated protein 9.

  4. Distinct activation thresholds of human conventional and innate-like memory T cells

    PubMed Central

    Slichter, Chloe K.; Miller, Hannah W.; Seymour, Brenda J.; McNevin, John P.; Diaz, Gabriela; Czartoski, Julie L.; McElrath, M. Juliana; Gottardo, Raphael

    2016-01-01

    Conventional memory CD8+ T cells and mucosal-associated invariant T cells (MAIT cells) are found in blood, liver, and mucosal tissues and have similar effector potential following activation, specifically expression of IFN-γ and granzyme B. To better understand each subset’s unique contributions to immunity and pathology, we interrogated inflammation- and TCR-driven activation requirements using human memory CD8+ T and MAIT cells isolated from blood and mucosal tissue biopsies in ex vivo functional assays and single cell gene expression experiments. We found that MAIT cells had a robust IFN-γ and granzyme B response to inflammatory signals but limited responsiveness when stimulated directly via their TCR. Importantly, this is not due to an overall hyporesponsiveness to TCR signals. When delivered together, TCR and inflammatory signals synergize to elicit potent effector function in MAIT cells. This unique control of effector function allows MAIT cells to respond to the same TCR signal in a dichotomous and situation-specific manner. We propose that this could serve to prevent responses to antigen in noninflamed healthy mucosal tissue, while maintaining responsiveness and great sensitivity to inflammation-eliciting infections. We discuss the implications of these findings in context of inflammation-inducing damage to tissues such as BM transplant conditioning or HIV infection. PMID:27331143

  5. The deubiquitinase activity of the Salmonella pathogenicity island 2 effector, SseL, prevents accumulation of cellular lipid droplets.

    PubMed

    Arena, Ellen T; Auweter, Sigrid D; Antunes, L Caetano M; Vogl, A Wayne; Han, Jun; Guttman, Julian A; Croxen, Matthew A; Menendez, Alfredo; Covey, Scott D; Borchers, Christoph H; Finlay, B Brett

    2011-11-01

    To cause disease, Salmonella enterica serovar Typhimurium requires two type III secretion systems that are encoded by Salmonella pathogenicity islands 1 and 2 (SPI-1 and -2). These secretion systems serve to deliver specialized proteins (effectors) into the host cell cytosol. While the importance of these effectors to promote colonization and replication within the host has been established, the specific roles of individual secreted effectors in the disease process are not well understood. In this study, we used an in vivo gallbladder epithelial cell infection model to study the function of the SPI-2-encoded type III effector, SseL. The deletion of the sseL gene resulted in bacterial filamentation and elongation and the unusual localization of Salmonella within infected epithelial cells. Infection with the ΔsseL strain also caused dramatic changes in host cell lipid metabolism and led to the massive accumulation of lipid droplets in infected cells. This phenotype was directly attributable to the deubiquitinase activity of SseL, as a Salmonella strain carrying a single point mutation in the catalytic cysteine also resulted in extensive lipid droplet accumulation. The excessive buildup of lipids due to the absence of a functional sseL gene also was observed in murine livers during S. Typhimurium infection. These results suggest that SseL alters host lipid metabolism in infected epithelial cells by modifying the ubiquitination patterns of cellular targets.

  6. Congruent and Incongruent Corticospinal Activations at the Level of Multiple Effectors.

    PubMed

    Sartori, Luisa; Betti, Sonia; Perrone, Chiara; Castiello, Umberto

    2015-10-01

    Motor resonance is defined as the subliminal activation of the motor system while observing actions performed by others. However, resonating with another person's actions is not always an appropriate response: In real life, people do not just imitate but rather respond in a suitable fashion. A growing body of neurophysiologic studies has demonstrated that motor resonance can be overridden by complementary motor responses (such as preparing a precision grip on a small object when seeing an open hand in sign of request). In this study, we investigated the relationship between congruent and incongruent corticospinal activations at the level of multiple effectors. The modulation of motor evoked potentials evoked by single-pulse TMS over the motor cortex was assessed in upper and lower limb muscles of participants observing a soccer player performing a penalty kick straight in their direction. Study results revealed a double dissociation: Seeing the soccer player kicking the ball triggered a motor resonance in the observer's lower limb, whereas the upper limb response afforded by the object was overridden. On the other hand, seeing the ball approaching the observers elicited a complementary motor activation in upper limbs while motor resonance in lower limbs disappeared. Control conditions showing lateral kicks, mimicked kicks, and a ball in penalty area were also included to test the motor coding of object affordances. Results point to a modulation of motor responses in different limbs over the course of action and in function of their relevance in different contexts. We contend that ecologically valid paradigms are now needed to shed light on the motor system functioning in complex forms of interaction. PMID:26102231

  7. Characterization of effector and memory T cell subsets in the immune response to bovine tuberculosis in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vaccine-elicited long-term cultured IFN-gamma ELISPOT responses correlate with protection against bovine tuberculosis in cattle. With humans, cultured IFN-gamma ELISPOT assays are primarily a measure of central memory T cell (Tcm) responses; however, this important subset of lymphocytes is poorly ch...

  8. E2~Ub conjugates regulate the kinase activity of Shigella effector OspG during pathogenesis

    SciTech Connect

    Pruneda, Jonathan N.; Smith, F. Donelson; Daurie, Angela; Swaney, Danielle L.; Villén, Judit; Scott, John D.; Stadnyk, Andrew W.; Le Trong, Isolde; Stenkamp, Ronald E.; Klevit, Rachel E.; Rohde, John R.; Brzovic, Peter S.

    2014-01-20

    Pathogenic bacteria introduce effector proteins directly into the cytosol of eukaryotic cells to promote invasion and colonization. OspG, a Shigella spp. effector kinase, plays a role in this process by helping to suppress the host inflammatory response. OspG has been reported to bind host E2 ubiquitin-conjugating enzymes activated with ubiquitin (E2~Ub), a key enzyme complex in ubiquitin transfer pathways. A cocrystal structure of the OspG/UbcH5c~Ub complex reveals that complex formation has important ramifications for the activity of both OspG and the UbcH5c~Ub conjugate. OspG is a minimal kinase domain containing only essential elements required for catalysis. UbcH5c~Ub binding stabilizes an active conformation of the kinase, greatly enhancing OspG kinase activity. In contrast, interaction with OspG stabilizes an extended, less reactive form of UbcH5c~Ub. Recognizing conserved E2 features, OspG can interact with at least ten distinct human E2s~Ub. Mouse oral infection studies indicate that E2~Ub conjugates act as novel regulators of OspG effector kinase function in eukaryotic host cells.

  9. E2∼Ub conjugates regulate the kinase activity of Shigella effector OspG during pathogenesis

    PubMed Central

    Pruneda, Jonathan N; Smith, F Donelson; Daurie, Angela; Swaney, Danielle L; Villén, Judit; Scott, John D; Stadnyk, Andrew W; Le Trong, Isolde; Stenkamp, Ronald E; Klevit, Rachel E; Rohde, John R; Brzovic, Peter S

    2014-01-01

    Pathogenic bacteria introduce effector proteins directly into the cytosol of eukaryotic cells to promote invasion and colonization. OspG, a Shigella spp. effector kinase, plays a role in this process by helping to suppress the host inflammatory response. OspG has been reported to bind host E2 ubiquitin-conjugating enzymes activated with ubiquitin (E2∼Ub), a key enzyme complex in ubiquitin transfer pathways. A co-crystal structure of the OspG/UbcH5c∼Ub complex reveals that complex formation has important ramifications for the activity of both OspG and the UbcH5c∼Ub conjugate. OspG is a minimal kinase domain containing only essential elements required for catalysis. UbcH5c∼Ub binding stabilizes an active conformation of the kinase, greatly enhancing OspG kinase activity. In contrast, interaction with OspG stabilizes an extended, less reactive form of UbcH5c∼Ub. Recognizing conserved E2 features, OspG can interact with at least ten distinct human E2s∼Ub. Mouse oral infection studies indicate that E2∼Ub conjugates act as novel regulators of OspG effector kinase function in eukaryotic host cells. PMID:24446487

  10. World War II Memorial Learning Activities.

    ERIC Educational Resources Information Center

    Tennessee State Dept. of Education, Nashville.

    These learning activities can help students get the most out of a visit to the Tennessee World War II Memorial, a group of ten pylons located in Nashville (Tennessee). Each pylon contains informational text about the events of World War II. The ten pylons are listed as: (1) "Pylon E-1--Terror: America Enters the War against Fascism, June 1940";…

  11. The Pseudomonas aeruginosa Type VI Secretion PGAP1-like Effector Induces Host Autophagy by Activating Endoplasmic Reticulum Stress.

    PubMed

    Jiang, Feng; Wang, Xia; Wang, Bei; Chen, Lihong; Zhao, Zhendong; Waterfield, Nicholas R; Yang, Guowei; Jin, Qi

    2016-08-01

    Pseudomonas aeruginosa is an opportunistic pathogen that regularly causes nosocomial infections in hospitalized patients. The type VI secretion system (T6SS) is responsible for the secretion of numerous virulence effector proteins that can both interfere with competing microbes and manipulate host cells. Here, we report a detailed investigation of a P. aeruginosa H2-T6SS-dependent phospholipase effector, TplE, which acts as a trans-kingdom toxin. Delivery of TplE to the periplasmic space of rival bacteria leads to growth inhibition. Importantly, TplE, also contains a eukaryotic PGAP1-like domain, which targets the host ER apparatus, ultimately leading to disruption of the ER. TplE activity leads to the activation of the unfolded protein response (UPR) through the IRE1α-XBP1 pathway, enhancing autophagic flux. These findings indicate that this T6SS-delivered phospholipase effector is active against both prokaryotic and eukaryotic cellular targets, highlighting the T6SS as a versatile weapon in the Pseudomonas arsenal. PMID:27477276

  12. Place cell activation predicts subsequent memory.

    PubMed

    Robitsek, R Jonathan; White, John A; Eichenbaum, Howard

    2013-10-01

    A major quandary in memory research is how hippocampal place cells, widely recognized as elements of a spatial map, contribute to episodic memory, our capacity to remember unique experiences that depends on hippocampal function. Here we recorded from hippocampal neurons as rats performed a T-maze alternation task in which they were required to remember a preceding experience over a delay in order to make a subsequent spatial choice. As it has been reported previously in other variations of this task, we observed differential firing that predicted correct subsequent choices, even as the animal traversed identical locations prior to the choice. Here we also observed that most place cells also fired differently on correct as compared to error trials. Among these cells, a large majority fired strongly before the delay or during the retrieval phase but were less active or failed to activate when the animal subsequently made an error. These findings join the place cell phenomenon with episodic memory performance dependent on the hippocampus, revealing that memory accuracy can be predicted by the activation of single place cells in the hippocampus.

  13. Tumor suppressor genes are larger than apoptosis-effector genes and have more regions of active chromatin: Connection to a stochastic paradigm for sequential gene expression programs.

    PubMed

    Garcia, Marlene; Mauro, James A; Ramsamooj, Michael; Blanck, George

    2015-08-01

    Apoptosis- and proliferation-effector genes are substantially regulated by the same transactivators, with E2F-1 and Oct-1 being notable examples. The larger proliferation-effector genes have more binding sites for the transactivators that regulate both sets of genes, and proliferation-effector genes have more regions of active chromatin, i.e, DNase I hypersensitive and histone 3, lysine-4 trimethylation sites. Thus, the size differences between the 2 classes of genes suggest a transcriptional regulation paradigm whereby the accumulation of transcription factors that regulate both sets of genes, merely as an aspect of stochastic behavior, accumulate first on the larger proliferation-effector gene "traps," and then accumulate on the apoptosis effector genes, thereby effecting sequential activation of the 2 different gene sets. As IRF-1 and p53 levels increase, tumor suppressor proteins are first activated, followed by the activation of apoptosis-effector genes, for example during S-phase pausing for DNA repair. Tumor suppressor genes are larger than apoptosis-effector genes and have more IRF-1 and p53 binding sites, thereby likewise suggesting a paradigm for transcription sequencing based on stochastic interactions of transcription factors with different gene classes. In this report, using the ENCODE database, we determined that tumor suppressor genes have a greater number of open chromatin regions and histone 3 lysine-4 trimethylation sites, consistent with the idea that a larger gene size can facilitate earlier transcriptional activation via the inclusion of more transactivator binding sites.

  14. Psychobiology of Active and Inactive Memory.

    ERIC Educational Resources Information Center

    Lewis, Donald J.

    1979-01-01

    Argues that the distinction between short-term memory and long-term memory is no longer adequate for either human or animal memory data. Recommends additional research on the physiological brain processes underlying memory interference and retrieval. (MP)

  15. Activated Ras Induces Cytoplasmic Vacuolation and Non-Apoptotic Death in Glioblastoma Cells via Novel Effector Pathways

    PubMed Central

    Kaul, Aparna; Overmeyer, Jean H.; Maltese, William A.

    2007-01-01

    Expression of activated H-Ras induces a unique form of non-apoptotic cell death in human glioblastoma cells and other specific tumor cell lines. The major cytopathological features of this form of death are the accumulation of large phase-lucent, LAMP1-positive, cytoplasmic vacuoles and increased autophagic activity. In this study we sought to determine if induction of cytoplasmic vacuolation a) depends on Ras farnesylation, b) is specific to H-Ras, and c) is mediated by signaling through the major known Ras effector pathways. We find that the unusual effects of activated H-Ras depend on farnesylation and membrane association of the GTPase. Both H-Ras(G12V) and K-Ras4B(G12V) stimulate vacuolation, but activated forms of Cdc42 and RhoA do not. Amino acid substitutions in the Ras effector domain, which are known to selectively impair its interactions with Raf kinase, class-I phosphatidylinositide 3-kinase (PI3K), or Ral nucleotide exchange factors, initially pointed to Raf as a possible mediator of cell vacuolation. However, the MEK inhibitor, PD98059, did not block the induction of vacuoles, and constitutively active Raf-Caax did not mimic the effects of Ras(G12V). Introduction of normal PTEN together with H-Ras(G12V) into U251 glioblastoma cells reduced the PI3K-dependent activation of Akt, but had no effect on vacuolation. Finally, co-expression of H-Ras(G12V) with a dominant-negative form of RalA did not suppress vacuolation. Taken together, the observations indicate that Ras activates non-conventional and perhaps unique effector pathways to induce cytoplasmic vacuolation in glioblastoma cells. Identification of the relevant signaling pathways may uncover specific molecular targets that can be manipulated to activate non-apoptotic cell death in this type of cancer. PMID:17210246

  16. Relation of Physical Activity to Memory Functioning in Older Adults: The Memory Workout Program.

    ERIC Educational Resources Information Center

    Rebok, George W.; Plude, Dana J.

    2001-01-01

    The Memory Workout, a CD-ROM program designed to help older adults increase changes in physical and cognitive activity influencing memory, was tested with 24 subjects. Results revealed a significant relationship between exercise time, exercise efficacy, and cognitive function, as well as interest in improving memory and physical activity.…

  17. Identification of Novel Coxiella burnetii Icm/Dot Effectors and Genetic Analysis of Their Involvement in Modulating a Mitogen-Activated Protein Kinase Pathway

    PubMed Central

    Lifshitz, Ziv; Burstein, David; Schwartz, Kierstyn; Shuman, Howard A.; Pupko, Tal

    2014-01-01

    Coxiella burnetii, the causative agent of Q fever, is a human intracellular pathogen that utilizes the Icm/Dot type IVB secretion system to translocate effector proteins into host cells. To identify novel C. burnetii effectors, we applied a machine-learning approach to predict C. burnetii effectors, and examination of 20 such proteins resulted in the identification of 13 novel effectors. To determine whether these effectors, as well as several previously identified effectors, modulate conserved eukaryotic pathways, they were expressed in Saccharomyces cerevisiae. The effects on yeast growth were examined under regular growth conditions and in the presence of caffeine, a known modulator of the yeast cell wall integrity (CWI) mitogen-activated protein (MAP) kinase pathway. In the presence of caffeine, expression of the effectors CBU0885 and CBU1676 caused an enhanced inhibition of yeast growth, and the growth inhibition of CBU0388 was suppressed. Furthermore, analysis of synthetic lethality effects and examination of the activity of the CWI MAP kinase transcription factor Rlm1 indicated that CBU0388 enhances the activation of this MAP kinase pathway in yeast, while CBU0885 and CBU1676 abolish this activation. Additionally, coexpression of CBU1676 and CBU0388 resulted in mutual suppression of their inhibition of yeast growth. These results strongly indicate that these three effectors modulate the CWI MAP kinase pathway in yeast. Moreover, both CBU1676 and CBU0885 were found to contain a conserved haloacid dehalogenase (HAD) domain, which was found to be required for their activity. Collectively, our results demonstrate that MAP kinase pathways are most likely targeted by C. burnetii Icm/Dot effectors. PMID:24958706

  18. Caspase-1 activation in macrophages infected with Yersinia pestis KIM requires the type III secretion system effector YopJ.

    PubMed

    Lilo, Sarit; Zheng, Ying; Bliska, James B

    2008-09-01

    Pathogenic Yersinia species utilize a type III secretion system (T3SS) to translocate effectors called Yersinia outer proteins (Yops) into infected host cells. Previous studies demonstrated a role for effector Yops in the inhibition of caspase-1-mediated cell death and secretion of interleukin-1beta (IL-1beta) in naïve macrophages infected with Yersinia enterocolitica. Naïve murine macrophages were infected with a panel of different Yersinia pestis and Yersinia pseudotuberculosis strains to determine whether Yops of these species inhibit caspase-1 activation. Cell death was measured by release of lactate dehydrogenase (LDH), and enzyme-linked immunosorbent assay for secreted IL-1beta was used to measure caspase-1 activation. Surprisingly, isolates derived from the Y. pestis KIM strain (e.g., KIM5) displayed an unusual ability to activate caspase-1 and kill infected macrophages compared to other Y. pestis and Y. pseudotuberculosis strains tested. Secretion of IL-1beta following KIM5 infection was reduced in caspase-1-deficient macrophages compared to wild-type macrophages. However, release of LDH was not reduced in caspase-1-deficient macrophages, indicating that cell death occurred independently of caspase-1. Analysis of KIM-derived strains defective for production of functional effector or translocator Yops indicated that translocation of catalytically active YopJ into macrophages was required for caspase-1 activation and cell death. Release of LDH and secretion of IL-1beta were not reduced when actin polymerization was inhibited in KIM5-infected macrophages, indicating that extracellular bacteria translocating YopJ could trigger cell death and caspase-1 activation. This study uncovered a novel role for YopJ in the activation of caspase-1 in macrophages.

  19. Caspase-1 Activation in Macrophages Infected with Yersinia pestis KIM Requires the Type III Secretion System Effector YopJ▿

    PubMed Central

    Lilo, Sarit; Zheng, Ying; Bliska, James B.

    2008-01-01

    Pathogenic Yersinia species utilize a type III secretion system (T3SS) to translocate effectors called Yersinia outer proteins (Yops) into infected host cells. Previous studies demonstrated a role for effector Yops in the inhibition of caspase-1-mediated cell death and secretion of interleukin-1β (IL-1β) in naïve macrophages infected with Yersinia enterocolitica. Naïve murine macrophages were infected with a panel of different Yersinia pestis and Yersinia pseudotuberculosis strains to determine whether Yops of these species inhibit caspase-1 activation. Cell death was measured by release of lactate dehydrogenase (LDH), and enzyme-linked immunosorbent assay for secreted IL-1β was used to measure caspase-1 activation. Surprisingly, isolates derived from the Y. pestis KIM strain (e.g., KIM5) displayed an unusual ability to activate caspase-1 and kill infected macrophages compared to other Y. pestis and Y. pseudotuberculosis strains tested. Secretion of IL-1β following KIM5 infection was reduced in caspase-1-deficient macrophages compared to wild-type macrophages. However, release of LDH was not reduced in caspase-1-deficient macrophages, indicating that cell death occurred independently of caspase-1. Analysis of KIM-derived strains defective for production of functional effector or translocator Yops indicated that translocation of catalytically active YopJ into macrophages was required for caspase-1 activation and cell death. Release of LDH and secretion of IL-1β were not reduced when actin polymerization was inhibited in KIM5-infected macrophages, indicating that extracellular bacteria translocating YopJ could trigger cell death and caspase-1 activation. This study uncovered a novel role for YopJ in the activation of caspase-1 in macrophages. PMID:18559430

  20. Identification of Phakopsora pachyrhizi Candidate Effectors with Virulence Activity in a Distantly Related Pathosystem

    PubMed Central

    Kunjeti, Sridhara G.; Iyer, Geeta; Johnson, Ebony; Li, Eric; Broglie, Karen E.; Rauscher, Gilda; Rairdan, Gregory J.

    2016-01-01

    Phakopsora pachyrhizi is the causal agent of Asian Soybean Rust, a disease that causes enormous economic losses, most markedly in South America. P. pachyrhizi is a biotrophic pathogen that utilizes specialized feeding structures called haustoria to colonize its hosts. In rusts and other filamentous plant pathogens, haustoria have been shown to secrete effector proteins into their hosts to permit successful completion of their life cycle. We have constructed a cDNA library from P. pachyrhizi haustoria using paramagnetic bead-based methodology and have identified 35 P. pachyrhizi candidate effector (CE) genes from this library which are described here. In addition, we quantified the transcript expression pattern of six of these genes and show that two of these CEs are able to greatly increase the susceptibility of Nicotiana benthamiana to Phytophthora infestans. This strongly suggests that these genes play an important role in P. pachyrhizi virulence on its hosts. PMID:27014295

  1. Identification of Phakopsora pachyrhizi Candidate Effectors with Virulence Activity in a Distantly Related Pathosystem.

    PubMed

    Kunjeti, Sridhara G; Iyer, Geeta; Johnson, Ebony; Li, Eric; Broglie, Karen E; Rauscher, Gilda; Rairdan, Gregory J

    2016-01-01

    Phakopsora pachyrhizi is the causal agent of Asian Soybean Rust, a disease that causes enormous economic losses, most markedly in South America. P. pachyrhizi is a biotrophic pathogen that utilizes specialized feeding structures called haustoria to colonize its hosts. In rusts and other filamentous plant pathogens, haustoria have been shown to secrete effector proteins into their hosts to permit successful completion of their life cycle. We have constructed a cDNA library from P. pachyrhizi haustoria using paramagnetic bead-based methodology and have identified 35 P. pachyrhizi candidate effector (CE) genes from this library which are described here. In addition, we quantified the transcript expression pattern of six of these genes and show that two of these CEs are able to greatly increase the susceptibility of Nicotiana benthamiana to Phytophthora infestans. This strongly suggests that these genes play an important role in P. pachyrhizi virulence on its hosts. PMID:27014295

  2. Characterization of cell death inducing Phytophthora capsici CRN effectors suggests diverse activities in the host nucleus

    PubMed Central

    Stam, Remco; Howden, Andrew J. M.; Delgado-Cerezo, Magdalena; M. M. Amaro, Tiago M.; Motion, Graham B.; Pham, Jasmine; Huitema, Edgar

    2013-01-01

    Plant-Microbe interactions are complex associations that feature recognition of Pathogen Associated Molecular Patterns by the plant immune system and dampening of subsequent responses by pathogen encoded secreted effectors. With large effector repertoires now identified in a range of sequenced microbial genomes, much attention centers on understanding their roles in immunity or disease. These studies not only allow identification of pathogen virulence factors and strategies, they also provide an important molecular toolset suited for studying immunity in plants. The Phytophthora intracellular effector repertoire encodes a large class of proteins that translocate into host cells and exclusively target the host nucleus. Recent functional studies have implicated the CRN protein family as an important class of diverse effectors that target distinct subnuclear compartments and modify host cell signaling. Here, we characterized three necrosis inducing CRNs and show that there are differences in the levels of cell death. We show that only expression of CRN20_624 has an additive effect on PAMP induced cell death but not AVR3a induced ETI. Given their distinctive phenotypes, we assessed localization of each CRN with a set of nuclear markers and found clear differences in CRN subnuclear distribution patterns. These assays also revealed that expression of CRN83_152 leads to a distinct change in nuclear chromatin organization, suggesting a distinct series of events that leads to cell death upon over-expression. Taken together, our results suggest diverse functions carried by CRN C-termini, which can be exploited to identify novel processes that take place in the host nucleus and are required for immunity or susceptibility. PMID:24155749

  3. [Emotional Memory and Electrocortical Activity in Schizophrenia].

    PubMed

    Lavoie, Marc E; Champagne, Julie; Glaser, Emma; Mendrek, Adrianna

    2016-01-01

    Context Abnormal emotion processing is frequent in schizophrenia and affects social and functional outcome. Past event-related potential (ERP) research investigating processing of affective stimuli in schizophrenia was done mainly with facial expressions and revealed impaired facial emotion recognition in patients relative to control subjects. Experimentations involving fMRI with this group of patients, showed alteration of limbic and frontal regions in response to emotional unpleasant images, compared to neutral stimuli during a memory task. Other studies have also noted an increase in brain activity when the activation of the stimuli was high compared to low arousal stimuli. This may indicate a different sensitivity threshold to emotional arousal and emotional valence involving frontal pathways in these patients. But very few studies attempted to separate the contributions of emotional valence and arousal within an episodic memory protocol with ERP, in that population.Goal The aim of the current research is to investigate brain electro-cortical activity in schizophrenia in response to emotional images during an episodic memory task.Method ERP components were analyzed in 16 schizophrenic and 17 control participants matched for age, sex and intelligence. ERPs were obtained from 56 EEG electrodes. The tasks consisted in a classical episodic memory task that presented 100 repeated old and 100 new photographic images divided into four categories (unpleasant-high arousal, unpleasant-low arousal, pleasant-high arousal and pleasant-low arousal) selected from the International Affective Picture System. The N200, P300 and late positive component (LPC) mean amplitude, were analyzed using repeated-measure analyses of variance (MANOVA).Results Patients with schizophrenia and control subjects gave comparable subjective evaluations of arousal and valence. However, the frontal N200 and the P300 both showed an interaction of the group x memory x valence x hemisphere (F [1

  4. Macrophages as effector cells of protective immunity in murine schistosomiasis: macrophage activation in mice vaccinated with radiation-attenuated cercariae.

    PubMed Central

    James, S L; Natovitz, P C; Farrar, W L; Leonard, E J

    1984-01-01

    Cell-mediated immune responses contributing to macrophage activation were compared in mice that demonstrated partial resistance to challenge Schistosoma mansoni infection as a result of vaccination with radiation-attenuated cercariae or of ongoing low-grade primary infection. Vaccinated mice developed significant delayed hypersensitivity reactions to soluble schistosome antigens in vivo. Splenocytes from vaccinated animals responded to in vitro culture with various specific antigens (soluble adult worm extract, living or disrupted schistosomula) by proliferation and production of macrophage-activating lymphokines as did lymphocytes from S. mansoni-infected animals. Macrophage-activating factors produced by spleen cells from vaccinated mice upon specific antigen stimulation eluted as a single peak on Sephadex G-100 with a molecular weight of approximately 50,000 and contained gamma interferon activity. Moreover, peritoneal macrophages with larvicidal and tumoricidal activity were recovered from vaccinated mice after intraperitoneal challenge with soluble schistosome antigens, a procedure also observed to elicit activated macrophages in S. mansoni-infected animals. These observations demonstrate that vaccination with irradiated cercariae stimulates many of the same cellular responses observed after primary S. mansoni infection, and suggest that lymphokine-activated macrophages may participate in the effector mechanism of vaccine-induced and concomitant immunity to challenge schistosome infection. This is the first demonstration of a potential immune effector mechanism in the irradiated vaccine model. PMID:6609885

  5. False memory for context activates the parahippocampal cortex.

    PubMed

    Karanian, Jessica M; Slotnick, Scott D

    2014-01-01

    Previous studies have reported greater activity in the parahippocampal cortex during true memory than false memory, which has been interpreted as reflecting greater sensory processing during true memory. However, in these studies, sensory detail and contextual information were confounded. In the present fMRI study, we employed a novel paradigm to dissociate these factors. During encoding, abstract shapes were presented in one of two contexts (i.e., moving or stationary). During retrieval, participants classified shapes as previously "moving" or "stationary." Critically, contextual processing was relatively greater during false memory ("moving" responses to stationary items), while sensory processing was relatively greater during true memory ("moving" responses to moving items). Within the medial temporal lobe, false memory versus true memory produced greater activity in the parahippocampal cortex, whereas true memory versus false memory produced greater activity in the hippocampus. The present results indicate that the parahippocampal cortex mediates contextual processing rather than sensory processing.

  6. Modeling Change in Memory Performance and Memory Perceptions: Findings from the ACTIVE Study

    PubMed Central

    Parisi, Jeanine M.; Gross, Alden L.; Rebok, George W.; Saczynski, Jane S.; Crowe, Michael; Cook, Sarah E.; Langbaum, Jessica B. S.; Sartori, Andrea; Unverzagt, Fredrick W.

    2011-01-01

    Within the context of the ACTIVE study, the current investigation explored the relationships between objective memory and two components of subjective memory (frequency of forgetting and use of external aids) over a five-year period. Relationships were assessed using parallel process latent growth curve models. Results indicated that changes in objective memory were associated with changes in perceived frequency of forgetting, but not with use of external aids (calendars, reminder notes) over time. Findings suggest that memory complaints may accurately reflect decline in objective memory performance, but that these memory changes are not necessarily related to compensatory behaviors. PMID:21463064

  7. The type III secretion effector NleF of enteropathogenic Escherichia coli activates NF-κB early during infection.

    PubMed

    Pallett, Mitchell A; Berger, Cedric N; Pearson, Jaclyn S; Hartland, Elizabeth L; Frankel, Gad

    2014-11-01

    The enteric pathogens enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli employ a type 3 secretion system (T3SS) to manipulate the host inflammatory response during infection. Previously, it has been reported that EPEC, in a T3SS-dependent manner, induces an early proinflammatory response through activation of NF-κB via extracellular signal-regulated kinases 1 and 2 (ERK1/2) and protein kinase Cζ (PKCζ). However, the activation of NF-κB during infection has not yet been attributed to an effector. At later time points postinfection, NF-κB signaling is inhibited through the translocation of multiple effectors, including NleE and NleC. Here we report that the highly conserved non-LEE (locus of enterocyte effacement)-encoded effector F (NleF) shows both diffuse and mitochondrial localization during ectopic expression. Moreover, NleF induces the nuclear translocation of NF-κB p65 and the expression of interleukin 8 (IL-8) following ectopic expression and during EPEC infection. Furthermore, the proinflammatory activity and localization of NleF were dependent on the C-terminal amino acids LQCG. While the C-terminal domain of NleF has previously been shown to be essential for interaction with caspase-4, caspase-8, and caspase-9, the proinflammatory activity of NleF was independent of interaction with caspase-4, -8, or -9. In conclusion, EPEC, through the T3SS-dependent translocation of NleF, induces a proinflammatory response in an NF-κB-dependent manner in the early stages of infection.

  8. The Type III Secretion Effector NleF of Enteropathogenic Escherichia coli Activates NF-κB Early during Infection

    PubMed Central

    Pallett, Mitchell A.; Berger, Cedric N.; Pearson, Jaclyn S.; Hartland, Elizabeth L.

    2014-01-01

    The enteric pathogens enteropathogenic Escherichia coli (EPEC) and enterohemorrhagic E. coli employ a type 3 secretion system (T3SS) to manipulate the host inflammatory response during infection. Previously, it has been reported that EPEC, in a T3SS-dependent manner, induces an early proinflammatory response through activation of NF-κB via extracellular signal-regulated kinases 1 and 2 (ERK1/2) and protein kinase Cζ (PKCζ). However, the activation of NF-κB during infection has not yet been attributed to an effector. At later time points postinfection, NF-κB signaling is inhibited through the translocation of multiple effectors, including NleE and NleC. Here we report that the highly conserved non-LEE (locus of enterocyte effacement)-encoded effector F (NleF) shows both diffuse and mitochondrial localization during ectopic expression. Moreover, NleF induces the nuclear translocation of NF-κB p65 and the expression of interleukin 8 (IL-8) following ectopic expression and during EPEC infection. Furthermore, the proinflammatory activity and localization of NleF were dependent on the C-terminal amino acids LQCG. While the C-terminal domain of NleF has previously been shown to be essential for interaction with caspase-4, caspase-8, and caspase-9, the proinflammatory activity of NleF was independent of interaction with caspase-4, -8, or -9. In conclusion, EPEC, through the T3SS-dependent translocation of NleF, induces a proinflammatory response in an NF-κB-dependent manner in the early stages of infection. PMID:25183730

  9. Clinically Relevant Reactivation of Polyomavirus BK (BKPyV) in HLA-A02-Positive Renal Transplant Recipients Is Associated with Impaired Effector-Memory Differentiation of BKPyV-Specific CD8+ T Cells

    PubMed Central

    Remmerswaal, Ester B. M.; Heutinck, Kirstin M.; ten Brinke, Anja; Feltkamp, Mariet C. W.; van der Weerd, Neelke C.; van der Pant, Karlijn A. M. I.; Bemelman, Frederike J.; van Lier, René A. W.; ten Berge, Ineke J. M.

    2016-01-01

    Polyomavirus BK (BKPyV) frequently reactivates in immunosuppressed renal transplant recipients (RTRs) and may lead to graft loss due to BKPyV-induced interstitial nephritis (BKVN). Little is known on the differentiation of CD8+ T cells targeting BKPyV in RTRs. Here we investigated whether BKPyV-specific CD8+ T cell differentiation differs in RTRs with varying degrees of BKPyV reactivation and/or BKVN. Using combinatorial encoding with tetramers carrying BKPyV major capsid protein (VP1) and large T antigen protein (LTAG) epitopes, we investigated CD8+ T cell responses to BKPyV in longitudinally obtained PBMC samples from 46 HLA-A02-positive RTRs and 20 healthy adults. We were also able to isolate BKPyV-specific CD8+ T cells from five renal allografts, two of which were affected by BKVN. Before transplantation, BKPyV-specific CD8+ T cells targeting VP1 and LTAG epitopes appeared predominantly as central-memory and CD27+/CD28+ effector-memory (TEM), and naïve-like PD-1-expressing cells, respectively. After viral reactivation, BKPyV-specific CD8+ T cells assumed CD28− TEM and TEMRA states in patients who were able to control BKPyV, whereas differentiation lagged behind in patients with severe viral reactivation or BKVN. Furthermore, VP1-specific CD69+/CD103+ tissue-resident memory (TRM) cells accumulated in BKVN-affected allografts but lacked signs of effector differentiation. In contrast, granzyme B-expressing effector cells were detected in allografts not affected by BKVN. In conclusion, effector-memory differentiation of BKPyV-specific CD8+ T cells in patients with high viral load or BKVN is impaired. Further characterization of the specific mechanisms behind this altered cellular differentiation is necessary to develop therapies that can prevent the emergence of BKVN. PMID:27723787

  10. Low numbers of CD8+ T lymphocytes in hereditary haemochromatosis are explained by a decrease of the most mature CD8+ effector memory T cells

    PubMed Central

    Macedo, M F; Porto, G; Costa, M; Vieira, C P; Rocha, B; Cruz, E

    2010-01-01

    Low CD8+ T lymphocyte numbers have long been described in hereditary haemochromatosis (HH). Recently, two conserved haplotypes localized near the microsatellite D6S105 at the major histocompatibility complex (MHC) class I region were described predicting the clinical expression of HH and the CD8+ T lymphocyte numbers. The A-A-T haplotype was associated with a severe clinical expression of HH and low CD8+ T lymphocyte numbers, while the G-G-G haplotype was associated with a milder clinical expression of HH and high CD8+ T lymphocyte numbers. As CD8+ T lymphocytes are a very heterogeneous population, in this study we analysed the CD8+ subpopulations of naive, central memory (TCM) and effector memory (TEM), and further subsets of CD8+ TEM cells in 47 HH patients and 68 controls. In addition, association studies were conducted between the conserved haplotypes and the CD8+ T cell subpopulations in HH. Variations of the numbers of naive and central memory cells with age were similar between HH patients and controls. For TEM cells and the TEM CD27−CD28− subset no effect of age was observed in HH [R2 = 0·001, not significant (n.s.) and R2 = 0·01, n.s., respectively] contrasting with the increasing of these subpopulations with age in controls (R2 = 0·09, P = 0·017 and R2 = 0·22, P = 0·0005, respectively). Interestingly, patients homozygous for the A-A-T haplotype have lower numbers of CD8+ TEM cells due especially to lower numbers of TEM CD27−CD28− (0·206 ± 0·119 and 0·066 ± 0·067 × 106 cells/ml, respectively) than patients carrying the G-G-G haplotype (0·358 ± 0·195 and 0·246 ± 0·202 × 106 cells/ml, respectively). This may suggest an inability of HH patients to differentiate the CD8+ T cells into the most mature phenotype. PMID:20015273

  11. Molecular Determinants of Resistance Activation and Suppression by Phytophthora infestans Effector IPI-O

    PubMed Central

    Chen, Yu; Liu, Zhenyu; Halterman, Dennis A.

    2012-01-01

    Despite intensive breeding efforts, potato late blight, caused by the oomycete pathogen Phytophthora infestans, remains a threat to potato production worldwide because newly evolved pathogen strains have consistently overcome major resistance genes. The potato RB gene, derived from the wild species Solanum bulbocastanum, confers resistance to most P. infestans strains through recognition of members of the pathogen effector family IPI-O. While the majority of IPI-O proteins are recognized by RB to elicit resistance (e.g. IPI-O1, IPI-O2), some family members are able to elude detection (e.g. IPI-O4). In addition, IPI-O4 blocks recognition of IPI-O1, leading to inactivation of RB-mediated programmed cell death. Here, we report results that elucidate molecular mechanisms governing resistance elicitation or suppression of RB by IPI-O. Our data indicate self-association of the RB coiled coil (CC) domain as well as a physical interaction between this domain and the effectors IPI-O4 and IPI-O1. We identified four amino acids within IPI-O that are critical for interaction with the RB CC domain and one of these amino acids, at position 129, determines hypersensitive response (HR) elicitation in planta. IPI-O1 mutant L129P fails to induce HR in presence of RB while IPI-O4 P129L gains the ability to induce an HR. Like IPI-O4, IPI-O1 L129P is also able to suppress the HR mediated by RB, indicating a critical step in the evolution of this gene family. Our results point to a model in which IPI-O effectors can affect RB function through interaction with the RB CC domain. PMID:22438813

  12. Physical Activity Is Positively Associated with Episodic Memory in Aging.

    PubMed

    Hayes, Scott M; Alosco, Michael L; Hayes, Jasmeet P; Cadden, Margaret; Peterson, Kristina M; Allsup, Kelly; Forman, Daniel E; Sperling, Reisa A; Verfaellie, Mieke

    2015-11-01

    Aging is associated with performance reductions in executive function and episodic memory, although there is substantial individual variability in cognition among older adults. One factor that may be positively associated with cognition in aging is physical activity. To date, few studies have objectively assessed physical activity in young and older adults, and examined whether physical activity is differentially associated with cognition in aging. Young (n=29, age 18-31 years) and older adults (n=31, ages 55-82 years) completed standardized neuropsychological testing to assess executive function and episodic memory capacities. An experimental face-name relational memory task was administered to augment assessment of episodic memory. Physical activity (total step count and step rate) was objectively assessed using an accelerometer, and hierarchical regressions were used to evaluate relationships between cognition and physical activity. Older adults performed more poorly on tasks of executive function and episodic memory. Physical activity was positively associated with a composite measure of visual episodic memory and face-name memory accuracy in older adults. Physical activity associations with cognition were independent of sedentary behavior, which was negatively correlated with memory performance. Physical activity was not associated with cognitive performance in younger adults. Physical activity is positively associated with episodic memory performance in aging. The relationship appears to be strongest for face-name relational memory and visual episodic memory, likely attributable to the fact that these tasks make strong demands on the hippocampus. The results suggest that physical activity relates to cognition in older, but not younger adults.

  13. Development of a Pre-Prototype Power Assisted Glove End Effector for Extravehicular Activity

    NASA Technical Reports Server (NTRS)

    1986-01-01

    The purpose of this program was to develop an EVA power tool which is capable of performing a variety of functions while at the same time increasing the EVA crewmember's effectiveness by reducing hand fatigue associated with gripping tools through a pressurized EMU glove. The Power Assisted Glove End Effector (PAGE) preprototype hardware met or exceeded all of its technical requirements and has incorporated acoustic feedback to allow the EVA crewmember to monitor motor loading and speed. If this tool is to be developed for flight use, several issues need to be addressed. These issues are listed.

  14. The Escherichia coli effector EspJ blocks Src kinase activity via amidation and ADP ribosylation

    PubMed Central

    Young, Joanna C.; Clements, Abigail; Lang, Alexander E.; Garnett, James A.; Munera, Diana; Arbeloa, Ana; Pearson, Jaclyn; Hartland, Elizabeth L.; Matthews, Stephen J.; Mousnier, Aurelie; Barry, David J.; Way, Michael; Schlosser, Andreas; Aktories, Klaus; Frankel, Gad

    2014-01-01

    The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actin-rich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases. This leads to recruitment of the Nck–WIP–N-WASP complex that triggers Arp2/3-dependent actin polymerization in the host cell. The same phosphorylation-mediated signalling network is also assembled downstream of the Vaccinia virus protein A36 and the phagocytic Fc-gamma receptor FcγRIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and FcγRIIa. Consistent with this, EspJ inhibits actin polymerization downstream of EPEC, Vaccinia virus and opsonized red blood cells. We identify EspJ as a unique adenosine diphosphate (ADP) ribosyltransferase that directly inhibits Src kinase by simultaneous amidation and ADP ribosylation of the conserved kinase-domain residue, Src E310, resulting in glutamine-ADP ribose. PMID:25523213

  15. The Escherichia coli effector EspJ blocks Src kinase activity via amidation and ADP ribosylation.

    PubMed

    Young, Joanna C; Clements, Abigail; Lang, Alexander E; Garnett, James A; Munera, Diana; Arbeloa, Ana; Pearson, Jaclyn; Hartland, Elizabeth L; Matthews, Stephen J; Mousnier, Aurelie; Barry, David J; Way, Michael; Schlosser, Andreas; Aktories, Klaus; Frankel, Gad

    2014-01-01

    The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actin-rich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases. This leads to recruitment of the Nck-WIP-N-WASP complex that triggers Arp2/3-dependent actin polymerization in the host cell. The same phosphorylation-mediated signalling network is also assembled downstream of the Vaccinia virus protein A36 and the phagocytic Fc-gamma receptor FcγRIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and FcγRIIa. Consistent with this, EspJ inhibits actin polymerization downstream of EPEC, Vaccinia virus and opsonized red blood cells. We identify EspJ as a unique adenosine diphosphate (ADP) ribosyltransferase that directly inhibits Src kinase by simultaneous amidation and ADP ribosylation of the conserved kinase-domain residue, Src E310, resulting in glutamine-ADP ribose.

  16. Cellular Renewal and Improvement of Local Cell Effector Activity in Peritoneal Cavity in Response to Infectious Stimuli

    PubMed Central

    dos Anjos Cassado, Alexandra; de Albuquerque, José Antônio Tavares; Sardinha, Luiz Roberto; de Lima Buzzo, Carina; Faustino, Lucas; Nascimento, Rogério; Ghosn, Eliver Eid Bou; D'Império Lima, Maria Regina; Alvarez, Jose Maria Mosig; Bortoluci, Karina Ramalho

    2011-01-01

    The peritoneal cavity (PerC) is a singular compartment where many cell populations reside and interact. Despite the widely adopted experimental approach of intraperitoneal (i.p.) inoculation, little is known about the behavior of the different cell populations within the PerC. To evaluate the dynamics of peritoneal macrophage (MØ) subsets, namely small peritoneal MØ (SPM) and large peritoneal MØ (LPM), in response to infectious stimuli, C57BL/6 mice were injected i.p. with zymosan or Trypanosoma cruzi. These conditions resulted in the marked modification of the PerC myelo-monocytic compartment characterized by the disappearance of LPM and the accumulation of SPM and monocytes. In parallel, adherent cells isolated from stimulated PerC displayed reduced staining for β-galactosidase, a biomarker for senescence. Further, the adherent cells showed increased nitric oxide (NO) and higher frequency of IL-12-producing cells in response to subsequent LPS and IFN-γ stimulation. Among myelo-monocytic cells, SPM rather than LPM or monocytes, appear to be the central effectors of the activated PerC; they display higher phagocytic activity and are the main source of IL-12. Thus, our data provide a first demonstration of the consequences of the dynamics between peritoneal MØ subpopulations by showing that substitution of LPM by a robust SPM and monocytes in response to infectious stimuli greatly improves PerC effector activity. PMID:21799778

  17. Effector Activity of Peanut Allergens: A critical role for Ara h 2, Ara h 6, and their variants

    PubMed Central

    Porterfield, HS; Murray, KS; Schlichting, DG; Chen, X.; Hansen, KC; Duncan, MW; Dreskin, SC

    2009-01-01

    Rationale An important property of allergens is their ability to cross-link IgE and activate mast cells and basophils. The effector activity of peanut allergens has not been well characterized. Methods Crude extracts of fresh peanut flour were fractionated by gel filtration. Effector function was assayed by measuring degranulation of RBL SX-38 cells sensitized with IgE from individual sera and from pools of sera of peanut allergic donors. Results Following gel filtration, 75±7% of the applied protein and 76±16% (n=3) of the applied activity (assayed with a pool of 11 sera) were recovered in the resultant fractions. The majority (85±2%; n=3) of the recovered activity resided in a fraction with a theoretical average molecular weight of ~20 kD and a range of 13–25 kD. When all the individual fractions were recombined, the measured activity was similar to that of the original extract (140±43% when measured with a pool of serum (n=2) and 66±7% when measured with individual sera (n=4)); when all individual fractions excluding the 20 kD fraction were recombined, the measured activity was only 8±2% (n=2) of the original extract when assayed with the serum pool and 10±4% (n=3) when assayed with the individual sera. Two dimensional gel electrophoresis of this biologically active fraction by revealed >60 protein spots . Analysis of 50 of the most prominent spots by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry and of the full mixture by automated tandem mass spectrometry coupled to online capillary liquid chromatography revealed that greater than 97% of the protein mass consisted of Ara h 2.0101, Ara h 2.0201, Ara h 6 isoforms, and variants of these proteins. Conclusions Ara h 2 and Ara h 6 account for the majority of the effector activity found in a crude peanut extract. PMID:19438581

  18. Chlamydial Protease-Like Activity Factor and Type III Secreted Effectors Cooperate in Inhibition of p65 Nuclear Translocation

    PubMed Central

    Patton, Michael John; McCorrister, Stuart; Grant, Chris; Westmacott, Garrett; Fariss, Robert; Hu, Pingzhao; Zhao, Kaiqiong; Blake, Mary; Whitmire, Bill; Yang, Chunfu

    2016-01-01

    ABSTRACT The chlamydial protease-like activity factor (CPAF) is hypothesized to be an important secreted virulence factor; however, challenges in denaturing its proteolytic activity have hampered attempts to identify its legitimate targets. Here, we use a genetic and proteomic approach to identify authentic CPAF targets. Human epithelial cells infected with CPAF-sufficient and CPAF-deficient chlamydiae were lysed using known CPAF-denaturing conditions. Their protein profiles were analyzed using isobaric mass tags and liquid chromatography-tandem mass spectrometry. Comparative analysis of CPAF-sufficient and CPAF-deficient infections identified a limited number of CPAF host and chlamydial protein targets. Host targets were primarily interferon-stimulated gene products, whereas chlamydial targets were type III secreted proteins. We provide evidence supporting a cooperative role for CPAF and type III secreted effectors in blocking NF-κB p65 nuclear translocation, resulting in decreased beta interferon and proinflammatory cytokine synthesis. Genetic complementation of null organisms with CPAF restored p65 nuclear translocation inhibition and proteolysis of chlamydial type III secreted effector proteins (T3SEs). We propose that CPAF and T3SEs cooperate in the inhibition of host innate immunity. PMID:27677792

  19. Hemispheric Asymmetries in the Activation and Monitoring of Memory Errors

    ERIC Educational Resources Information Center

    Giammattei, Jeannette; Arndt, Jason

    2012-01-01

    Previous research on the lateralization of memory errors suggests that the right hemisphere's tendency to produce more memory errors than the left hemisphere reflects hemispheric differences in semantic activation. However, all prior research that has examined the lateralization of memory errors has used self-paced recognition judgments. Because…

  20. Adenovirus Vector-Induced CD8+ T Effector Memory Cell Differentiation and Recirculation, But Not Proliferation, Are Important for Protective Immunity Against Experimental Trypanosoma cruzi Infection

    PubMed Central

    Vasconcelos, José Ronnie; Dominguez, Mariana R.; Neves, Ramon L.; Ersching, Jonatan; Araújo, Adriano; Santos, Luara I.; Virgilio, Fernando S.; Machado, Alexandre V.; Bruna-Romero, Oscar; Gazzinelli, Ricardo T.

    2014-01-01

    Abstract Heterologous prime-boost vaccination using plasmid DNA followed by replication-defective adenovirus vector generates a large number of specific CD8+ T effector memory (TEM) cells that provide long-term immunity against a variety of pathogens. In the present study, we initially characterized the frequency, phenotype, and function of these T cells in vaccinated mice that were subjected to infectious challenge with the human protozoan parasite Trypanosoma cruzi. We observed that the frequency of the specific CD8+ T cells in the spleens of the vaccinated mice increased after challenge. Specific TEM cells differentiated into cells with a KLRG1High CD27Low CD43Low CD183LowT-betHigh EomesLow phenotype and capable to produce simultaneously the antiparasitic mediators IFNγ and TNF. Using the gzmBCreERT2/ROSA26EYFP transgenic mouse line, in which the cells that express Granzyme B after immunization, are indelibly labeled with enhanced yellow fluorescent protein, we confirmed that CD8+ T cells present after challenge were indeed TEM cells that had been induced by vaccination. Subsequently, we observed that the in vivo increase in the frequency of the specific CD8+ T cells was not because of an anamnestic immune response. Most importantly, after challenge, the increase in the frequency of specific cells and the protective immunity they mediate were insensitive to treatment with the cytostatic toxic agent hydroxyurea. We have previously described that the administration of the drug FTY720, which reduces lymphocyte recirculation, severely impairs protective immunity, and our evidence supports the model that when large amounts of antigen-experienced CD8+ TEM cells are present after heterologous prime-boost vaccination, differentiation, and recirculation, rather than proliferation, are key for the resultant protective immunity. PMID:24568548

  1. Patient-adapted, specific activation of HIV-1 by customized TAL effectors (TALEs), a proof of principle study.

    PubMed

    Geissler, Rene; Hauber, Ilona; Funk, Nancy; Richter, Annekatrin; Behrens, Martina; Renner, Ivonne; Chemnitz, Jan; Hofmann-Sieber, Helga; Baum, Heidi; van Lunzen, Jan; Boch, Jens; Hauber, Joachim; Behrens, Sven-Erik

    2015-12-01

    The major obstacle to cure infections with human immunodeficiency virus (HIV-1) is integrated proviral genomes, which are not eliminated by antiretroviral therapies (ART). Treatment approaches with latency-reversing agents (LRAs) aim at inducing provirus expression to tag latently-infected cells for clearance through viral cytopathic effects or cytotoxic T cell (CTL) responses. However, the currently tested LRAs reveal evident drawbacks as gene expression is globally induced and viral outgrowth is insecure. Here, we present transcription activator-like effector (TALE) proteins as potent tools to activate HIV-1 specifically. The large variety of circulating HIV-1 strains and, accordingly, integrated proviruses was addressed by the programmable DNA-specificity of TALEs. Using customized engineered TALEs, a substantial transcription activation and viral outgrowth was achieved with cells obtained from different HIV-1 patients. Our data suggest that TALEs may be useful tools in future strategies aimed at removing HIV-1 reservoirs. PMID:26474371

  2. Implications of Spatiotemporal Regulation of Shigella flexneri Type Three Secretion Activity on Effector Functions: Think Globally, Act Locally.

    PubMed

    Campbell-Valois, F-X; Pontier, Stéphanie M

    2016-01-01

    Shigella spp. are Gram-negative bacterial pathogens that infect human colonic epithelia and cause bacterial dysentery. These bacteria express multiple copies of a syringe-like protein complex, the Type Three Secretion apparatus (T3SA), which is instrumental in the etiology of the disease. The T3SA triggers the plasma membrane (PM) engulfment of the bacteria by host cells during the initial entry process. It then enables bacteria to escape the resulting phagocytic-like vacuole. Freed bacteria form actin comets to move in the cytoplasm, which provokes bacterial collision with the inner leaflet of the PM. This phenomenon culminates in T3SA-dependent secondary uptake and vacuolar rupture in neighboring cells in a process akin to what is observed during entry and named cell-to-cell spread. The activity of the T3SA of Shigella flexneri was recently demonstrated to display an on/off regulation during the infection. While the T3SA is active when bacteria are in contact with PM-derived compartments, it switches to an inactive state when bacteria are released within the cytosol. These observations indicate that effector proteins transiting through the T3SA are therefore translocated in a highly time and space constrained fashion, likely impacting on their cellular distribution. Herein, we present what is currently known about the composition, the assembly and the regulation of the T3SA activity and discuss the consequences of the on/off regulation of T3SA on Shigella effector properties and functions during the infection. Specific examples that will be developed include the role of effectors IcsB and VirA in the escape from LC3/ATG8-positive vacuoles formed during cell-to-cell spread and of IpaJ protease activity against N-miristoylated proteins. The conservation of a similar regulation of T3SA activity in other pathogens such as Salmonella or Enteropathogenic Escherichia coli will also be briefly discussed. PMID:27014638

  3. Implications of Spatiotemporal Regulation of Shigella flexneri Type Three Secretion Activity on Effector Functions: Think Globally, Act Locally

    PubMed Central

    Campbell-Valois, F.-X.; Pontier, Stéphanie M.

    2016-01-01

    Shigella spp. are Gram-negative bacterial pathogens that infect human colonic epithelia and cause bacterial dysentery. These bacteria express multiple copies of a syringe-like protein complex, the Type Three Secretion apparatus (T3SA), which is instrumental in the etiology of the disease. The T3SA triggers the plasma membrane (PM) engulfment of the bacteria by host cells during the initial entry process. It then enables bacteria to escape the resulting phagocytic-like vacuole. Freed bacteria form actin comets to move in the cytoplasm, which provokes bacterial collision with the inner leaflet of the PM. This phenomenon culminates in T3SA-dependent secondary uptake and vacuolar rupture in neighboring cells in a process akin to what is observed during entry and named cell-to-cell spread. The activity of the T3SA of Shigella flexneri was recently demonstrated to display an on/off regulation during the infection. While the T3SA is active when bacteria are in contact with PM-derived compartments, it switches to an inactive state when bacteria are released within the cytosol. These observations indicate that effector proteins transiting through the T3SA are therefore translocated in a highly time and space constrained fashion, likely impacting on their cellular distribution. Herein, we present what is currently known about the composition, the assembly and the regulation of the T3SA activity and discuss the consequences of the on/off regulation of T3SA on Shigella effector properties and functions during the infection. Specific examples that will be developed include the role of effectors IcsB and VirA in the escape from LC3/ATG8-positive vacuoles formed during cell-to-cell spread and of IpaJ protease activity against N-miristoylated proteins. The conservation of a similar regulation of T3SA activity in other pathogens such as Salmonella or Enteropathogenic Escherichia coli will also be briefly discussed. PMID:27014638

  4. Multiple Activities of the Plant Pathogen Type III Effector Proteins WtsE and AvrE1 require WxxxE Motifs

    PubMed Central

    Ham, Jong Hyun; Majerczak, Doris R.; Nomura, Kinya; Mecey, Christy; Uribe, Francisco; He, Sheng-Yang; Mackey, David; Coplin, David L.

    2009-01-01

    The broadly conserved AvrE-family of type III effectors from Gram-negative plant pathogenic bacteria includes important virulence factors, yet little is known about the mechanisms by which these effectors function inside plant cells to promote disease. We have identified two conserved motifs in AvrE-family effectors: a WxxxE motif and a putative C-terminal endoplasmic reticulum membrane retention/retrieval signal (ERMRS). The WxxxE and ERMRS motifs are both required for the virulence activities of WtsE and AvrE1, which are major virulence factors of the corn pathogen Pantoea stewartii subsp. stewartii and the tomato/Arabidopsis pathogen Pseudomonas syringae pv. tomato, respectively. The WxxxE and the predicted ERMRS motifs are also required for other biological activities of WtsE, including elicitation of the hypersensitive response in nonhost plants and suppression of defense responses in Arabidopsis. A family of type III effectors from mammalian bacterial pathogens requires WxxxE and sub-cellular targeting motifs for virulence functions that involve their ability to mimic activated G-proteins. The conservation of related motifs and their necessity for the function of type III effectors from plant pathogens indicates that disturbing host pathways by mimicking activated host G-proteins may be a virulence mechanism employed by plant pathogens as well. PMID:19445595

  5. The Influence of Acute Physical Activity on Working Memory.

    PubMed

    Zach, Sima; Shalom, Eyal

    2016-04-01

    The effect of three types of physical activity on two types of working memory were investigated. Participants were 20 adult males who trained twice a week in volleyball two hours per session. Procedures included two pre and post intervention tests of working memory: the Digit span and Visual Memory Span subtests of the Wechsler Memory Scale-Revised. Interventions included tactical volleyball formation, body-weight resistance exercises, 15 minutes of running, and sub-maximal aerobic activity. Volleyball activity improved memory performance to a greater extent than the other two activities. Results indicate that immediately after acute exercise there is an increase in working memory function, more evident after physical activity in which cognitive functioning is inherent. PMID:27166321

  6. The Influence of Acute Physical Activity on Working Memory.

    PubMed

    Zach, Sima; Shalom, Eyal

    2016-04-01

    The effect of three types of physical activity on two types of working memory were investigated. Participants were 20 adult males who trained twice a week in volleyball two hours per session. Procedures included two pre and post intervention tests of working memory: the Digit span and Visual Memory Span subtests of the Wechsler Memory Scale-Revised. Interventions included tactical volleyball formation, body-weight resistance exercises, 15 minutes of running, and sub-maximal aerobic activity. Volleyball activity improved memory performance to a greater extent than the other two activities. Results indicate that immediately after acute exercise there is an increase in working memory function, more evident after physical activity in which cognitive functioning is inherent.

  7. The Activation of Phytophthora Effector Avr3b by Plant Cyclophilin is Required for the Nudix Hydrolase Activity of Avr3b.

    PubMed

    Kong, Guanghui; Zhao, Yao; Jing, Maofeng; Huang, Jie; Yang, Jin; Xia, Yeqiang; Kong, Liang; Ye, Wenwu; Xiong, Qin; Qiao, Yongli; Dong, Suomeng; Ma, Wenbo; Wang, Yuanchao

    2015-08-01

    Plant pathogens secrete an arsenal of effector proteins to impair host immunity. Some effectors possess enzymatic activities that can modify their host targets. Previously, we demonstrated that a Phytophthora sojae RXLR effector Avr3b acts as a Nudix hydrolase when expressed in planta; and this enzymatic activity is required for full virulence of P. sojae strain P6497 in soybean (Glycine max). Interestingly, recombinant Avr3b produced by E. coli does not have the hydrolase activity unless it was incubated with plant protein extracts. Here, we report the activation of Avr3b by a prolyl-peptidyl isomerase (PPIase), cyclophilin, in plant cells. Avr3b directly interacts with soybean cyclophilin GmCYP1, which activates the hydrolase activity of Avr3b in a PPIase activity-dependent manner. Avr3b contains a putative Glycine-Proline (GP) motif; which is known to confer cyclophilin-binding in other protein substrates. Substitution of the Proline (P132) in the putative GP motif impaired the interaction of Avr3b with GmCYP1; as a result, the mutant Avr3bP132A can no longer be activated by GmCYP1, and is also unable to promote Phytophthora infection. Avr3b elicits hypersensitive response (HR) in soybean cultivars producing the resistance protein Rps3b, but Avr3bP132A lost its ability to trigger HR. Furthermore, silencing of GmCYP1 rendered reduced cell death triggered by Avr3b, suggesting that GmCYP1-mediated Avr3b maturation is also required for Rps3b recognition. Finally, cyclophilins of Nicotiana benthamiana can also interact with Avr3b and activate its enzymatic activity. Overall, our results demonstrate that cyclophilin is a "helper" that activates the enzymatic activity of Avr3b after it is delivered into plant cells; as such, cyclophilin is required for the avirulence and virulence functions of Avr3b.

  8. Subthreshold IKK activation modulates the effector functions of primary mast cells and allows specific targeting of transformed mast cells.

    PubMed

    Drube, Sebastian; Weber, Franziska; Loschinski, Romy; Beyer, Mandy; Rothe, Mandy; Rabenhorst, Anja; Göpfert, Christiane; Meininger, Isabel; Diamanti, Michaela A; Stegner, David; Häfner, Norman; Böttcher, Martin; Reinecke, Kirstin; Herdegen, Thomas; Greten, Florian R; Nieswandt, Bernhard; Hartmann, Karin; Krämer, Oliver H; Kamradt, Thomas

    2015-03-10

    Mast cell differentiation and proliferation depends on IL-3. IL-3 induces the activation of MAP-kinases and STATs and consequently induces proliferation and survival. Dysregulation of IL-3 signaling pathways also contribute to inflammation and tumorigenesis. We show here that IL-3 induces a SFK- and Ca²⁺-dependent activation of the inhibitor of κB kinases 2 (IKK2) which results in mast cell proliferation and survival but does not induce IκBα-degradation and NFκB activation. Therefore we propose the term "subthreshold IKK activation".This subthreshold IKK activation also primes mast cells for enhanced responsiveness to IL-33R signaling. Consequently, co-stimulation with IL-3 and IL-33 increases IKK activation and massively enhances cytokine production induced by IL-33.We further reveal that in neoplastic mast cells expressing constitutively active Ras, subthreshold IKK activation is associated with uncontrolled proliferation. Consequently, pharmacological IKK inhibition reduces tumor growth selectively by inducing apoptosis in vivo.Together, subthreshold IKK activation is crucial to mediate the full IL-33-induced effector functions in primary mast cells and to mediate uncontrolled proliferation of neoplastic mast cells. Thus, IKK2 is a new molecularly defined target structure. PMID:25749030

  9. Heritability of working memory brain activation.

    PubMed

    Blokland, Gabriëlla A M; McMahon, Katie L; Thompson, Paul M; Martin, Nicholas G; de Zubicaray, Greig I; Wright, Margaret J

    2011-07-27

    Although key to understanding individual variation in task-related brain activation, the genetic contribution to these individual differences remains largely unknown. Here we report voxel-by-voxel genetic model fitting in a large sample of 319 healthy, young adult, human identical and fraternal twins (mean ± SD age, 23.6 ± 1.8 years) who performed an n-back working memory task during functional magnetic resonance imaging (fMRI) at a high magnetic field (4 tesla). Patterns of task-related brain response (BOLD signal difference of 2-back minus 0-back) were significantly heritable, with the highest estimates (40-65%) in the inferior, middle, and superior frontal gyri, left supplementary motor area, precentral and postcentral gyri, middle cingulate cortex, superior medial gyrus, angular gyrus, superior parietal lobule, including precuneus, and superior occipital gyri. Furthermore, high test-retest reliability for a subsample of 40 twins indicates that nongenetic variance in the fMRI brain response is largely due to unique environmental influences rather than measurement error. Individual variations in activation of the working memory network are therefore significantly influenced by genetic factors. By establishing the heritability of cognitive brain function in a large sample that affords good statistical power, and using voxel-by-voxel analyses, this study provides the necessary evidence for task-related brain activation to be considered as an endophenotype for psychiatric or neurological disorders, and represents a substantial new contribution to the field of neuroimaging genetics. These genetic brain maps should facilitate discovery of gene variants influencing cognitive brain function through genome-wide association studies, potentially opening up new avenues in the treatment of brain disorders. PMID:21795540

  10. The Agrobacterium VirE3 effector protein: a potential plant transcriptional activator.

    PubMed

    García-Rodríguez, Fernando M; Schrammeijer, Barbara; Hooykaas, Paul J J

    2006-01-01

    During the infection of plants, Agrobacterium tumefaciens introduces several Virulence proteins including VirE2, VirF, VirD5 and VirE3 into plant cells in addition to the T-DNA. Here, we report that double mutation of virF and virE3 leads to strongly diminished tumor formation on tobacco, tomato and sunflower. The VirE3 protein is translated from a polycistronic mRNA containing the virE1, virE2 and virE3 genes, in Agrobacterium. The VirE3 protein has nuclear localization sequences, which suggests that it is transported into the plant cell nucleus upon translocation. Indeed we show here that VirE3 interacts in vitro with importin-alpha and that a VirE3-GFP fusion protein is localized in the nucleus. VirE3 also interacts with two other proteins, viz. pCsn5, a component of the COP9 signalosome and pBrp, a plant specific general transcription factor belonging to the TFIIB family. We found that VirE3 is able to induce transcription in yeast when bound to DNA through the GAL4-BD. Our data indicate that the translocated effector protein VirE3 is transported into the nucleus and there it may interact with the transcription factor pBrp to induce the expression of genes needed for tumor development. PMID:17130174

  11. Association of lymph-node antigens with lower Gag-specific central-memory and higher Env-specific effector-memory CD8+ T-cell frequencies in a macaque AIDS model

    PubMed Central

    Ishii, Hiroshi; Matsuoka, Saori; Nomura, Takushi; Nakamura, Midori; Shiino, Teiichiro; Sato, Yuko; Iwata-Yoshikawa, Naoko; Hasegawa, Hideki; Mizuta, Kazuta; Sakawaki, Hiromi; Miura, Tomoyuki; Koyanagi, Yoshio; Naruse, Taeko K.; Kimura, Akinori; Matano, Tetsuro

    2016-01-01

    Virus-specific CD8+ T cells exert strong suppressive pressure on human/simian immunodeficiency virus (HIV/SIV) replication. These responses have been intensively examined in peripheral blood mononuclear cells (PBMCs) but not fully analyzed in lymph nodes (LNs), where interaction between CD8+ T cells and HIV/SIV-infected cells occurs. Here, we investigated target antigen specificity of CD8+ T cells in LNs in a macaque AIDS model. Analysis of virus antigen-specific CD8+ T-cell responses in the inguinal LNs obtained from twenty rhesus macaques in the chronic phase of SIV infection showed an inverse correlation between viral loads and frequencies of CD8+ T cells with CD28+ CD95+ central memory phenotype targeting the N-terminal half of SIV core antigen (Gag-N). In contrast, analysis of LNs but not PBMCs revealed a positive correlation between viral loads and frequencies of CD8+ T cells with CD28−CD95+ effector memory phenotype targeting the N-terminal half of SIV envelope (Env-N), soluble antigen. Indeed, LNs with detectable SIV capsid p27 antigen in the germinal center exhibited significantly lower Gag-N-specific CD28+ CD95+ CD8+ T-cell and higher Env-N-specific CD28−CD95+ CD8+ T-cell responses than those without detectable p27. These results imply that core and envelope antigen-specific CD8+ T cells show different patterns of interactions with HIV/SIV-infected cells. PMID:27452272

  12. Subthreshold IKK activation modulates the effector functions of primary mast cells and allows specific targeting of transformed mast cells

    PubMed Central

    Drube, Sebastian; Beyer, Mandy; Rothe, Mandy; Rabenhorst, Anja; Göpfert, Christiane; Meininger, Isabel; Diamanti, Michaela A.; Stegner, David; Häfner, Norman; Böttcher, Martin; Reinecke, Kirstin; Herdegen, Thomas; Greten, Florian R.; Nieswandt, Bernhard; Hartmann, Karin; Krämer, Oliver H.; Kamradt, Thomas

    2015-01-01

    Mast cell differentiation and proliferation depends on IL-3. IL-3 induces the activation of MAP-kinases and STATs and consequently induces proliferation and survival. Dysregulation of IL-3 signaling pathways also contribute to inflammation and tumorigenesis. We show here that IL-3 induces a SFK- and Ca2+-dependent activation of the inhibitor of κB kinases 2 (IKK2) which results in mast cell proliferation and survival but does not induce IκBα-degradation and NFκB activation. Therefore we propose the term “subthreshold IKK activation”. This subthreshold IKK activation also primes mast cells for enhanced responsiveness to IL-33R signaling. Consequently, co-stimulation with IL-3 and IL-33 increases IKK activation and massively enhances cytokine production induced by IL-33. We further reveal that in neoplastic mast cells expressing constitutively active Ras, subthreshold IKK activation is associated with uncontrolled proliferation. Consequently, pharmacological IKK inhibition reduces tumor growth selectively by inducing apoptosis in vivo. Together, subthreshold IKK activation is crucial to mediate the full IL-33-induced effector functions in primary mast cells and to mediate uncontrolled proliferation of neoplastic mast cells. Thus, IKK2 is a new molecularly defined target structure. PMID:25749030

  13. Shape memory polymers for active cell culture.

    PubMed

    Davis, Kevin A; Luo, Xiaofan; Mather, Patrick T; Henderson, James H

    2011-07-04

    Shape memory polymers (SMPs) are a class of "smart" materials that have the ability to change from a fixed, temporary shape to a pre-determined permanent shape upon the application of a stimulus such as heat(1-5). In a typical shape memory cycle, the SMP is first deformed at an elevated temperature that is higher than its transition temperature, T(trans;) [either the melting temperature (T(m;)) or the glass transition temperature (T(g;))]. The deformation is elastic in nature and mainly leads to a reduction in conformational entropy of the constituent network chains (following the rubber elasticity theory). The deformed SMP is then cooled to a temperature below its T(trans;) while maintaining the external strain or stress constant. During cooling, the material transitions to a more rigid state (semi-crystalline or glassy), which kinetically traps or "freezes" the material in this low-entropy state leading to macroscopic shape fixing. Shape recovery is triggered by continuously heating the material through T(trans;) under a stress-free (unconstrained) condition. By allowing the network chains (with regained mobility) to relax to their thermodynamically favored, maximal-entropy state, the material changes from the temporary shape to the permanent shape. Cells are capable of surveying the mechanical properties of their surrounding environment(6). The mechanisms through which mechanical interactions between cells and their physical environment control cell behavior are areas of active research. Substrates of defined topography have emerged as powerful tools in the investigation of these mechanisms. Mesoscale, microscale, and nanoscale patterns of substrate topography have been shown to direct cell alignment, cell adhesion, and cell traction forces(7-14). These findings have underscored the potential for substrate topography to control and assay the mechanical interactions between cells and their physical environment during cell culture, but the substrates used to date

  14. Stress-Induced Out-of-Context Activation of Memory

    PubMed Central

    Ježek, Karel; Lee, Benjamin B.; Kelemen, Eduard; McCarthy, Katharine M.; McEwen, Bruce S.; Fenton, André A.

    2010-01-01

    Inappropriate recollections and responses in stressful conditions are hallmarks of post-traumatic stress disorder and other anxiety and mood disorders, but how stress contributes to the disorders is unclear. Here we show that stress itself reactivates memories even if the memory is unrelated to the stressful experience. Forced-swim stress one day after learning enhanced memory recall. One-day post-learning amnestic treatments were ineffective unless administered soon after the swim, indicating that a stressful experience itself can reactivate unrelated consolidated memories. The swim also triggered inter-hemispheric transfer of a lateralized memory, confirming stress reactivates stable memories. These novel effects of stress on memory required the hippocampus although the memories themselves did not, indicating hippocampus-dependent modulation of extrahippocampal memories. These findings that a stressful experience itself can activate memory suggest the novel hypothesis that traumatic stress reactivates pre-trauma memories, linking them to memory for the trauma and pathological facilitation of post-traumatic recall. PMID:21203585

  15. False memory for face in short-term memory and neural activity in human amygdala.

    PubMed

    Iidaka, Tetsuya; Harada, Tokiko; Sadato, Norihiro

    2014-12-01

    Human memory is often inaccurate. Similar to words and figures, new faces are often recognized as seen or studied items in long- and short-term memory tests; however, the neural mechanisms underlying this false memory remain elusive. In a previous fMRI study using morphed faces and a standard false memory paradigm, we found that there was a U-shaped response curve of the amygdala to old, new, and lure items. This indicates that the amygdala is more active in response to items that are salient (hit and correct rejection) compared to items that are less salient (false alarm), in terms of memory retrieval. In the present fMRI study, we determined whether the false memory for faces occurs within the short-term memory range (a few seconds), and assessed which neural correlates are involved in veridical and illusory memories. Nineteen healthy participants were scanned by 3T MRI during a short-term memory task using morphed faces. The behavioral results indicated that the occurrence of false memories was within the short-term range. We found that the amygdala displayed a U-shaped response curve to memory items, similar to those observed in our previous study. These results suggest that the amygdala plays a common role in both long- and short-term false memory for faces. We made the following conclusions: First, the amygdala is involved in detecting the saliency of items, in addition to fear, and supports goal-oriented behavior by modulating memory. Second, amygdala activity and response time might be related with a subject's response criterion for similar faces.

  16. Activated macrophage conditioned medium: identification of the soluble factors inducing cytotoxicity and the L-arginine dependent effector mechanism.

    PubMed

    Amber, I J; Hibbs, J B; Parker, C J; Johnson, B B; Taintor, R R; Vavrin, Z

    1991-06-01

    Conditioned medium (CM) from cultures of cytotoxic activated macrophages causes inhibition of mitochondrial respiration, DNA synthesis, and aconitase activity in murine EMT-6 mammary adenocarcinoma cells by an L-arginine dependent effector mechanism. CM induces cytotoxicity and nitrite synthesis in EMT-6 cells in a dose dependent manner. We have identified the soluble factors in CM that induce cytotoxicity and synthesis of inorganic nitrogen oxides from L-arginine by EMT-6 cells. Using functional inhibition experiments, the activity of lipopolysaccharide (LPS), tumor necrosis factor alpha (TNF alpha), and interferon gamma (IFN gamma) in CM was investigated. The LPS inhibitor polymyxin B and TNF alpha antibody produced a modest decrease in nitrite production, while IFN gamma antibody markedly inhibited both nitrite production and cytostasis. Simultaneous treatment with polymyxin B, TNF alpha antibody, and IFN gamma antibody reduced EMT-6 cell nitrite production by 81%, and cytostasis by 74%. By Western blot, IFN gamma and TNF alpha were shown to be present in CM. When CM was subjected to hydrophobic interaction chromatography, a single peak of activity was eluted, and Western blot showed that the active fractions contained IFN gamma. Furthermore, IFN gamma antibody neutralized the activity in these chromatographic fractions. We conclude that induction of inorganic nitrogen oxide synthesis from L-arginine by the synergistic combination of IFN gamma, TNF alpha, and LPS accounts for most of the biologic activity of CM, and that IFN gamma is the major priming factor. PMID:1902865

  17. Physical Activity Is Positively Associated with Episodic Memory in Aging

    PubMed Central

    Hayes, Scott M.; Alosco, Michael L.; Hayes, Jasmeet P.; Cadden, Margaret; Peterson, Kristina M.; Allsup, Kelly; Forman, Daniel E.; Sperling, Reisa A.; Verfaellie, Mieke

    2016-01-01

    Aging is associated with performance reductions in executive function and episodic memory, although there is substantial individual variability in cognition among older adults. One factor that may be positively associated with cognition in aging is physical activity. To date, few studies have objectively assessed physical activity in young and older adults, and examined whether physical activity is differentially associated with cognition in aging. Young (n = 29, age 18–31 years) and older adults (n = 31, ages 55–82 years) completed standardized neuropsychological testing to assess executive function and episodic memory capacities. An experimental face-name relational memory task was administered to augment assessment of episodic memory. Physical activity (total step count and step rate) was objectively assessed using an accelerometer, and hierarchical regressions were used to evaluate relationships between cognition and physical activity. Older adults performed more poorly on tasks of executive function and episodic memory. Physical activity was positively associated with a composite measure of visual episodic memory and face-name memory accuracy in older adults. Physical activity associations with cognition were independent of sedentary behavior, which was negatively correlated with memory performance. Physical activity was not associated with cognitive performance in younger adults. Physical activity is positively associated with episodic memory performance in aging. The relationship appears to be strongest for face-name relational memory and visual episodic memory, likely attributable to the fact that these tasks make strong demands on the hippocampus. The results suggest that physical activity relates to cognition in older, but not younger adults. PMID:26581790

  18. Parallel effects of memory set activation and search on timing and working memory capacity

    PubMed Central

    Schweickert, Richard; Fortin, Claudette; Xi, Zhuangzhuang; Viau-Quesnel, Charles

    2014-01-01

    Accurately estimating a time interval is required in everyday activities such as driving or cooking. Estimating time is relatively easy, provided a person attends to it. But a brief shift of attention to another task usually interferes with timing. Most processes carried out concurrently with timing interfere with it. Curiously, some do not. Literature on a few processes suggests a general proposition, the Timing and Complex-Span Hypothesis: A process interferes with concurrent timing if and only if process performance is related to complex span. Complex-span is the number of items correctly recalled in order, when each item presented for study is followed by a brief activity. Literature on task switching, visual search, memory search, word generation and mental time travel supports the hypothesis. Previous work found that another process, activation of a memory set in long term memory, is not related to complex-span. If the Timing and Complex-Span Hypothesis is true, activation should not interfere with concurrent timing in dual-task conditions. We tested such activation in single-task memory search task conditions and in dual-task conditions where memory search was executed with concurrent timing. In Experiment 1, activating a memory set increased reaction time, with no significant effect on time production. In Experiment 2, set size and memory set activation were manipulated. Activation and set size had a puzzling interaction for time productions, perhaps due to difficult conditions, leading us to use a related but easier task in Experiment 3. In Experiment 3 increasing set size lengthened time production, but memory activation had no significant effect. Results here and in previous literature on the whole support the Timing and Complex-Span Hypotheses. Results also support a sequential organization of activation and search of memory. This organization predicts activation and set size have additive effects on reaction time and multiplicative effects on percent

  19. Maintenance of Serological Memory by Polyclonal Activation of Human Memory B Cells

    NASA Astrophysics Data System (ADS)

    Bernasconi, Nadia L.; Traggiai, Elisabetta; Lanzavecchia, Antonio

    2002-12-01

    Production of antibodies can last for a lifetime, through mechanisms that remain poorly understood. Here, we show that human memory B lymphocytes proliferate and differentiate into plasma cells in response to polyclonal stimuli, such as bystander T cell help and CpG DNA. Furthermore, plasma cells secreting antibodies to recall antigens are produced in vivo at levels proportional to the frequency of specific memory B cells, even several years after antigenic stimulation. Although antigen boosting leads to a transient increase in specific antibody levels, ongoing polyclonal activation of memory B cells offers a means to maintain serological memory for a human lifetime.

  20. Developmental Differences in Prefrontal Activation during Working Memory Maintenance and Manipulation for Different Memory Loads

    ERIC Educational Resources Information Center

    Jolles, Dietsje D.; Kleibeuker, Sietske W.; Rombouts, Serge A. R. B.; Crone, Eveline A.

    2011-01-01

    The ability to keep information active in working memory is one of the cornerstones of cognitive development. Prior studies have demonstrated that regions which are important for working memory performance in adults, such as dorsolateral prefrontal cortex (DLPFC), ventrolateral prefrontal cortex (VLPFC), and superior parietal cortex, become…

  1. The Rac GTPase effector p21-activated kinase is essential for hematopoietic stem/progenitor cell migration and engraftment.

    PubMed

    Dorrance, Adrienne M; De Vita, Serena; Radu, Maria; Reddy, Pavankumar N G; McGuinness, Meaghan K; Harris, Chad E; Mathieu, Ronald; Lane, Steven W; Kosoff, Rachelle; Milsom, Michael D; Chernoff, Jonathan; Williams, David A

    2013-03-28

    The p21-activated kinases (Paks) are serine/threonine kinases that are major effectors of the Rho guanosine 5'\\x{2011}triphosphatase, Rac, and Cdc42. Rac and Cdc42 are known regulators of hematopoietic stem and progenitor cell (HSPC) function, however, a direct role for Paks in HSPCs has yet to be elucidated. Lin(-)Sca1(+)c-kit(+) (LSK) cells from wild-type mice were transduced with retrovirus expressing Pak inhibitory domain (PID), a well-characterized inhibitor of Pak activation. Defects in marrow homing and in vitro cell migration, assembly of the actin cytoskeleton, proliferation, and survival were associated with engraftment failure of PID-LSK. The PID-LSK demonstrated decreased phosphorylation of extracellular signal-regulated kinase (ERK), whereas constitutive activation of ERK in these cells led to rescue of hematopoietic progenitor cell proliferation in vitro and partial rescue of Pak-deficient HSPC homing and engraftment in vivo. Using conditional knock-out mice, we demonstrate that among group A Paks, Pak2(-/-) HSPC show reduced homing to the bone marrow and altered cell shape similar to PID-LSK cells in vitro and are completely defective in HSPC engraftment. These data demonstrate that Pak proteins are key components of multiple engraftment-associated HSPC functions and play a direct role in activation of ERK in HSPCs, and that Pak2 is specifically essential for HSPC engraftment.

  2. Molecular and Cellular Mechanisms for Trapping and Activating Emotional Memories

    PubMed Central

    Cai, Denise J.; Sano, Yoshitake; Lee, Yong-Seok; Zhou, Yu; Bekal, Pallavi; Deisseroth, Karl; Silva, Alcino J.

    2016-01-01

    Recent findings suggest that memory allocation to specific neurons (i.e., neuronal allocation) in the amygdala is not random, but rather the transcription factor cAMP-response element binding protein (CREB) modulates this process, perhaps by regulating the transcription of channels that control neuronal excitability. Here, optogenetic studies in the mouse lateral amygdala (LA) were used to demonstrate that CREB and neuronal excitability regulate which neurons encode an emotional memory. To test the role of CREB in memory allocation, we overexpressed CREB in the lateral amygdala to recruit the encoding of an auditory-fear conditioning (AFC) memory to a subset of neurons. Then, post-training activation of these neurons with Channelrhodopsin-2 was sufficient to trigger recall of the memory for AFC, suggesting that CREB regulates memory allocation. To test the role of neuronal excitability in memory allocation, we used a step function opsin (SFO) to transiently increase neuronal excitability in a subset of LA neurons during AFC. Post-training activation of these neurons with Volvox Channelrhodopsin-1 was able to trigger recall of that memory. Importantly, our studies show that activation of the SFO did not affect AFC by either increasing anxiety or by strengthening the unconditioned stimulus. Our findings strongly support the hypothesis that CREB regulates memory allocation by modulating neuronal excitability. PMID:27579481

  3. Molecular and Cellular Mechanisms for Trapping and Activating Emotional Memories.

    PubMed

    Rogerson, Thomas; Jayaprakash, Balaji; Cai, Denise J; Sano, Yoshitake; Lee, Yong-Seok; Zhou, Yu; Bekal, Pallavi; Deisseroth, Karl; Silva, Alcino J

    2016-01-01

    Recent findings suggest that memory allocation to specific neurons (i.e., neuronal allocation) in the amygdala is not random, but rather the transcription factor cAMP-response element binding protein (CREB) modulates this process, perhaps by regulating the transcription of channels that control neuronal excitability. Here, optogenetic studies in the mouse lateral amygdala (LA) were used to demonstrate that CREB and neuronal excitability regulate which neurons encode an emotional memory. To test the role of CREB in memory allocation, we overexpressed CREB in the lateral amygdala to recruit the encoding of an auditory-fear conditioning (AFC) memory to a subset of neurons. Then, post-training activation of these neurons with Channelrhodopsin-2 was sufficient to trigger recall of the memory for AFC, suggesting that CREB regulates memory allocation. To test the role of neuronal excitability in memory allocation, we used a step function opsin (SFO) to transiently increase neuronal excitability in a subset of LA neurons during AFC. Post-training activation of these neurons with Volvox Channelrhodopsin-1 was able to trigger recall of that memory. Importantly, our studies show that activation of the SFO did not affect AFC by either increasing anxiety or by strengthening the unconditioned stimulus. Our findings strongly support the hypothesis that CREB regulates memory allocation by modulating neuronal excitability. PMID:27579481

  4. Role of Prefrontal Persistent Activity in Working Memory

    PubMed Central

    Riley, Mitchell R.; Constantinidis, Christos

    2016-01-01

    The prefrontal cortex is activated during working memory, as evidenced by fMRI results in human studies and neurophysiological recordings in animal models. Persistent activity during the delay period of working memory tasks, after the offset of stimuli that subjects are required to remember, has traditionally been thought of as the neural correlate of working memory. In the last few years several findings have cast doubt on the role of this activity. By some accounts, activity in other brain areas, such as the primary visual and posterior parietal cortex, is a better predictor of information maintained in visual working memory and working memory performance; dynamic patterns of activity may convey information without requiring persistent activity at all; and prefrontal neurons may be ill-suited to represent non-spatial information about the features and identity of remembered stimuli. Alternative interpretations about the role of the prefrontal cortex have thus been suggested, such as that it provides a top-down control of information represented in other brain areas, rather than maintaining a working memory trace itself. Here we review evidence for and against the role of prefrontal persistent activity, with a focus on visual neurophysiology. We show that persistent activity predicts behavioral parameters precisely in working memory tasks. We illustrate that prefrontal cortex represents features of stimuli other than their spatial location, and that this information is largely absent from early cortical areas during working memory. We examine memory models not dependent on persistent activity, and conclude that each of those models could mediate only a limited range of memory-dependent behaviors. We review activity decoded from brain areas other than the prefrontal cortex during working memory and demonstrate that these areas alone cannot mediate working memory maintenance, particularly in the presence of distractors. We finally discuss the discrepancy between

  5. Ralstonia solanacearum Type III Effector RipAY Is a Glutathione-Degrading Enzyme That Is Activated by Plant Cytosolic Thioredoxins and Suppresses Plant Immunity

    PubMed Central

    Hatanaka, Tadashi; Nakano, Masahito; Oda, Kenji

    2016-01-01

    ABSTRACT The plant pathogen Ralstonia solanacearum uses a large repertoire of type III effector proteins to succeed in infection. To clarify the function of effector proteins in host eukaryote cells, we expressed effectors in yeast cells and identified seven effector proteins that interfere with yeast growth. One of the effector proteins, RipAY, was found to share homology with the ChaC family proteins that function as γ-glutamyl cyclotransferases, which degrade glutathione (GSH), a tripeptide that plays important roles in the plant immune system. RipAY significantly inhibited yeast growth and simultaneously induced rapid GSH depletion when expressed in yeast cells. The in vitro GSH degradation activity of RipAY is specifically activated by eukaryotic factors in the yeast and plant extracts. Biochemical purification of the yeast protein identified that RipAY is activated by thioredoxin TRX2. On the other hand, RipAY was not activated by bacterial thioredoxins. Interestingly, RipAY was activated by plant h-type thioredoxins that exist in large amounts in the plant cytosol, but not by chloroplastic m-, f-, x-, y- and z-type thioredoxins, in a thiol-independent manner. The transient expression of RipAY decreased the GSH level in plant cells and affected the flg22-triggered production of reactive oxygen species (ROS) and expression of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) marker genes in Nicotiana benthamiana leaves. These results indicate that RipAY is activated by host cytosolic thioredoxins and degrades GSH specifically in plant cells to suppress plant immunity. PMID:27073091

  6. The Glucose Transporter Glut1 is Selectively Essential for CD4 T Cell Activation and Effector Function

    PubMed Central

    Nichols, Amanda G.; Michalek, Ryan D.; Rudolph, Michael C.; Deoliveira, Divino; Anderson, Steven M.; Abel, E. Dale; Chen, Benny J.; Hale, Laura P.; Rathmell, Jeffrey C.

    2014-01-01

    SUMMARY CD4 T cell activation leads to rapid proliferation and differentiation into effector (Teff) or regulatory (Treg) cells that mediate or control immunity. While Teff and Treg prefer distinct glycolytic or oxidative metabolic programs in vitro, requirements and mechanisms that control T cell glucose uptake and metabolism in vivo are poorly understood. Despite expression of multiple glucose transporters, Glut1-deficiency selectively impaired metabolism and function of thymocytes and Teff. Resting T cells were normal until activated, when Glut1-deficiency prevented increased glucose uptake and glycolysis, growth, proliferation, and decreased cell survival and Teff differentiation. Importantly, Glut1-deficiency decreased Teff expansion and ability to induce inflammatory disease in vivo. Treg, in contrast, were enriched in vivo and appeared functionally unaffected by Glut1-deficiency and able to suppress Teff irrespective of Glut1 expression. These data show a selective in vivo requirement for Glut1 in metabolic reprogramming of CD4 T cell activation and Teff expansion and survival. PMID:24930970

  7. CTLA4 blockade expands FoxP3+ regulatory and activated effector CD4+ T cells in a dose-dependent fashion

    PubMed Central

    Kavanagh, Brian; O'Brien, Shaun; Lee, David; Hou, Yafei; Weinberg, Vivian; Rini, Brian; Allison, James P.; Small, Eric J.

    2008-01-01

    Cytotoxic T lymphocyte–associated antigen 4 (CTLA4) delivers inhibitory signals to activated T cells. CTLA4 is constitutively expressed on regulatory CD4+ T cells (Tregs), but its role in these cells remains unclear. CTLA4 blockade has been shown to induce antitumor immunity. In this study, we examined the effects of anti-CTLA4 antibody on the endogenous CD4+ T cells in cancer patients. We show that CTLA4 blockade induces an increase not only in the number of activated effector CD4+ T cells, but also in the number of CD4+ FoxP3+ Tregs. Although the effects were dose-dependent, CD4+ FoxP3+ regulatory T cells could be expanded at lower antibody doses. In contrast, expansion of effector T cells was seen only at the highest dose level studied. Moreover, these expanded CD4+ FoxP3+ regulatory T cells are induced to proliferate with treatment and possess suppressor function. Our results demonstrate that treatment with anti-CTLA4 antibody does not deplete human CD4+ FoxP3+ Tregs in vivo, but rather may mediate its effects through the activation of effector T cells. Our results also suggest that CTLA4 may inhibit Treg proliferation similar to its role on effector T cells. This study is registered at http://www.clinicaltrials.gov/ct2/show/NCT00064129, registry number NCT00064129. PMID:18523152

  8. The Activation of Phytophthora Effector Avr3b by Plant Cyclophilin is Required for the Nudix Hydrolase Activity of Avr3b

    PubMed Central

    Kong, Guanghui; Zhao, Yao; Jing, Maofeng; Huang, Jie; Yang, Jin; Xia, Yeqiang; Kong, Liang; Ye, Wenwu; Xiong, Qin; Qiao, Yongli; Dong, Suomeng; Ma, Wenbo; Wang, Yuanchao

    2015-01-01

    Plant pathogens secrete an arsenal of effector proteins to impair host immunity. Some effectors possess enzymatic activities that can modify their host targets. Previously, we demonstrated that a Phytophthora sojae RXLR effector Avr3b acts as a Nudix hydrolase when expressed in planta; and this enzymatic activity is required for full virulence of P. sojae strain P6497 in soybean (Glycine max). Interestingly, recombinant Avr3b produced by E. coli does not have the hydrolase activity unless it was incubated with plant protein extracts. Here, we report the activation of Avr3b by a prolyl-peptidyl isomerase (PPIase), cyclophilin, in plant cells. Avr3b directly interacts with soybean cyclophilin GmCYP1, which activates the hydrolase activity of Avr3b in a PPIase activity-dependent manner. Avr3b contains a putative Glycine-Proline (GP) motif; which is known to confer cyclophilin-binding in other protein substrates. Substitution of the Proline (P132) in the putative GP motif impaired the interaction of Avr3b with GmCYP1; as a result, the mutant Avr3bP132A can no longer be activated by GmCYP1, and is also unable to promote Phytophthora infection. Avr3b elicits hypersensitive response (HR) in soybean cultivars producing the resistance protein Rps3b, but Avr3bP132A lost its ability to trigger HR. Furthermore, silencing of GmCYP1 rendered reduced cell death triggered by Avr3b, suggesting that GmCYP1-mediated Avr3b maturation is also required for Rps3b recognition. Finally, cyclophilins of Nicotiana benthamiana can also interact with Avr3b and activate its enzymatic activity. Overall, our results demonstrate that cyclophilin is a “helper” that activates the enzymatic activity of Avr3b after it is delivered into plant cells; as such, cyclophilin is required for the avirulence and virulence functions of Avr3b. PMID:26317500

  9. Regulation of selected genome loci using de novo-engineered transcription activator-like effector (TALE)-type transcription factors.

    PubMed

    Morbitzer, Robert; Römer, Patrick; Boch, Jens; Lahaye, Thomas

    2010-12-14

    Proteins that can be tailored to bind desired DNA sequences are key tools for molecular biology. Previous studies suggested that DNA-binding specificity of transcription activator-like effectors (TALEs) from the bacterial genus Xanthomonas is defined by repeat-variable diresidues (RVDs) of tandem-arranged 34/35-amino acid repeat units. We have studied chimeras of two TALEs differing in RVDs and non-RVDs and found that, in contrast to the critical contributions by RVDs, non-RVDs had no major effect on the DNA-binding specificity of the chimeras. This finding suggests that one needs only to modify the RVDs to generate designer TALEs (dTALEs) to activate transcription of user-defined target genes. We used the scaffold of the TALE AvrBs3 and changed its RVDs to match either the tomato Bs4, the Arabidopsis EGL3, or the Arabidopsis KNAT1 promoter. All three dTALEs transcriptionally activated the desired promoters in a sequence-specific manner as mutations within the targeted DNA sequences abolished promoter activation. This study is unique in showing that chromosomal loci can be targeted specifically by dTALEs. We also engineered two AvrBs3 derivatives with four additional repeat units activating specifically either the pepper Bs3 or UPA20 promoter. Because AvrBs3 activates both promoters, our data show that addition of repeat units facilitates TALE-specificity fine-tuning. Finally, we demonstrate that the RVD NK mediates specific interaction with G nucleotides that thus far could not be targeted specifically by any known RVD type. In summary, our data demonstrate that the TALE scaffold can be tailored to target user-defined DNA sequences in whole genomes.

  10. Efficient Gene Editing in Pluripotent Stem Cells by Bacterial Injection of Transcription Activator-Like Effector Nuclease Proteins

    PubMed Central

    Jia, Jingyue; Bai, Fang; Jin, Yongxin; Santostefano, Katherine E.; Ha, Un-Hwan; Wu, Donghai

    2015-01-01

    The type III secretion system (T3SS) of Pseudomonas aeruginosa is a powerful tool for direct protein delivery into mammalian cells and has successfully been used to deliver various exogenous proteins into mammalian cells. In the present study, transcription activator-like effector nuclease (TALEN) proteins have been efficiently delivered using the P. aeruginosa T3SS into mouse embryonic stem cells (mESCs), human ESCs (hESCs), and human induced pluripotent stem cells (hiPSCs) for genome editing. This bacterial delivery system offers an alternative method of TALEN delivery that is highly efficient in cleavage of the chromosomal target and presumably safer by avoiding plasmid DNA introduction. We combined the method of bacterial T3SS-mediated TALEN protein injection and transfection of an oligonucleotide template to effectively generate precise genetic modifications in the stem cells. Initially, we efficiently edited a single-base in the gfp gene of a mESC line to silence green fluorescent protein (GFP) production. The resulting GFP-negative mESC was cloned from a single cell and subsequently mutated back to a GFP-positive mESC line. Using the same approach, the gfp gene was also effectively knocked out in hESCs. In addition, a defined single-base edition was effectively introduced into the X-chromosome-linked HPRT1 gene in hiPSCs, generating an in vitro model of Lesch-Nyhan syndrome. T3SS-mediated TALEN protein delivery provides a highly efficient alternative for introducing precise gene editing within pluripotent stem cells for the purpose of disease genotype-phenotype relationship studies and cellular replacement therapies. Significance The present study describes a novel and powerful tool for the delivery of the genome editing enzyme transcription activator-like effector nuclease (TALEN) directly into pluripotent stem cells (PSCs), achieving desired base changes on the genomes of PSCs with high efficiency. This novel approach uses bacteria as a protein delivery

  11. Intracellular Complement Activation Sustains T Cell Homeostasis and Mediates Effector Differentiation

    PubMed Central

    Liszewski, M. Kathryn; Kolev, Martin; Le Friec, Gaelle; Leung, Marilyn; Bertram, Paula G.; Fara, Antonella F.; Subias, Marta; Pickering, Matthew C.; Drouet, Christian; Meri, Seppo; Arstila, T. Petteri; Pekkarinen, Pirkka T.; Ma, Margaret; Cope, Andrew; Reinheckel, Thomas; Rodriguez de Cordoba, Santiago; Afzali, Behdad; Atkinson, John P.; Kemper, Claudia

    2013-01-01

    Summary Complement is viewed as a critical serum-operative component of innate immunity, with processing of its key component, C3, into activation fragments C3a and C3b confined to the extracellular space. We report here that C3 activation also occurred intracellularly. We found that the T cell-expressed protease cathepsin L (CTSL) processed C3 into biologically active C3a and C3b. Resting T cells contained stores of endosomal and lysosomal C3 and CTSL and substantial amounts of CTSL-generated C3a. While “tonic” intracellular C3a generation was required for homeostatic T cell survival, shuttling of this intracellular C3-activation-system to the cell surface upon T cell stimulation induced autocrine proinflammatory cytokine production. Furthermore, T cells from patients with autoimmune arthritis demonstrated hyperactive intracellular complement activation and interferon-γ production and CTSL inhibition corrected this deregulated phenotype. Importantly, intracellular C3a was observed in all examined cell populations, suggesting that intracellular complement activation might be of broad physiological significance. PMID:24315997

  12. Intracellular complement activation sustains T cell homeostasis and mediates effector differentiation.

    PubMed

    Liszewski, M Kathryn; Kolev, Martin; Le Friec, Gaelle; Leung, Marilyn; Bertram, Paula G; Fara, Antonella F; Subias, Marta; Pickering, Matthew C; Drouet, Christian; Meri, Seppo; Arstila, T Petteri; Pekkarinen, Pirkka T; Ma, Margaret; Cope, Andrew; Reinheckel, Thomas; Rodriguez de Cordoba, Santiago; Afzali, Behdad; Atkinson, John P; Kemper, Claudia

    2013-12-12

    Complement is viewed as a critical serum-operative component of innate immunity, with processing of its key component, C3, into activation fragments C3a and C3b confined to the extracellular space. We report here that C3 activation also occurred intracellularly. We found that the T cell-expressed protease cathepsin L (CTSL) processed C3 into biologically active C3a and C3b. Resting T cells contained stores of endosomal and lysosomal C3 and CTSL and substantial amounts of CTSL-generated C3a. While "tonic" intracellular C3a generation was required for homeostatic T cell survival, shuttling of this intracellular C3-activation-system to the cell surface upon T cell stimulation induced autocrine proinflammatory cytokine production. Furthermore, T cells from patients with autoimmune arthritis demonstrated hyperactive intracellular complement activation and interferon-γ production and CTSL inhibition corrected this deregulated phenotype. Importantly, intracellular C3a was observed in all examined cell populations, suggesting that intracellular complement activation might be of broad physiological significance. PMID:24315997

  13. Active versus passive maintenance of visual nonverbal memory.

    PubMed

    McKeown, Denis; Holt, Jessica; Delvenne, Jean-Francois; Smith, Amy; Griffiths, Benjamin

    2014-08-01

    Forgetting over the short term has challenged researchers for more than a century, largely because of the difficulty of controlling what goes on within the memory retention interval. But the "recent-negative-probe" procedure offers a valuable paradigm, by examining the influences of (presumably) unattended memoranda from prior trials. Here we used a recent-probe task to investigate forgetting for visual nonverbal short-term memory. The target stimuli (two visually presented abstract shapes) on a trial were followed after a retention interval by a probe, and participants indicated whether the probe matched one of the target items. Proactive interference, and hence memory for old trial probes, was observed, whereby participants were slowed in rejecting a nonmatching probe on the current trial that nevertheless matched a target item on the previous trial (a recent-negative probe). The attraction of the paradigm is that, by uncovering proactive influences of past-trial probe stimuli, it can be argued that active maintenance in memory of those probes is unlikely. In two experiments, we recorded such proactive interference of prior-trial items over a range of interstimulus (ISI) and intertrial (ITI) intervals (between 1 and 6 s, respectively). Consistent with a proposed two-process memory conception (the active-passive memory model, or APM), actively maintained memories on current trials decayed, but passively "maintained," or unattended, visual memories of stimuli on past trials did not. PMID:24390797

  14. Human Tyr-tRNA synthetase is a potent PARP-1 activating effector target for resveratrol

    PubMed Central

    Sajish, Mathew; Schimmel, Paul

    2014-01-01

    Resveratrol (RSV) is reported to extend life span1,2 and provide cardio-neuro-protective3, anti-diabetic4, and anti-cancer effects3,5 by initiating a stress response2 that induces survival genes. Because human tyrosyl tRNA synthetase (TyrRS) translocates to the nucleus under stress conditions6, we considered the possibility that the tyrosine-like phenolic ring of RSV might fit into the active site pocket to effect a nuclear role. Here we present a 2.1Å co-crystal structure of RSV bound to the active site of TyrRS. RSV nullified the catalytic activity and redirected TyrRS to a nuclear function, stimulating NAD+-dependent auto-poly-ADP-ribosylation of PARP-1. Downstream activation of key stress signaling pathways were causally connected to TyrRS-PARP-1-NAD+ collaboration. This collaboration was also demonstrated in the mouse, and was specifically blocked in vivo by a RSV-displacing tyrosyl adenylate analog. In contrast to functionally diverse tRNA synthetase catalytic nulls created by alternative splicing events that ablate active sites7, here a non-spliced TyrRS catalytic null reveals a new PARP-1- and NAD+-dependent dimension to the physiological mechanism of RSV. PMID:25533949

  15. Death effector activation in the subventricular zone subsequent to perinatal hypoxia/ischemia.

    PubMed

    Romanko, Michael J; Zhu, Changlian; Bahr, Ben A; Blomgren, Klas; Levison, Steven W

    2007-11-01

    Perinatal hypoxia/ischemia (H/I) is the leading cause of neurological injury resulting from birth complications and pre-maturity. Our studies have demonstrated that this injury depletes the subventricular zone (SVZ) of progenitors. In this study, we sought to reveal which cell death pathways are activated within these progenitors after H/I. We found that calpain activity is detected as early as 4 h of reperfusion and is sustained for 48 h, while caspase 3 activation does not occur until 8 h and peaks at 24 h post-insult. Activated calpains and caspase 3 co-localized within precursors situated in the lateral aspects of the SVZ (which coincides with progenitor cell death), whereas neither enzyme was activated in the medial SVZ (which harbors the neural stem cells that are resilient to this insult). These studies reveal targets for neuroprotective agents to protect precursors from cell death towards the goal of restoring normal brain development after H/I.

  16. Structural Mechanism of WASP Activation by the Enterohaemorrhagic E. coli Effector EspFU

    PubMed Central

    Cheng, Hui-Chun; Skehan, Brian M.; Campellone, Kenneth G.; Leong, John M.; Rosen, Michael K.

    2008-01-01

    During infection enterohaemorrhagic Escherichia coli (EHEC) usurp the actin cytoskeleton of eukaryotic cells by injecting the EspFU protein into the host cytoplasm1, 2. EspFU controls actin by activating members of the Wiskott-Aldrich Syndrome Protein (WASP) family1–5. Here we show that EspFU binds the autoinhibitory GTPase binding domain (GBD) in WASP proteins and displaces it from the activity-bearing VCA domain. This interaction potently activates WASP and N-WASP in vitro and induces localized actin assembly in cells. In the solution structure of the GBD-EspFU complex, EspFU forms an amphipathic helix that binds the GBD, mimicking interactions of the VCA in autoinhibited WASP. Thus, EspFU activates by competing directly for the VCA binding site on the GBD. This mechanism is distinct from that used by the eukaryotic activators Cdc42 and SH2 domains, which globally destabilize the GBD fold to release the VCA6–8. Such diversity of mechanism in WASP proteins is distinct from other multi-modular systems, and may result from the intrinsically unstructured nature of the isolated GBD and VCA elements. The structural incompatibility of the GBD complexes with EspFU and Cdc42/SH2, plus high affinity EspFU binding, enable EHEC to potently hijack the eukaryotic cytoskeletal machinery. PMID:18650809

  17. Single molecule real-time sequencing of Xanthomonas oryzae genomes reveals a dynamic structure and complex TAL (transcription activator-like) effector gene relationships

    PubMed Central

    Booher, Nicholas J.; Carpenter, Sara C. D.; Sebra, Robert P.; Wang, Li; Salzberg, Steven L.; Leach, Jan E.; Bogdanove, Adam J.

    2016-01-01

    Pathogen-injected, direct transcriptional activators of host genes, TAL (transcription activator-like) effectors play determinative roles in plant diseases caused by Xanthomonas spp. A large domain of nearly identical, 33–35 aa repeats in each protein mediates DNA recognition. This modularity makes TAL effectors customizable and thus important also in biotechnology. However, the repeats render TAL effector (tal) genes nearly impossible to assemble using next-generation, short reads. Here, we demonstrate that long-read, single molecule real-time (SMRT) sequencing solves this problem. Taking an ensemble approach to first generate local, tal gene contigs, we correctly assembled de novo the genomes of two strains of the rice pathogen X. oryzae completed previously using the Sanger method and even identified errors in those references. Sequencing two more strains revealed a dynamic genome structure and a striking plasticity in tal gene content. Our results pave the way for population-level studies to inform resistance breeding, improve biotechnology and probe TAL effector evolution. PMID:27148456

  18. Jet Engine Exhaust Nozzle Flow Effector

    NASA Technical Reports Server (NTRS)

    Turner, Travis L. (Inventor); Cano, Roberto J. (Inventor); Silcox, Richard J. (Inventor); Buehrle, Ralph D. (Inventor); Cagle, Christopher M. (Inventor); Cabell, Randolph H. (Inventor); Hilton, George C. (Inventor)

    2011-01-01

    A jet engine exhaust nozzle flow effector is a chevron formed with a radius of curvature with surfaces of the flow effector being defined and opposing one another. At least one shape memory alloy (SMA) member is embedded in the chevron closer to one of the chevron's opposing surfaces and substantially spanning from at least a portion of the chevron's root to the chevron's tip.

  19. Jet Engine Exhaust Nozzle Flow Effector

    NASA Technical Reports Server (NTRS)

    Turner, Travis L. (Inventor); Cano, Roberto J. (Inventor); Silox, Richard J. (Inventor); Buehrle, Ralph D. (Inventor); Cagle, Christopher M. (Inventor); Cabell, Randolph H. (Inventor); Hilton, George C. (Inventor)

    2014-01-01

    A jet engine exhaust nozzle flow effector is a chevron formed with a radius of curvature with surfaces of the flow effector being defined and opposing one another. At least one shape memory alloy (SMA) member is embedded in the chevron closer to one of the chevron's opposing surfaces and substantially spanning from at least a portion of the chevron's root to the chevron's tip.

  20. Understanding the Mysterious M2 Macrophage through Activation Markers and Effector Mechanisms

    PubMed Central

    Rőszer, Tamás

    2015-01-01

    The alternatively activated or M2 macrophages are immune cells with high phenotypic heterogeneity and are governing functions at the interface of immunity, tissue homeostasis, metabolism, and endocrine signaling. Today the M2 macrophages are identified based on the expression pattern of a set of M2 markers. These markers are transmembrane glycoproteins, scavenger receptors, enzymes, growth factors, hormones, cytokines, and cytokine receptors with diverse and often yet unexplored functions. This review discusses whether these M2 markers can be reliably used to identify M2 macrophages and define their functional subdivisions. Also, it provides an update on the novel signals of the tissue environment and the neuroendocrine system which shape the M2 activation. The possible evolutionary roots of the M2 macrophage functions are also discussed. PMID:26089604

  1. Transcription activator-like effector nuclease (TALEN)-mediated female-specific sterility in the silkworm, Bombyx mori.

    PubMed

    Xu, J; Wang, Y; Li, Z; Ling, L; Zeng, B; James, A A; Tan, A; Huang, Y

    2014-12-01

    Engineering sex-specific sterility is critical for developing transgene-based sterile insect technology. Targeted genome engineering achieved by customized zinc-finger nuclease, transcription activator-like effector nuclease (TALEN) or clustered, regularly interspaced, short palindromic repeats/Cas9 systems has been exploited extensively in a variety of model organisms; however, screening mutated individuals without a detectable phenotype is still challenging. In addition, genetically recessive mutations only detectable in homozygotes make the experiments time-consuming. In the present study, we model a novel genetic system in the silkworm, Bombyx mori, that results in female-specific sterility by combining transgenesis with TALEN technologies. This system induces sex-specific sterility at a high efficiency by targeting the female-specific exon of the B. mori doublesex (Bmdsx) gene, which has sex-specific splicing isoforms regulating somatic sexual development. Transgenic animals co-expressing TALEN left and right arms targeting the female-specific Bmdsx exon resulted in somatic mutations and female mutants lost fecundity because of lack of egg storage and abnormal external genitalia. The wild-type sexual dimorphism of abdominal segment was not evident in mutant females. In contrast, there were no deleterious effects in mutant male moths. The current somatic TALEN technologies provide a promising approach for future insect functional genetics, thus providing the basis for the development of attractive genetic alternatives for insect population management. PMID:25125145

  2. A One-Step System for Convenient and Flexible Assembly of Transcription Activator-Like Effector Nucleases (TALENs).

    PubMed

    Zhao, Jinlong; Sun, Wenye; Liang, Jing; Jiang, Jing; Wu, Zhao

    2016-09-01

    Transcription activator-like effector nucleases (TALENs) are powerful tools for targeted genome editing in diverse cell types and organisms. However, the highly identical TALE repeat sequences make it challenging to assemble TALEs using conventional cloning approaches, and multiple repeats in one plasmid are easily catalyzed for homologous recombination in bacteria. Although the methods for TALE assembly are constantly improving, these methods are not convenient because of laborious assembly steps or large module libraries, limiting their broad utility. To overcome the barrier of multiple assembly steps, we report a one-step system for the convenient and flexible assembly of a 180 TALE module library. This study is the first demonstration to ligate 9 mono-/dimer modules and one circular TALEN backbone vector in a one step process, generating 9.5 to 18.5 repeat sequences with an overall assembly rate higher than 50%. This system makes TALEN assembly much simpler than the conventional cloning of two DNA fragments because this strategy combines digestion and ligation into one step using circular vectors and different modules to avoid gel extraction. Therefore, this system provides a convenient tool for the application of TALEN-mediated genome editing in scientific studies and clinical trials. PMID:27604899

  3. Allele-Specific Reduction of the Mutant Huntingtin Allele Using Transcription Activator-Like Effectors in Human Huntington's Disease Fibroblasts.

    PubMed

    Fink, Kyle D; Deng, Peter; Gutierrez, Josh; Anderson, Joseph S; Torrest, Audrey; Komarla, Anvita; Kalomoiris, Stefanos; Cary, Whitney; Anderson, Johnathon D; Gruenloh, William; Duffy, Alexandra; Tempkin, Teresa; Annett, Geralyn; Wheelock, Vicki; Segal, David J; Nolta, Jan A

    2016-01-01

    Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder caused by an abnormal expansion of CAG repeats. Although pathogenesis has been attributed to this polyglutamine expansion, the underlying mechanisms through which the huntingtin protein functions have yet to be elucidated. It has been suggested that postnatal reduction of mutant huntingtin through protein interference or conditional gene knockout could prove to be an effective therapy for patients suffering from HD. For allele-specific targeting, transcription activator-like effectors (TALE) were designed to target single-nucleotide polymorphisms (SNP) in the mutant allele and packaged into a vector backbone containing KRAB to promote transcriptional repression of the disease-associated allele. Additional TALEs were packaged into a vector backbone containing heterodimeric FokI and were designed to be used as nucleases (TALEN) to cause a CAG-collapse in the mutant allele. Human HD fibroblasts were treated with each TALE-SNP or TALEN. Allele-expression was measured using a SNP-genotyping assay and mutant protein aggregation was quantified with Western blots for anti-ubiquitin. The TALE-SNP and TALEN significantly reduced mutant allele expression (p < 0.05) when compared to control transfections while not affecting expression of the nondisease allele. This study demonstrates the potential of allele-specific gene modification using TALE proteins, and provides a foundation for targeted treatment for individuals suffering from Huntington's or other genetically linked diseases. PMID:26850319

  4. A One-Step System for Convenient and Flexible Assembly of Transcription Activator-Like Effector Nucleases (TALENs)

    PubMed Central

    Zhao, Jinlong; Sun, Wenye; Liang, Jing; Jiang, Jing; Wu, Zhao

    2016-01-01

    Transcription activator-like effector nucleases (TALENs) are powerful tools for targeted genome editing in diverse cell types and organisms. However, the highly identical TALE repeat sequences make it challenging to assemble TALEs using conventional cloning approaches, and multiple repeats in one plasmid are easily catalyzed for homologous recombination in bacteria. Although the methods for TALE assembly are constantly improving, these methods are not convenient because of laborious assembly steps or large module libraries, limiting their broad utility. To overcome the barrier of multiple assembly steps, we report a one-step system for the convenient and flexible assembly of a 180 TALE module library. This study is the first demonstration to ligate 9 mono-/dimer modules and one circular TALEN backbone vector in a one step process, generating 9.5 to 18.5 repeat sequences with an overall assembly rate higher than 50%. This system makes TALEN assembly much simpler than the conventional cloning of two DNA fragments because this strategy combines digestion and ligation into one step using circular vectors and different modules to avoid gel extraction. Therefore, this system provides a convenient tool for the application of TALEN-mediated genome editing in scientific studies and clinical trials. PMID:27604899

  5. Signal transducer and activator of transcription 5 (STAT5) paralog dose governs T cell effector and regulatory functions

    PubMed Central

    Villarino, Alejandro; Laurence, Arian; Robinson, Gertraud W; Bonelli, Michael; Dema, Barbara; Afzali, Behdad; Shih, Han-Yu; Sun, Hong-Wei; Brooks, Stephen R; Hennighausen, Lothar; Kanno, Yuka; O'Shea, John J

    2016-01-01

    The transcription factor STAT5 is fundamental to the mammalian immune system. However, the relationship between its two paralogs, STAT5A and STAT5B, and the extent to which they are functionally distinct, remain uncertain. Using mouse models of paralog deficiency, we demonstrate that they are not equivalent for CD4+ 'helper' T cells, the principal orchestrators of adaptive immunity. Instead, we find that STAT5B is dominant for both effector and regulatory (Treg) responses and, therefore, uniquely necessary for immunological tolerance. Comparative analysis of genomic distribution and transcriptomic output confirm that STAT5B has fargreater impact but, surprisingly, the data point towards asymmetric expression (i.e. paralog dose), rather than distinct functional properties, as the key distinguishing feature. Thus, we propose a quantitative model of STAT5 paralog activity whereby relative abundance imposes functional specificity (or dominance) in the face of widespread structural homology. DOI: http://dx.doi.org/10.7554/eLife.08384.001 PMID:26999798

  6. Salicylic acid receptors activate jasmonic acid signalling through a non-canonical pathway to promote effector-triggered immunity

    PubMed Central

    Liu, Lijing; Sonbol, Fathi-Mohamed; Huot, Bethany; Gu, Yangnan; Withers, John; Mwimba, Musoki; Yao, Jian; He, Sheng Yang; Dong, Xinnian

    2016-01-01

    It is an apparent conundrum how plants evolved effector-triggered immunity (ETI), involving programmed cell death (PCD), as a major defence mechanism against biotrophic pathogens, because ETI-associated PCD could leave them vulnerable to necrotrophic pathogens that thrive on dead host cells. Interestingly, during ETI, the normally antagonistic defence hormones, salicylic acid (SA) and jasmonic acid (JA) associated with defence against biotrophs and necrotrophs respectively, both accumulate to high levels. In this study, we made the surprising finding that JA is a positive regulator of RPS2-mediated ETI. Early induction of JA-responsive genes and de novo JA synthesis following SA accumulation is activated through the SA receptors NPR3 and NPR4, instead of the JA receptor COI1. We provide evidence that NPR3 and NPR4 may mediate this effect by promoting degradation of the JA transcriptional repressor JAZs. This unique interplay between SA and JA offers a possible explanation of how plants can mount defence against a biotrophic pathogen without becoming vulnerable to necrotrophic pathogens. PMID:27725643

  7. Transcription activator-like effector nucleases mediated metabolic engineering for enhanced fatty acids production in Saccharomyces cerevisiae.

    PubMed

    Aouida, Mustapha; Li, Lixin; Mahjoub, Ali; Alshareef, Sahar; Ali, Zahir; Piatek, Agnieszka; Mahfouz, Magdy M

    2015-10-01

    Targeted engineering of microbial genomes holds much promise for diverse biotechnological applications. Transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/Cas9 systems are capable of efficiently editing microbial genomes, including that of Saccharomyces cerevisiae. Here, we demonstrate the use of TALENs to edit the genome of S. cerevisiae with the aim of inducing the overproduction of fatty acids. Heterodimeric TALENs were designed to simultaneously edit the FAA1 and FAA4 genes encoding acyl-CoA synthetases in S. cerevisiae. Functional yeast double knockouts generated using these TALENs over-produce large amounts of free fatty acids into the cell. This study demonstrates the use of TALENs for targeted engineering of yeast and demonstrates that this technology can be used to stimulate the enhanced production of free fatty acids, which are potential substrates for biofuel production. This proof-of-principle study extends the utility of TALENs as excellent genome editing tools and highlights their potential use for metabolic engineering of yeast and other organisms, such as microalgae and plants, for biofuel production.

  8. Working memory delay activity predicts individual differences in cognitive abilities.

    PubMed

    Unsworth, Nash; Fukuda, Keisuke; Awh, Edward; Vogel, Edward K

    2015-05-01

    A great deal of prior research has examined the relation between estimates of working memory and cognitive abilities. Yet, the neural mechanisms that account for these relations are still not very well understood. The current study explored whether individual differences in working memory delay activity would be a significant predictor of cognitive abilities. A large number of participants performed multiple measures of capacity, attention control, long-term memory, working memory span, and fluid intelligence, and latent variable analyses were used to examine the data. During two working memory change detection tasks, we acquired EEG data and examined the contralateral delay activity. The results demonstrated that the contralateral delay activity was significantly related to cognitive abilities, and importantly these relations were because of individual differences in both capacity and attention control. These results suggest that individual differences in working memory delay activity predict individual differences in a broad range of cognitive abilities, and this is because of both differences in the number of items that can be maintained and the ability to control access to working memory. PMID:25436671

  9. Working Memory Delay Activity Predicts Individual Differences in Cognitive Abilities

    PubMed Central

    Unsworth, Nash; Fukuda, Keisuke; Awh, Edward; Vogel, Edward K.

    2015-01-01

    A great deal of prior research has examined the relation between estimates of working memory and cognitive abilities. Yet, the neural mechanisms that account for these relations are still not very well understood. The current study explored whether individual differences in working memory delay activity would be a significant predictor of cognitive abilities. A large number of participants performed multiple measures of capacity, attention control, long-term memory, working memory span, and fluid intelligence, and latent variable analyses were used to examine the data. During two working memory change detection tasks, we acquired EEG data and examined the contra-lateral delay activity. The results demonstrated that the contralateral delay activity was significantly related to cognitive abilities, and importantly these relations were because of individual differences in both capacity and attention control. These results suggest that individual differences in working memory delay activity predict individual differences in a broad range of cognitive abilities, and this is because of both differences in the number of items that can be maintained and the ability to control access to working memory. PMID:25436671

  10. Human temporal cortical single neuron activity during working memory maintenance.

    PubMed

    Zamora, Leona; Corina, David; Ojemann, George

    2016-06-01

    The Working Memory model of human memory, first introduced by Baddeley and Hitch (1974), has been one of the most influential psychological constructs in cognitive psychology and human neuroscience. However the neuronal correlates of core components of this model have yet to be fully elucidated. Here we present data from two studies where human temporal cortical single neuron activity was recorded during tasks differentially affecting the maintenance component of verbal working memory. In Study One we vary the presence or absence of distracting items for the entire period of memory storage. In Study Two we vary the duration of storage so that distractors filled all, or only one-third of the time the memory was stored. Extracellular single neuron recordings were obtained from 36 subjects undergoing awake temporal lobe resections for epilepsy, 25 in Study one, 11 in Study two. Recordings were obtained from a total of 166 lateral temporal cortex neurons during performance of one of these two tasks, 86 study one, 80 study two. Significant changes in activity with distractor manipulation were present in 74 of these neurons (45%), 38 Study one, 36 Study two. In 48 (65%) of those there was increased activity during the period when distracting items were absent, 26 Study One, 22 Study Two. The magnitude of this increase was greater for Study One, 47.6%, than Study Two, 8.1%, paralleling the reduction in memory errors in the absence of distracters, for Study One of 70.3%, Study Two 26.3% These findings establish that human lateral temporal cortex is part of the neural system for working memory, with activity during maintenance of that memory that parallels performance, suggesting it represents active rehearsal. In 31 of these neurons (65%) this activity was an extension of that during working memory encoding that differed significantly from the neural processes recorded during overt and silent language tasks without a recent memory component, 17 Study one, 14 Study two

  11. Human temporal cortical single neuron activity during working memory maintenance.

    PubMed

    Zamora, Leona; Corina, David; Ojemann, George

    2016-06-01

    The Working Memory model of human memory, first introduced by Baddeley and Hitch (1974), has been one of the most influential psychological constructs in cognitive psychology and human neuroscience. However the neuronal correlates of core components of this model have yet to be fully elucidated. Here we present data from two studies where human temporal cortical single neuron activity was recorded during tasks differentially affecting the maintenance component of verbal working memory. In Study One we vary the presence or absence of distracting items for the entire period of memory storage. In Study Two we vary the duration of storage so that distractors filled all, or only one-third of the time the memory was stored. Extracellular single neuron recordings were obtained from 36 subjects undergoing awake temporal lobe resections for epilepsy, 25 in Study one, 11 in Study two. Recordings were obtained from a total of 166 lateral temporal cortex neurons during performance of one of these two tasks, 86 study one, 80 study two. Significant changes in activity with distractor manipulation were present in 74 of these neurons (45%), 38 Study one, 36 Study two. In 48 (65%) of those there was increased activity during the period when distracting items were absent, 26 Study One, 22 Study Two. The magnitude of this increase was greater for Study One, 47.6%, than Study Two, 8.1%, paralleling the reduction in memory errors in the absence of distracters, for Study One of 70.3%, Study Two 26.3% These findings establish that human lateral temporal cortex is part of the neural system for working memory, with activity during maintenance of that memory that parallels performance, suggesting it represents active rehearsal. In 31 of these neurons (65%) this activity was an extension of that during working memory encoding that differed significantly from the neural processes recorded during overt and silent language tasks without a recent memory component, 17 Study one, 14 Study two

  12. Neutrophil activation induced by ArtinM: release of inflammatory mediators and enhancement of effector functions.

    PubMed

    Toledo, Karina Alves; Scwartz, Carolina; Oliveira, Aline Ferreira; Conrado, Marina Cavalcanti Albuquerque Veiga; Bernardes, Emerson Soares; Fernandes, Luiz Cláudio; Roque-Barreira, Maria Cristina; Pereira-da-Silva, Gabriela; Moreno, Andréa Novais

    2009-03-24

    The D-mannose binding lectin ArtinM from Artocarpus integrifolia, previously known as KM+ and artocarpin, is considered a stimulant of Th1-type immunity, which is able to confer resistance to some intracellular pathogens. In addition, ArtinM induces neutrophil migration by haptotaxis through simultaneous interactions of its carbohydrate recognition domains (CRDs) with glycans expressed on the extracellular matrix and the neutrophil surface. In the present study, we have expanded the characterization of ArtinM as a neutrophil activator. Exposure of neutrophils to ArtinM for 15 min resulted in tyrosine phosphorylation of intracellular proteins, a process that was selectively inhibited by d-mannose or mannotriose. Shortly after stimulation, neutrophils secreted high levels of LTB(4) and underwent shedding of L-selectin from their surface. Exposure to ArtinM enhanced neutrophil functions, such as respiratory burst and zymozan and Listeria monocytogenes phagocytosis. In addition, ArtinM-stimulated neutrophils displayed increased CXCL-8 secretion and TLR2 gene transcription. These results demonstrate that ArtinM is able to induce potent neutrophil activation, a feature that should be strongly considered in the assessment of the lectin capacity to confer resistance against infections.

  13. FusX: A Rapid One-Step Transcription Activator-Like Effector Assembly System for Genome Science.

    PubMed

    Ma, Alvin C; McNulty, Melissa S; Poshusta, Tanya L; Campbell, Jarryd M; Martínez-Gálvez, Gabriel; Argue, David P; Lee, Han B; Urban, Mark D; Bullard, Cassandra E; Blackburn, Patrick R; Man, Toni K; Clark, Karl J; Ekker, Stephen C

    2016-06-01

    Transcription activator-like effectors (TALEs) are extremely effective, single-molecule DNA-targeting molecular cursors used for locus-specific genome science applications, including high-precision molecular medicine and other genome engineering applications. TALEs are used in genome engineering for locus-specific DNA editing and imaging, as artificial transcriptional activators and repressors, and for targeted epigenetic modification. TALEs as nucleases (TALENs) are effective editing tools and offer high binding specificity and fewer sequence constraints toward the targeted genome than other custom nuclease systems. One bottleneck of broader TALE use is reagent accessibility. For example, one commonly deployed method uses a multitube, 5-day assembly protocol. Here we describe FusX, a streamlined Golden Gate TALE assembly system that (1) is backward compatible with popular TALE backbones, (2) is functionalized as a single-tube 3-day TALE assembly process, (3) requires only commonly used basic molecular biology reagents, and (4) is cost-effective. More than 100 TALEN pairs have been successfully assembled using FusX, and 27 pairs were quantitatively tested in zebrafish, with each showing high somatic and germline activity. Furthermore, this assembly system is flexible and is compatible with standard molecular biology laboratory tools, but can be scaled with automated laboratory support. To demonstrate, we use a highly accessible and commercially available liquid-handling robot to rapidly and accurately assemble TALEs using the FusX TALE toolkit. Together, the FusX system accelerates TALE-based genomic science applications from basic science screening work for functional genomics testing and molecular medicine applications. PMID:26854857

  14. Ras regulates SCF(β-TrCP) protein activity and specificity via its effector protein NORE1A.

    PubMed

    Schmidt, M Lee; Donninger, Howard; Clark, Geoffrey J

    2014-11-01

    Ras is the most frequently activated oncogene found in human cancer, but its mechanisms of action remain only partially understood. Ras activates multiple signaling pathways to promote transformation. However, Ras can also exhibit a potent ability to induce growth arrest and death. NORE1A (RASSF5) is a direct Ras effector that acts as a tumor suppressor by promoting apoptosis and cell cycle arrest. Expression of NORE1A is frequently lost in human tumors, and its mechanism of action remains unclear. Here we show that NORE1A forms a direct, Ras-regulated complex with β-TrCP, the substrate recognition component of the SCF(β-TrCP) ubiquitin ligase complex. This interaction allows Ras to stimulate the ubiquitin ligase activity of SCF(β-TrCP) toward its target β-catenin, resulting in degradation of β-catenin by the 26 S proteasome. However, the action of Ras/NORE1A/β-TrCP is substrate-specific because IκB, another substrate of SCF(β-TrCP), is not sensitive to NORE1A-promoted degradation. We identify a completely new signaling mechanism for Ras that allows for the specific regulation of SCF(β-TrCP) targets. We show that the NORE1A levels in a cell may dictate the effects of Ras on the Wnt/β-catenin pathway. Moreover, because NORE1A expression is frequently impaired in tumors, we provide an explanation for the observation that β-TrCP can act as a tumor suppressor or an oncogene in different cell systems.

  15. FusX: A Rapid One-Step Transcription Activator-Like Effector Assembly System for Genome Science

    PubMed Central

    Ma, Alvin C.; McNulty, Melissa S.; Poshusta, Tanya L.; Campbell, Jarryd M.; Martínez-Gálvez, Gabriel; Argue, David P.; Lee, Han B.; Urban, Mark D.; Bullard, Cassandra E.; Blackburn, Patrick R.; Man, Toni K.; Clark, Karl J.; Ekker, Stephen C.

    2016-01-01

    Transcription activator-like effectors (TALEs) are extremely effective, single-molecule DNA-targeting molecular cursors used for locus-specific genome science applications, including high-precision molecular medicine and other genome engineering applications. TALEs are used in genome engineering for locus-specific DNA editing and imaging, as artificial transcriptional activators and repressors, and for targeted epigenetic modification. TALEs as nucleases (TALENs) are effective editing tools and offer high binding specificity and fewer sequence constraints toward the targeted genome than other custom nuclease systems. One bottleneck of broader TALE use is reagent accessibility. For example, one commonly deployed method uses a multitube, 5-day assembly protocol. Here we describe FusX, a streamlined Golden Gate TALE assembly system that (1) is backward compatible with popular TALE backbones, (2) is functionalized as a single-tube 3-day TALE assembly process, (3) requires only commonly used basic molecular biology reagents, and (4) is cost-effective. More than 100 TALEN pairs have been successfully assembled using FusX, and 27 pairs were quantitatively tested in zebrafish, with each showing high somatic and germline activity. Furthermore, this assembly system is flexible and is compatible with standard molecular biology laboratory tools, but can be scaled with automated laboratory support. To demonstrate, we use a highly accessible and commercially available liquid-handling robot to rapidly and accurately assemble TALEs using the FusX TALE toolkit. Together, the FusX system accelerates TALE-based genomic science applications from basic science screening work for functional genomics testing and molecular medicine applications. PMID:26854857

  16. Long-term memories in online users' selecting activities

    NASA Astrophysics Data System (ADS)

    Pan, Xue; Hou, Lei; Stephen, Mutua; Yang, Huijie

    2014-07-01

    In this paper, we investigate the long-term memory effect in the behavior of online users. Two user-oriented online movie systems are used in this study. Due to the short length of the series, the balanced estimation of diffusion entropy approach is used to evaluate scaling-invariance in selecting activities of users in the two online movie systems. Our results indicate that persistence (long-term memory) exists widely in the movie selecting series. However, there is generally significant difference between a user's objective and subjective behaviors. Additionally, statistically, the long-term memory depends on activity levels, as results show that the much more active a users' group, the stronger the long-term memory will be. These findings provide a new criterion for constructing reasonable models, and can help understand how individuals' behaviors form a collective behavior of an online society.

  17. A survey of commercially available manipulators, end-effectors, and delivery systems for reactor decommissioning activities

    SciTech Connect

    Henley, D.R.; Litka, T.J.

    1996-05-01

    Numerous nuclear facilities owned by the U.S. Department of Energy (DOE) are under consideration for decommissioning. Currently, there are no standardized, automated, remote systems designed to dismantle and thereby reduce the size of activated reactor components and vessels so that they can be packaged and shipped to disposal sites. Existing dismantling systems usually consist of customized, facility-specific tooling that has been developed to dismantle a specific reactor system. Such systems have a number of drawbacks. Generally, current systems cannot be disassembled, moved, and reused. Developing and deploying the tooling for current systems is expensive and time-consuming. In addition, the amount of manual work is significant because long-handled tools must be used; as a result, personnel are exposed to excessive radiation. A standardized, automated, remote system is therefore needed to deliver the tooling necessary to dismantle nuclear facilities at different locations. Because this system would be reusable, it would produce less waste. The system would also save money because of its universal design, and it would be more reliable than current systems.

  18. SseK3 Is a Salmonella Effector That Binds TRIM32 and Modulates the Host’s NF-κB Signalling Activity

    PubMed Central

    Yang, Zhe; Soderholm, Amelia; Lung, Tania Wong Fok; Giogha, Cristina; Hill, Michelle M.; Brown, Nathaniel F.; Hartland, Elizabeth; Teasdale, Rohan D.

    2015-01-01

    Salmonella Typhimurium employs an array of type III secretion system effectors that facilitate intracellular survival and replication during infection. The Salmonella effector SseK3 was originally identified due to amino acid sequence similarity with NleB; an effector secreted by EPEC/EHEC that possesses N-acetylglucoasmine (GlcNAc) transferase activity and modifies death domain containing proteins to block extrinsic apoptosis. In this study, immunoprecipitation of SseK3 defined a novel molecular interaction between SseK3 and the host protein, TRIM32, an E3 ubiquitin ligase. The conserved DxD motif within SseK3, which is essential for the GlcNAc transferase activity of NleB, was required for TRIM32 binding and for the capacity of SseK3 to suppress TNF-stimulated activation of NF-κB pathway. However, we did not detect GlcNAc modification of TRIM32 by SseK3, nor did the SseK3-TRIM32 interaction impact on TRIM32 ubiquitination that is associated with its activation. In addition, lack of sseK3 in Salmonella had no effect on production of the NF-κB dependent cytokine, IL-8, in HeLa cells even though TRIM32 knockdown suppressed TNF-induced NF-κB activity. Ectopically expressed SseK3 partially co-localises with TRIM32 at the trans-Golgi network, but SseK3 is not recruited to Salmonella induced vacuoles or Salmonella induced filaments during Salmonella infection. Our study has identified a novel effector-host protein interaction and suggests that SseK3 may influence NF-κB activity. However, the lack of GlcNAc modification of TRIM32 suggests that SseK3 has further, as yet unidentified, host targets. PMID:26394407

  19. SseK3 Is a Salmonella Effector That Binds TRIM32 and Modulates the Host's NF-κB Signalling Activity.

    PubMed

    Yang, Zhe; Soderholm, Amelia; Lung, Tania Wong Fok; Giogha, Cristina; Hill, Michelle M; Brown, Nathaniel F; Hartland, Elizabeth; Teasdale, Rohan D

    2015-01-01

    Salmonella Typhimurium employs an array of type III secretion system effectors that facilitate intracellular survival and replication during infection. The Salmonella effector SseK3 was originally identified due to amino acid sequence similarity with NleB; an effector secreted by EPEC/EHEC that possesses N-acetylglucoasmine (GlcNAc) transferase activity and modifies death domain containing proteins to block extrinsic apoptosis. In this study, immunoprecipitation of SseK3 defined a novel molecular interaction between SseK3 and the host protein, TRIM32, an E3 ubiquitin ligase. The conserved DxD motif within SseK3, which is essential for the GlcNAc transferase activity of NleB, was required for TRIM32 binding and for the capacity of SseK3 to suppress TNF-stimulated activation of NF-κB pathway. However, we did not detect GlcNAc modification of TRIM32 by SseK3, nor did the SseK3-TRIM32 interaction impact on TRIM32 ubiquitination that is associated with its activation. In addition, lack of sseK3 in Salmonella had no effect on production of the NF-κB dependent cytokine, IL-8, in HeLa cells even though TRIM32 knockdown suppressed TNF-induced NF-κB activity. Ectopically expressed SseK3 partially co-localises with TRIM32 at the trans-Golgi network, but SseK3 is not recruited to Salmonella induced vacuoles or Salmonella induced filaments during Salmonella infection. Our study has identified a novel effector-host protein interaction and suggests that SseK3 may influence NF-κB activity. However, the lack of GlcNAc modification of TRIM32 suggests that SseK3 has further, as yet unidentified, host targets.

  20. AC Electric Field Activated Shape Memory Polymer Composite

    NASA Technical Reports Server (NTRS)

    Kang, Jin Ho; Siochi, Emilie J.; Penner, Ronald K.; Turner, Travis L.

    2011-01-01

    Shape memory materials have drawn interest for applications like intelligent medical devices, deployable space structures and morphing structures. Compared to other shape memory materials like shape memory alloys (SMAs) or shape memory ceramics (SMCs), shape memory polymers (SMPs) have high elastic deformation that is amenable to tailored of mechanical properties, have lower density, and are easily processed. However, SMPs have low recovery stress and long response times. A new shape memory thermosetting polymer nanocomposite (LaRC-SMPC) was synthesized with conductive fillers to enhance its thermo-mechanical characteristics. A new composition of shape memory thermosetting polymer nanocomposite (LaRC-SMPC) was synthesized with conductive functionalized graphene sheets (FGS) to enhance its thermo-mechanical characteristics. The elastic modulus of LaRC-SMPC is approximately 2.7 GPa at room temperature and 4.3 MPa above its glass transition temperature. Conductive FGSs-doped LaRC-SMPC exhibited higher conductivity compared to pristine LaRC SMP. Applying an electric field at between 0.1 Hz and 1 kHz induced faster heating to activate the LaRC-SMPC s shape memory effect relative to applying DC electric field or AC electric field at frequencies exceeding1 kHz.

  1. Activity in prelimbic cortex subserves fear memory reconsolidation over time

    PubMed Central

    Stern, Cristina A.J.; Gazarini, Lucas; Vanvossen, Ana C.; Hames, Mayara S.; Bertoglio, Leandro J.

    2014-01-01

    The prelimbic cortex has been implicated in the consolidation of previously learned fear. Herein, we report that temporarily inactivating this medial prefrontal cortex subregion with the GABAA agonist muscimol (4.0 nmol in 0.2 μL per hemisphere) was able to equally disrupt 1-, 7-, and 21-d-old contextual fear memories after their brief retrieval in rats. In all cases, this effect was prevented when memory reactivation was omitted. These results indicate that recent and remote fear memories are susceptible to reconsolidation blockade induced by prelimbic cortex inactivation. It was also demonstrated that the disrupting effect of prelimbic cortex inactivation on fear memory persisted over 11 d, and did not show extinction-related features, such as reinstatement. Infusing the same dose and volume of muscimol bilaterally into the infralimbic cortex after brief retrieval/reactivation of the fear memory did not disrupt it, as seen in prelimbic cortex-inactivated animals. The expression of Zif268/Egr1, the product of an immediate early gene related to memory reconsolidation, was also less pronounced in the infralimbic cortex than in prelimbic cortex following memory retrieval/reactivation. Altogether, the present findings highlight that activity in the prelimbic cortex may reestablish reactivated aversive memories and, therefore, contribute to their maintenance over time. PMID:24344180

  2. Activating human genes with zinc finger proteins, transcription activator-like effectors and CRISPR/Cas9 for gene therapy and regenerative medicine.

    PubMed

    Gersbach, Charles A; Perez-Pinera, Pablo

    2014-08-01

    New technologies have recently been developed to control the expression of human genes in their native genomic context by engineering synthetic transcription factors that can be targeted to any DNA sequence. The ability to precisely regulate any gene as it occurs naturally in the genome provides a means to address a variety of diseases and disorders. This approach also circumvents some of the traditional challenges of gene therapy. In this editorial, we review the technologies that have enabled targeted human gene activation, including the engineering of transcription factors based on zinc finger proteins, transcription activator-like effectors and the CRISPR/Cas9 system. Additionally, we highlight examples in which these methods have been developed for therapeutic applications and discuss challenges and opportunities.

  3. A library of synthetic transcription activator-like effector-activated promoters for coordinated orthogonal gene expression in plants

    PubMed Central

    Brückner, Kathleen; Schäfer, Petra; Weber, Ernst; Grützner, Ramona; Marillonnet, Sylvestre; Tissier, Alain

    2015-01-01

    A library of synthetic promoters containing the binding site of a single designer transcription activator-like effector (dTALE) was constructed. The promoters contain a constant sequence, consisting of an 18-base long dTALE-binding site and a TATA box, flanked by degenerate sequences of 49 bases downstream and 19 bases upstream. Forty-three of these promoters were sequenced and tested in transient assays in Nicotiana benthamiana using a GUS reporter gene. The strength of expression of the promoters ranged from around 5% to almost 100% of the viral 35S promoter activity. We then demonstrated the utility of these promoters for metabolic engineering by transiently expressing three genes for the production of a plant diterpenoid in N. benthamiana. The simplicity of the promoter structure shows great promise for the development of genetic circuits, with wide potential applications in plant synthetic biology and metabolic engineering. PMID:25846505

  4. The Yersinia Virulence Factor YopM Hijacks Host Kinases to Inhibit Type III Effector-Triggered Activation of the Pyrin Inflammasome.

    PubMed

    Chung, Lawton K; Park, Yong Hwan; Zheng, Yueting; Brodsky, Igor E; Hearing, Patrick; Kastner, Daniel L; Chae, Jae Jin; Bliska, James B

    2016-09-14

    Pathogenic Yersinia, including Y. pestis, the agent of plague in humans, and Y. pseudotuberculosis, the related enteric pathogen, deliver virulence effectors into host cells via a prototypical type III secretion system to promote pathogenesis. These effectors, termed Yersinia outer proteins (Yops), modulate multiple host signaling responses. Studies in Y. pestis and Y. pseudotuberculosis have shown that YopM suppresses infection-induced inflammasome activation; however, the underlying molecular mechanism is largely unknown. Here we show that YopM specifically restricts the pyrin inflammasome, which is triggered by the RhoA-inactivating enzymatic activities of YopE and YopT, in Y. pseudotuberculosis-infected macrophages. The attenuation of a yopM mutant is fully reversed in pyrin knockout mice, demonstrating that YopM inhibits pyrin to promote virulence. Mechanistically, YopM recruits and activates the host kinases PRK1 and PRK2 to negatively regulate pyrin by phosphorylation. These results show how a virulence factor can hijack host kinases to inhibit effector-triggered pyrin inflammasome activation. PMID:27569559

  5. Repeated nitrous oxide exposure in rats causes a thermoregulatory sign-reversal with concurrent activation of opposing thermoregulatory effectors

    PubMed Central

    Ramsay, Douglas S.; Woods, Stephen C.; Kaiyala, Karl J.

    2015-01-01

    Initial administration of 60% nitrous oxide (N2O) to rats at an ambient temperature of 21°C decreases core temperature (Tc), primarily via increased heat loss (HL). Over repeated N2O administrations, rats first develop tolerance to this hypothermia and subsequently exhibit hyperthermia (a sign-reversal) due primarily to progressive increases in heat production (HP). When rats initially receive 60% N2O in a thermal gradient, they become hypothermic while selecting cooler ambient temperatures that facilitate HL. This study investigated whether rats repeatedly administered 60% N2O in a thermal gradient would use the gradient to behaviorally facilitate, or oppose, the development of chronic tolerance and a hyperthermic sign-reversal. Male Long-Evans rats (N=16) received twelve 3-h administrations of 60% N2O in a gas-tight, live-in thermal gradient. Hypothermia (Sessions 1-3), complete chronic tolerance (Sessions 4-6), and a subsequent transient hyperthermic sign-reversal (Sessions 7-12) sequentially developed. Despite the progressive recovery and eventual hyperthermic sign-reversal of Tc, rats consistently selected cooler ambient temperatures during all N2O administrations. A final 60% N2O administration in a total calorimeter indicated that the hyperthermic sign-reversal resulted primarily from increased HP. Thus, rats did not facilitate chronic tolerance development by moving to warmer locations in the gradient, and instead selected cooler ambient temperatures while simultaneously increasing autonomic HP. The inefficient concurrent activation of opposing effectors and the development of a sign-reversal are incompatible with homeostatic models of drug-adaptation and may be better interpreted using a model of drug-induced allostasis. PMID:25938127

  6. The role of REM sleep theta activity in emotional memory.

    PubMed

    Hutchison, Isabel C; Rathore, Shailendra

    2015-01-01

    While non-REM (NREM) sleep has been strongly implicated in the reactivation and consolidation of memory traces, the role of rapid-eye movement (REM) sleep remains unclear. A growing body of research on humans and animals provide behavioral evidence for a role of REM sleep in the strengthening and modulation of emotional memories. Theta activity-which describes low frequency oscillations in the local field potential within the hippocampus, amygdala and neocortex-is a prominent feature of both wake and REM sleep in humans and rodents. Theta coherence between the hippocampus and amygdala drives large-scale pontine-geniculo-occipital (PGO) waves, the density of which predicts increases in plasticity-related gene expression. This could potentially facilitate the processing of emotional memory traces within the hippocampus during REM sleep. Further, the timing of hippocampal activity in relation to theta phase is vital in determining subsequent potentiation of neuronal activity. This could allow the emotionally modulated strengthening of novel and gradual weakening of consolidated hippocampal memory traces during REM sleep. Hippocampal theta activity is also correlated with REM sleep levels of achetylcholine - which is thought to reduce hippocampal inputs in the neocortex. The additional low levels of noradrenaline during REM sleep, which facilitate feedback within the neocortex, could allow the integration of novel memory traces previously consolidated during NREM sleep. We therefore propose that REM sleep mediates the prioritized processing of emotional memories within the hippocampus, the integration of previously consolidated memory traces within the neocortex, as well as the disengagement of consolidated neocortical memory traces from the hippocampus. PMID:26483709

  7. The role of REM sleep theta activity in emotional memory.

    PubMed

    Hutchison, Isabel C; Rathore, Shailendra

    2015-01-01

    While non-REM (NREM) sleep has been strongly implicated in the reactivation and consolidation of memory traces, the role of rapid-eye movement (REM) sleep remains unclear. A growing body of research on humans and animals provide behavioral evidence for a role of REM sleep in the strengthening and modulation of emotional memories. Theta activity-which describes low frequency oscillations in the local field potential within the hippocampus, amygdala and neocortex-is a prominent feature of both wake and REM sleep in humans and rodents. Theta coherence between the hippocampus and amygdala drives large-scale pontine-geniculo-occipital (PGO) waves, the density of which predicts increases in plasticity-related gene expression. This could potentially facilitate the processing of emotional memory traces within the hippocampus during REM sleep. Further, the timing of hippocampal activity in relation to theta phase is vital in determining subsequent potentiation of neuronal activity. This could allow the emotionally modulated strengthening of novel and gradual weakening of consolidated hippocampal memory traces during REM sleep. Hippocampal theta activity is also correlated with REM sleep levels of achetylcholine - which is thought to reduce hippocampal inputs in the neocortex. The additional low levels of noradrenaline during REM sleep, which facilitate feedback within the neocortex, could allow the integration of novel memory traces previously consolidated during NREM sleep. We therefore propose that REM sleep mediates the prioritized processing of emotional memories within the hippocampus, the integration of previously consolidated memory traces within the neocortex, as well as the disengagement of consolidated neocortical memory traces from the hippocampus.

  8. The role of REM sleep theta activity in emotional memory

    PubMed Central

    Hutchison, Isabel C.; Rathore, Shailendra

    2015-01-01

    While non-REM (NREM) sleep has been strongly implicated in the reactivation and consolidation of memory traces, the role of rapid-eye movement (REM) sleep remains unclear. A growing body of research on humans and animals provide behavioral evidence for a role of REM sleep in the strengthening and modulation of emotional memories. Theta activity—which describes low frequency oscillations in the local field potential within the hippocampus, amygdala and neocortex—is a prominent feature of both wake and REM sleep in humans and rodents. Theta coherence between the hippocampus and amygdala drives large-scale pontine-geniculo-occipital (PGO) waves, the density of which predicts increases in plasticity-related gene expression. This could potentially facilitate the processing of emotional memory traces within the hippocampus during REM sleep. Further, the timing of hippocampal activity in relation to theta phase is vital in determining subsequent potentiation of neuronal activity. This could allow the emotionally modulated strengthening of novel and gradual weakening of consolidated hippocampal memory traces during REM sleep. Hippocampal theta activity is also correlated with REM sleep levels of achetylcholine - which is thought to reduce hippocampal inputs in the neocortex. The additional low levels of noradrenaline during REM sleep, which facilitate feedback within the neocortex, could allow the integration of novel memory traces previously consolidated during NREM sleep. We therefore propose that REM sleep mediates the prioritized processing of emotional memories within the hippocampus, the integration of previously consolidated memory traces within the neocortex, as well as the disengagement of consolidated neocortical memory traces from the hippocampus. PMID:26483709

  9. Marker for type VI secretion system effectors

    PubMed Central

    Salomon, Dor; Kinch, Lisa N.; Trudgian, David C.; Guo, Xiaofeng; Klimko, John A.; Grishin, Nick V.; Mirzaei, Hamid; Orth, Kim

    2014-01-01

    Bacteria use diverse mechanisms to kill, manipulate, and compete with other cells. The recently discovered type VI secretion system (T6SS) is widespread in bacterial pathogens and used to deliver virulence effector proteins into target cells. Using comparative proteomics, we identified two previously unidentified T6SS effectors that contained a conserved motif. Bioinformatic analyses revealed that this N-terminal motif, named MIX (marker for type six effectors), is found in numerous polymorphic bacterial proteins that are primarily located in the T6SS genome neighborhood. We demonstrate that several MIX-containing proteins are T6SS effectors and that they are not required for T6SS activity. Thus, we propose that MIX-containing proteins are T6SS effectors. Our findings allow for the identification of numerous uncharacterized T6SS effectors that will undoubtedly lead to the discovery of new biological mechanisms. PMID:24927539

  10. Making memories without trying: medial temporal lobe activity associated with incidental memory formation during recognition.

    PubMed

    Stark, Craig E L; Okado, Yoko

    2003-07-30

    Structures in the medial portions of the human temporal lobes (MTL) play a vital role in the ability to learn new facts and events, whether such learning is intentional or incidental. We examined neural activity in the MTL both while participants studied pictures of novel scenes and while they attempted to recognize which scenes had been previously presented. In a second surprise test we assessed participants' memory for items that were presented only during the previous recognition memory test. We present a novel approach to cross-participant alignment of neuroimaging data that provides more precise localization and enhanced statistical power within regions such as the MTL. Using this technique, we observed that the amount of MTL activity predicted participants' ability to subsequently remember scenes not only during the intentional study task, but also during the first memory retrieval test when only incidental encoding occurred. This encoding-related activity during memory retrieval was in the same subregions of the MTL as encoding-related activity during intentional study and is hypothesized to be one of the primary reasons why retrieval-related activity is often difficult to observe with neuroimaging techniques. PMID:12890767

  11. Memory

    MedlinePlus

    ... it has to decide what is worth remembering. Memory is the process of storing and then remembering this information. There are different types of memory. Short-term memory stores information for a few ...

  12. Pre-incubation of human monocytes results in loss of effector activity and diminished stimulation of the autologous mixed lymphocyte reaction.

    PubMed Central

    Lederman, M M; Liebman, M L; Hassid, A I; Berk, G I

    1983-01-01

    Human monocytes were cultured at 37 degrees C for 72 h, washed, adjusted for viability and compared to freshly prepared monocytes for stimulation of the autologous mixed lymphocyte reaction (AMLR) and effector function. Pre-incubated monocytes were less potent AMLR stimulators than were freshly prepared cells. Pre-incubated monocytes demonstrated less antibody-dependent tumour killing of CCRF-CEM, less killing of Staphylococci and less spontaneous tumour killing of K-562 than did fresh monocytes. Pre-incubated monocytes produced less prostaglandin E2, demonstrated less surface Ia antigen and were less efficient accessory cells for antigen presentation than were fresh monocytes. AMLR stimulation correlated with monocyte killing (r = 0.95) and PGE2 production (r = 0.98). Thus, monocytes pre-incubated for 3 days are less active effector cells, display less surface Ia antigen and are less potent stimulators of the AMLR than fresh monocytes. Moreover, in this system, monocyte effector activity correlates with ability to stimulate the AMLR. PMID:6224613

  13. A Yersinia effector with enhanced inhibitory activity on the NF-κB pathway activates the NLRP3/ASC/caspase-1 inflammasome in macrophages.

    PubMed

    Zheng, Ying; Lilo, Sarit; Brodsky, Igor E; Zhang, Yue; Medzhitov, Ruslan; Marcu, Kenneth B; Bliska, James B

    2011-04-01

    A type III secretion system (T3SS) in pathogenic Yersinia species functions to translocate Yop effectors, which modulate cytokine production and regulate cell death in macrophages. Distinct pathways of T3SS-dependent cell death and caspase-1 activation occur in Yersinia-infected macrophages. One pathway of cell death and caspase-1 activation in macrophages requires the effector YopJ. YopJ is an acetyltransferase that inactivates MAPK kinases and IKKβ to cause TLR4-dependent apoptosis in naïve macrophages. A YopJ isoform in Y. pestis KIM (YopJ(KIM)) has two amino acid substitutions, F177L and K206E, not present in YopJ proteins of Y. pseudotuberculosis and Y. pestis CO92. As compared to other YopJ isoforms, YopJ(KIM) causes increased apoptosis, caspase-1 activation, and secretion of IL-1β in Yersinia-infected macrophages. The molecular basis for increased apoptosis and activation of caspase-1 by YopJ(KIM) in Yersinia-infected macrophages was studied. Site directed mutagenesis showed that the F177L and K206E substitutions in YopJ(KIM) were important for enhanced apoptosis, caspase-1 activation, and IL-1β secretion. As compared to YopJ(CO92), YopJ(KIM) displayed an enhanced capacity to inhibit phosphorylation of IκB-α in macrophages and to bind IKKβ in vitro. YopJ(KIM) also showed a moderately increased ability to inhibit phosphorylation of MAPKs. Increased caspase-1 cleavage and IL-1β secretion occurred in IKKβ-deficient macrophages infected with Y. pestis expressing YopJ(CO92), confirming that the NF-κB pathway can negatively regulate inflammasome activation. K+ efflux, NLRP3 and ASC were important for secretion of IL-1β in response to Y. pestis KIM infection as shown using macrophages lacking inflammasome components or by the addition of exogenous KCl. These data show that caspase-1 is activated in naïve macrophages in response to infection with a pathogen that inhibits IKKβ and MAPK kinases and induces TLR4-dependent apoptosis. This pro

  14. A Yersinia Effector with Enhanced Inhibitory Activity on the NF-κB Pathway Activates the NLRP3/ASC/Caspase-1 Inflammasome in Macrophages

    PubMed Central

    Brodsky, Igor E.; Zhang, Yue; Medzhitov, Ruslan; Marcu, Kenneth B.; Bliska, James B.

    2011-01-01

    A type III secretion system (T3SS) in pathogenic Yersinia species functions to translocate Yop effectors, which modulate cytokine production and regulate cell death in macrophages. Distinct pathways of T3SS-dependent cell death and caspase-1 activation occur in Yersinia-infected macrophages. One pathway of cell death and caspase-1 activation in macrophages requires the effector YopJ. YopJ is an acetyltransferase that inactivates MAPK kinases and IKKβ to cause TLR4-dependent apoptosis in naïve macrophages. A YopJ isoform in Y. pestis KIM (YopJKIM) has two amino acid substitutions, F177L and K206E, not present in YopJ proteins of Y. pseudotuberculosis and Y. pestis CO92. As compared to other YopJ isoforms, YopJKIM causes increased apoptosis, caspase-1 activation, and secretion of IL-1β in Yersinia-infected macrophages. The molecular basis for increased apoptosis and activation of caspase-1 by YopJKIM in Yersinia-infected macrophages was studied. Site directed mutagenesis showed that the F177L and K206E substitutions in YopJKIM were important for enhanced apoptosis, caspase-1 activation, and IL-1β secretion. As compared to YopJCO92, YopJKIM displayed an enhanced capacity to inhibit phosphorylation of IκB-α in macrophages and to bind IKKβ in vitro. YopJKIM also showed a moderately increased ability to inhibit phosphorylation of MAPKs. Increased caspase-1 cleavage and IL-1β secretion occurred in IKKβ-deficient macrophages infected with Y. pestis expressing YopJCO92, confirming that the NF-κB pathway can negatively regulate inflammasome activation. K+ efflux, NLRP3 and ASC were important for secretion of IL-1β in response to Y. pestis KIM infection as shown using macrophages lacking inflammasome components or by the addition of exogenous KCl. These data show that caspase-1 is activated in naïve macrophages in response to infection with a pathogen that inhibits IKKβ and MAPK kinases and induces TLR4-dependent apoptosis. This pro-inflammatory form of apoptosis

  15. Communication: Memory effects and active Brownian diffusion

    SciTech Connect

    Ghosh, Pulak K.; Li, Yunyun; Marchegiani, Giampiero; Marchesoni, Fabio

    2015-12-07

    A self-propelled artificial microswimmer is often modeled as a ballistic Brownian particle moving with constant speed aligned along one of its axis, but changing direction due to random collisions with the environment. Similarly to thermal noise, its angular randomization is described as a memoryless stochastic process. Here, we speculate that finite-time correlations in the orientational dynamics can affect the swimmer’s diffusivity. To this purpose, we propose and solve two alternative models. In the first one, we simply assume that the environmental fluctuations governing the swimmer’s propulsion are exponentially correlated in time, whereas in the second one, we account for possible damped fluctuations of the propulsion velocity around the swimmer’s axis. The corresponding swimmer’s diffusion constants are predicted to get, respectively, enhanced or suppressed upon increasing the model memory time. Possible consequences of this effect on the interpretation of the experimental data are discussed.

  16. Genetic diversity of transcriptional activator-like effector genes in Chinese isolates of Xanthomonas oryzae pv. oryzicola.

    PubMed

    Ji, Zhi-Yuan; Zakria, Muhammad; Zou, Li-Fang; Xiong, Li; Li, Zheng; Ji, Guang-Hai; Chen, Gong-You

    2014-07-01

    Xanthomonas oryzae pv. oryzicola causes bacterial leaf streak (BLS), a devastating disease of rice in Asia countries. X. oryzae pv. oryzicola utilizes repertoires of transcriptional activator-like effectors (TALEs) to manipulate host resistance or susceptibility; thus, TALEs can determine the outcome of BLS. In this report, we studied genetic diversity in putative tale genes of 65 X. oryzae pv. oryzicola strains that originated from nine provinces of southern China. Genomic DNAs from the 65 strains were digested with BamHI and hybridized with an internal fragment of avrXa3, a tale gene originating from the related pathogen, X. oryzae pv. oryzae, which causes bacterial leaf blight (BLB). Southern blot analysis indicated that the strains contained a variable number (9 to 22) of avrXa3-hybridizing fragments (e.g., putative tale genes). Based on the number and size of hybridizing bands, strains were classified into 14 genotypes (designated 1 to 14), and genotypes 3 and 10 represented 29.23 and 24.64% of the total, respectively. A high molecular weight BamHI fragment (HMWB; ≈6.0 kb) was present in 12 of the 14 genotypes, and sequence analysis of the HMWB revealed the presence of a C-terminally truncated tale, an insertion element related to IS1403, and genes encoding phosphoglycerate mutase and endonuclease V. Primers were developed from the 6.0-kb HMWB fragment and showed potential in genotyping X. oryzae pv. oryzicola strains by polymerase chain reaction. Virulence of X. oryzae pv. oryzicola strains was assessed on 23 rice cultivars containing different resistance genes for BLB. The X. oryzae pv. oryzicola strains could be grouped into 14 pathotypes (I to XIV), and the grouping of strains was almost identical to the categories determined by genotypic analysis. In general, strains containing higher numbers of putative tale genes were more virulent on rice than strains containing fewer tales. The results also indicate that there are no gene-for-gene relationships

  17. Xanthomonas axonopodis virulence is promoted by a transcription activator-like effector-mediated induction of a SWEET sugar transporter in cassava.

    PubMed

    Cohn, Megan; Bart, Rebecca S; Shybut, Mikel; Dahlbeck, Douglas; Gomez, Michael; Morbitzer, Robert; Hou, Bi-Huei; Frommer, Wolf B; Lahaye, Thomas; Staskawicz, Brian J

    2014-11-01

    The gene-for-gene concept has historically been applied to describe a specific resistance interaction wherein single genes from the host and the pathogen dictate the outcome. These interactions have been observed across the plant kingdom and all known plant microbial pathogens. In recent years, this concept has been extended to susceptibility phenotypes in the context of transcription activator-like (TAL) effectors that target SWEET sugar transporters. However, because this interaction has only been observed in rice, it was not clear whether the gene-for-gene susceptibility was unique to that system. Here, we show, through a combined systematic analysis of the TAL effector complement of Xanthomonas axonopodis pv. manihotis and RNA sequencing to identify targets in cassava, that TAL20Xam668 specifically induces the sugar transporter MeSWEET10a to promote virulence. Designer TAL effectors (dTALE) complement TAL20Xam668 mutant phenotypes, demonstrating that MeSWEET10a is a susceptibility gene in cassava. Sucrose uptake-deficient X. axonopodis pv. manihotis bacteria do not lose virulence, indicating that sucrose may be cleaved extracellularly and taken up as hexoses into X. axonopodis pv. manihotis. Together, our data suggest that pathogen hijacking of plant nutrients is not unique to rice blight but also plays a role in bacterial blight of the dicot cassava.

  18. Xanthomonas axonopodis virulence is promoted by a transcription activator-like effector-mediated induction of a SWEET sugar transporter in cassava.

    PubMed

    Cohn, Megan; Bart, Rebecca S; Shybut, Mikel; Dahlbeck, Douglas; Gomez, Michael; Morbitzer, Robert; Hou, Bi-Huei; Frommer, Wolf B; Lahaye, Thomas; Staskawicz, Brian J

    2014-11-01

    The gene-for-gene concept has historically been applied to describe a specific resistance interaction wherein single genes from the host and the pathogen dictate the outcome. These interactions have been observed across the plant kingdom and all known plant microbial pathogens. In recent years, this concept has been extended to susceptibility phenotypes in the context of transcription activator-like (TAL) effectors that target SWEET sugar transporters. However, because this interaction has only been observed in rice, it was not clear whether the gene-for-gene susceptibility was unique to that system. Here, we show, through a combined systematic analysis of the TAL effector complement of Xanthomonas axonopodis pv. manihotis and RNA sequencing to identify targets in cassava, that TAL20Xam668 specifically induces the sugar transporter MeSWEET10a to promote virulence. Designer TAL effectors (dTALE) complement TAL20Xam668 mutant phenotypes, demonstrating that MeSWEET10a is a susceptibility gene in cassava. Sucrose uptake-deficient X. axonopodis pv. manihotis bacteria do not lose virulence, indicating that sucrose may be cleaved extracellularly and taken up as hexoses into X. axonopodis pv. manihotis. Together, our data suggest that pathogen hijacking of plant nutrients is not unique to rice blight but also plays a role in bacterial blight of the dicot cassava. PMID:25083909

  19. Autonomic activity during sleep predicts memory consolidation in humans.

    PubMed

    Whitehurst, Lauren N; Cellini, Nicola; McDevitt, Elizabeth A; Duggan, Katherine A; Mednick, Sara C

    2016-06-28

    Throughout history, psychologists and philosophers have proposed that good sleep benefits memory, yet current studies focusing on the relationship between traditionally reported sleep features (e.g., minutes in sleep stages) and changes in memory performance show contradictory findings. This discrepancy suggests that there are events occurring during sleep that have not yet been considered. The autonomic nervous system (ANS) shows strong variation across sleep stages. Also, increases in ANS activity during waking, as measured by heart rate variability (HRV), have been correlated with memory improvement. However, the role of ANS in sleep-dependent memory consolidation has never been examined. Here, we examined whether changes in cardiac ANS activity (HRV) during a daytime nap were related to performance on two memory conditions (Primed and Repeated) and a nonmemory control condition on the Remote Associates Test. In line with prior studies, we found sleep-dependent improvement in the Primed condition compared with the Quiet Wake control condition. Using regression analyses, we compared the proportion of variance in performance associated with traditionally reported sleep features (model 1) vs. sleep features and HRV during sleep (model 2). For both the Primed and Repeated conditions, model 2 (sleep + HRV) predicted performance significantly better (73% and 58% of variance explained, respectively) compared with model 1 (sleep only, 46% and 26% of variance explained, respectively). These findings present the first evidence, to our knowledge, that ANS activity may be one potential mechanism driving sleep-dependent plasticity. PMID:27298366

  20. Memory.

    ERIC Educational Resources Information Center

    McKean, Kevin

    1983-01-01

    Discusses current research (including that involving amnesiacs and snails) into the nature of the memory process, differentiating between and providing examples of "fact" memory and "skill" memory. Suggests that three brain parts (thalamus, fornix, mammilary body) are involved in the memory process. (JN)

  1. Modeling Active Aging and Explicit Memory: An Empirical Study.

    PubMed

    Ponce de León, Laura Ponce; Lévy, Jean Pierre; Fernández, Tomás; Ballesteros, Soledad

    2015-08-01

    The rapid growth of the population of older adults and their concomitant psychological status and health needs have captured the attention of researchers and health professionals. To help fill the void of literature available to social workers interested in mental health promotion and aging, the authors provide a model for active aging that uses psychosocial variables. Structural equation modeling was used to examine the relationships among the latent variables of the state of explicit memory, the perception of social resources, depression, and the perception of quality of life in a sample of 184 older adults. The results suggest that explicit memory is not a direct indicator of the perception of quality of life, but it could be considered an indirect indicator as it is positively correlated with perception of social resources and negatively correlated with depression. These last two variables influenced the perception of quality of life directly, the former positively and the latter negatively. The main outcome suggests that the perception of social support improves explicit memory and quality of life and reduces depression in active older adults. The findings also suggest that gerontological professionals should design memory training programs, improve available social resources, and offer environments with opportunities to exercise memory.

  2. The type 3 effector NopL of Sinorhizobium sp. strain NGR234 is a mitogen-activated protein kinase substrate.

    PubMed

    Ge, Ying-Ying; Xiang, Qi-Wang; Wagner, Christian; Zhang, Di; Xie, Zhi-Ping; Staehelin, Christian

    2016-04-01

    Pathogenic bacteria utilize type 3 secretion systems to inject type 3 effectors (T3Es) into host cells, thereby subverting host defense reactions. Similarly, T3Es of symbiotic nitrogen-fixing rhizobia can affect nodule formation on roots of legumes. Previous work showed that NopL (nodulation outer protein L) of Sinorhizobium(Ensifer) sp. strain NGR234 is multiply phosphorylated in eukaryotic cells and that this T3E suppresses responses mediated by mitogen-activated protein (MAP) kinase signaling in yeast (mating pheromone signaling) and plant cells (expression of pathogenesis-related defense proteins). Here, we show that NopL is a MAP kinase substrate. Microscopic observations of fluorescent fusion proteins and bimolecular fluorescence complementation analysis in onion cells indicated that NopL is targeted to the nucleus and forms a complex with SIPK (salicylic acid-induced protein kinase), a MAP kinase of tobacco. In vitro experiments demonstrated that NopL is phosphorylatyed by SIPK. At least nine distinct spots were observed after two-dimensional gel electrophoresis, indicating that NopL can be hyperphosphorylated by MAP kinases. Senescence symptoms in nodules of beans (Phaseolus vulgaris cv. Tendergreen) were analyzed to determine the symbiotic effector activity of different NopL variants with serine to alanine substitutions at identified and predicted phosphorylation sites (serine-proline motif). NopL variants with six or eight serine to alanine substitutions were partially active, whereas NopL forms with 10 or 12 substituted serine residues were inactive. In conclusion, our findings provide evidence that NopL interacts with MAP kinases and reveals the importance of serine-proline motifs for effector activity during symbiosis. PMID:26931172

  3. Structural analysis of effector functions related motifs, complement activation and hemagglutinating activities in Lama glama heavy chain antibodies.

    PubMed

    Saccodossi, Natalia; De Simone, Emilio A; Leoni, Juliana

    2012-01-15

    Heavy chain antibodies (HCAbs), devoid of the light chains and the CH(1) domain, are present in the serum of camelids. IgG(2) and IgG(3) are HCAbs; whereas IgG(1) has the conventional structure. In order to study the immunological properties of llama HCAbs, from which to date little is known, llamas (Lama glama) HCAbs cDNA were cloned, sequenced and compared with other mammalian Igs. The sequence analysis showed that llama HCAbs cDNA organization is similar to other mammalian Igs and the presence of conserved binding motifs to Protein A, Protein G, FcγRI, FcγRIII and C1q in HCAbs were observed. In a previous work, different IgG isotypes purified by Protein A and Protein G chromatography, were assayed for their ability to fix complement. Both IgG(1) and the total serum were able to fix complement, whereas IgG(2) and IgG(3) fixed complement even in the absence of antigen (anti-complementary activity). Therefore, in this work we performed the complement activating activity of the different IgG isotypes purified under physiological conditions using Sephadex G-150 and their ability to induce hemagglutination. Llamas were immunized with sheep red blood cells (RBC) stroma and the different isotypes were purified from sera. Whole serum and IgG(1) could activate complement; however, HCAbs (IgG(2)+IgG(3)) could not, despite the presence of the C1q binding motif in their primary sequence. Unlike IgG(1), the fraction corresponding to IgG(2)+IgG(3) did not display hemagglutinating activity. Our findings suggest that HCAbs cannot crosslink efficiently with different antigens and that the C1q binding site might be hindered by the proximity of the variable domains. PMID:22197565

  4. Working Memory Training: Improving Intelligence--Changing Brain Activity

    ERIC Educational Resources Information Center

    Jausovec, Norbert; Jausovec, Ksenija

    2012-01-01

    The main objectives of the study were: to investigate whether training on working memory (WM) could improve fluid intelligence, and to investigate the effects WM training had on neuroelectric (electroencephalography--EEG) and hemodynamic (near-infrared spectroscopy--NIRS) patterns of brain activity. In a parallel group experimental design,…

  5. Primer: making sense of T-cell memory.

    PubMed

    Beverley, Peter C L

    2008-01-01

    Protective memory is a key property of the immune system. Pathogen-associated molecular patterns of invading organisms deliver signals to pattern-recognition receptors that activate the innate immune system. Ligation of the T-cell receptor by peptides bound to MHC antigens and presented by dendritic cells, together with signals produced by the activated innate immune system, initiate T-cell responses. The nature of the T-cell response, consisting of phases of clonal expansion and contraction, and differentiation to effector and memory cells, however, is determined both by the properties of the antigen and the co-stimuli produced by the innate immune system. Short-lived effector and longer-lived memory T cells are generated during primary responses; after the death of most of the effectors, memory cells remain. Memory cells are heterogeneous in phenotype and function; subsets include the relatively quiescent central and more activated effector memory cells, as well as cells able to promote inflammation, help antibody production or regulate other immune responses. Understanding the properties of memory cells will help in the rational design of vaccines for 'difficult' organisms or cancer, as well as immunotherapies for autoimmune diseases. PMID:18172448

  6. Effector proteins of rust fungi.

    PubMed

    Petre, Benjamin; Joly, David L; Duplessis, Sébastien

    2014-01-01

    Rust fungi include many species that are devastating crop pathogens. To develop resistant plants, a better understanding of rust virulence factors, or effector proteins, is needed. Thus far, only six rust effector proteins have been described: AvrP123, AvrP4, AvrL567, AvrM, RTP1, and PGTAUSPE-10-1. Although some are well established model proteins used to investigate mechanisms of immune receptor activation (avirulence activities) or entry into plant cells, how they work inside host tissues to promote fungal growth remains unknown. The genome sequences of four rust fungi (two Melampsoraceae and two Pucciniaceae) have been analyzed so far. Genome-wide analyses of these species, as well as transcriptomics performed on a broader range of rust fungi, revealed hundreds of small secreted proteins considered as rust candidate secreted effector proteins (CSEPs). The rust community now needs high-throughput approaches (effectoromics) to accelerate effector discovery/characterization and to better understand how they function in planta. However, this task is challenging due to the non-amenability of rust pathosystems (obligate biotrophs infecting crop plants) to traditional molecular genetic approaches mainly due to difficulties in culturing these species in vitro. The use of heterologous approaches should be promoted in the future.

  7. The generation effect: activating broad neural circuits during memory encoding.

    PubMed

    Rosner, Zachary A; Elman, Jeremy A; Shimamura, Arthur P

    2013-01-01

    The generation effect is a robust memory phenomenon in which actively producing material during encoding acts to improve later memory performance. In a functional magnetic resonance imaging (fMRI) analysis, we explored the neural basis of this effect. During encoding, participants generated synonyms from word-fragment cues (e.g., GARBAGE-W_ST_) or read other synonym pairs (e.g., GARBAGE-WASTE). Compared to simply reading target words, generating target words significantly improved later recognition memory performance. During encoding, this benefit was associated with a broad neural network that involved both prefrontal (inferior frontal gyrus, middle frontal gyrus) and posterior cortex (inferior temporal gyrus, lateral occipital cortex, parahippocampal gyrus, ventral posterior parietal cortex). These findings define the prefrontal-posterior cortical dynamics associated with the mnemonic benefits underlying the generation effect.

  8. Thermally activated retainer means utilizing shape memory alloy

    NASA Technical Reports Server (NTRS)

    Grimaldi, Margaret E. (Inventor); Hartz, Leslie S. (Inventor)

    1993-01-01

    A retainer member suitable for retaining a gap filler placed in gaps between adjacent tile members is presented. One edge of the retainer member may be attached to the gap filler and another edge may be provided with a plurality of tab members which in an intermediate position do not interfere with placement or removal of the gap filler between tile members. The retainer member may be fabricated from a shape memory alloy which when heated to a specified memory temperature will thermally activate the tab members to predetermined memory positions engaging the tile members to retain the gap filler in the gap. This invention has particular application to the thermal tiles on space vehicles such as the Space Shuttle Orbiter.

  9. Working memory training: improving intelligence--changing brain activity.

    PubMed

    Jaušovec, Norbert; Jaušovec, Ksenija

    2012-07-01

    The main objectives of the study were: to investigate whether training on working memory (WM) could improve fluid intelligence, and to investigate the effects WM training had on neuroelectric (electroencephalography - EEG) and hemodynamic (near-infrared spectroscopy - NIRS) patterns of brain activity. In a parallel group experimental design, respondents of the working memory group after 30 h of training significantly increased performance on all tests of fluid intelligence. By contrast, respondents of the active control group (participating in a 30-h communication training course) showed no improvements in performance. The influence of WM training on patterns of neuroelectric brain activity was most pronounced in the theta and alpha bands. Theta and lower-1 alpha band synchronization was accompanied by increased lower-2 and upper alpha desynchronization. The hemodynamic patterns of brain activity after the training changed from higher right hemispheric activation to a balanced activity of both frontal areas. The neuroelectric as well as hemodynamic patterns of brain activity suggest that the training influenced WM maintenance functions as well as processes directed by the central executive. The changes in upper alpha band desynchronization could further indicate that processes related to long term memory were also influenced.

  10. Postnatal TLR2 activation impairs learning and memory in adulthood.

    PubMed

    Madar, Ravit; Rotter, Aviva; Waldman Ben-Asher, Hiba; Mughal, Mohamed R; Arumugam, Thiruma V; Wood, W H; Becker, K G; Mattson, Mark P; Okun, Eitan

    2015-08-01

    Neuroinflammation in the central nervous system is detrimental for learning and memory, as evident form epidemiological studies linking developmental defects and maternal exposure to harmful pathogens. Postnatal infections can also induce neuroinflammatory responses with long-term consequences. These inflammatory responses can lead to motor deficits and/or behavioral disabilities. Toll like receptors (TLRs) are a family of innate immune receptors best known as sensors of microbial-associated molecular patterns, and are the first responders to infection. TLR2 forms heterodimers with either TLR1 or TLR6, is activated in response to gram-positive bacterial infections, and is expressed in the brain during embryonic development. We hypothesized that early postnatal TLR2-mediated neuroinflammation would adversely affect cognitive behavior in the adult. Our data indicate that postnatal TLR2 activation affects learning and memory in adult mice in a heterodimer-dependent manner. TLR2/6 activation improved motor function and fear learning, while TLR2/1 activation impaired spatial learning and enhanced fear learning. Moreover, developmental TLR2 deficiency significantly impairs spatial learning and enhances fear learning, stressing the involvement of the TLR2 pathway in learning and memory. Analysis of the transcriptional effects of TLR2 activation reveals both common and unique transcriptional programs following heterodimer-specific TLR2 activation. These results imply that adult cognitive behavior could be influenced in part, by activation or alterations in the TLR2 pathway at birth. PMID:26021559

  11. Postnatal TLR2 activation impairs learning and memory in adulthood

    PubMed Central

    Madar, Ravit; Rotter, Aviva; Ben-Asher, Hiba Waldman; Mughal, Mohamed R.; Arumugam, Thiruma V.; Wood, WH; Becker, KG; Mattson, Mark P.; Okun, Eitan

    2015-01-01

    Neuroinflammation in the central nervous system is detrimental for learning and memory, as evident form epidemiological studies linking developmental defects and maternal exposure to harmful pathogens. Postnatal infections can also induce neuroinflammatory responses with long-term consequences. These inflammatory responses can lead to motor deficits and/or behavioral disabilities. Toll like receptors (TLRs) are a family of innate immune receptors best known as sensors of microbial-associated molecular patterns, and are the first responders to infection. TLR2 forms heterodimers with either TLR1 or TLR6, is activated in response to gram-positive bacterial infections, and is expressed in the brain during embryonic development. We hypothesized that early postnatal TLR2-mediated neuroinflammation would adversely affect cognitive behavior in the adult. Our data indicate that postnatal TLR2 activation affects learning and memory in adult mice in a heterodimer-dependent manner. TLR2/6 activation improved motor function and fear learning, while TLR2/1 activation impaired spatial learning and enhanced fear learning. Moreover, developmental TLR2 deficiency significantly impairs spatial learning and enhances fear learning, stressing the involvement of the TLR2 pathway in learning and memory. Analysis of the transcriptional effects of TLR2 activation reveals both common and unique transcriptional programs following heterodimer-specific TLR2 activation. These results imply that adult cognitive behavior could be influenced in part, by activation or alterations in the TLR2 pathway at birth. PMID:26021559

  12. Postnatal TLR2 activation impairs learning and memory in adulthood.

    PubMed

    Madar, Ravit; Rotter, Aviva; Waldman Ben-Asher, Hiba; Mughal, Mohamed R; Arumugam, Thiruma V; Wood, W H; Becker, K G; Mattson, Mark P; Okun, Eitan

    2015-08-01

    Neuroinflammation in the central nervous system is detrimental for learning and memory, as evident form epidemiological studies linking developmental defects and maternal exposure to harmful pathogens. Postnatal infections can also induce neuroinflammatory responses with long-term consequences. These inflammatory responses can lead to motor deficits and/or behavioral disabilities. Toll like receptors (TLRs) are a family of innate immune receptors best known as sensors of microbial-associated molecular patterns, and are the first responders to infection. TLR2 forms heterodimers with either TLR1 or TLR6, is activated in response to gram-positive bacterial infections, and is expressed in the brain during embryonic development. We hypothesized that early postnatal TLR2-mediated neuroinflammation would adversely affect cognitive behavior in the adult. Our data indicate that postnatal TLR2 activation affects learning and memory in adult mice in a heterodimer-dependent manner. TLR2/6 activation improved motor function and fear learning, while TLR2/1 activation impaired spatial learning and enhanced fear learning. Moreover, developmental TLR2 deficiency significantly impairs spatial learning and enhances fear learning, stressing the involvement of the TLR2 pathway in learning and memory. Analysis of the transcriptional effects of TLR2 activation reveals both common and unique transcriptional programs following heterodimer-specific TLR2 activation. These results imply that adult cognitive behavior could be influenced in part, by activation or alterations in the TLR2 pathway at birth.

  13. Gene targeting technologies in rats: zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats.

    PubMed

    Mashimo, Tomoji

    2014-01-01

    The laboratory rat has been widely used as an animal model in biomedical science for more than 150 years. Applying zinc-finger nucleases or transcription activator-like effector nucleases to rat embryos via microinjection is an efficient genome editing tool for generating targeted knockout rats. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated endonucleases have been used as an effective tool for precise and multiplex genome editing in mice and rats. In this review, the advantages and disadvantages of these site-specific nuclease technologies for genetic analysis and manipulation in rats are discussed.

  14. Melatonin controls experimental autoimmune encephalomyelitis by altering the T effector/regulatory balance.

    PubMed

    Álvarez-Sánchez, Nuria; Cruz-Chamorro, Ivan; López-González, Antonio; Utrilla, José C; Fernández-Santos, José M; Martínez-López, Alicia; Lardone, Patricia J; Guerrero, Juan M; Carrillo-Vico, Antonio

    2015-11-01

    Experimental autoimmune encephalomyelitis (EAE), the experimental model for multiple sclerosis (MS), is triggered by myelin-specific Th1 and Th17 cells. The immunomodulatory activities of melatonin have been shown to be beneficial under several conditions in which the immune system is exacerbated. Here, we sought to elucidate the basis of the melatonin protective effect on EAE by characterizing the T effector/regulatory responses, particularly those of the memory cell subsets. Melatonin was tested for its effect on Th1, Th17 and T regulatory (Treg) cells in the lymph nodes and CNS of immunodominant peptide of myelin oligodendrocyte glycoprotein (pMOG)-immunized and EAE mice, respectively. The capacity of melatonin to ameliorate EAE as well as modifying both T cell response and effector/regulatory balance was surveyed. T cell memory subsets and CD44, a key activation marker involved in the EAE pathogenesis, were also examined. Melatonin protected from EAE by decreasing peripheral and central Th1/Th17 responses and enhancing both the Treg frequency and IL-10 synthesis in the CNS. Melatonin reduced the T effector memory population and its pro-inflammatory response and regulated CD44 expression, which was decreased in T effector cells and increased in Tregs. The alterations in the T cell subpopulations were associated with a reduced mononuclear infiltration (CD4 and CD11b cells) of the melatonin-treated mice CNS. For the first time, we report that melatonin protects against EAE by controlling peripheral and central T effector/regulatory responses, effects that might be partially mediated by CD44. This immunomodulatory effect on EAE suggests that melatonin may represent an effective treatment option for MS.

  15. The relationship between memory complaints, activity and perceived health status.

    PubMed

    Lee, P-L

    2014-04-01

    Subjective memory complaints (SMC) is a possible symptom of mild cognitive impairment which may progress to dementia. The present study examines the relationship of physical activity (PA), cognitive activity (CA), social activity (SA), and perceived health status (HS) with SMC for middle age and older adults. Participants were from the MIDUS II study (Midlife in the United States) recruited in 2004-2006 (Mean age = 55.99; N = 3030). Hierarchical multiple regression was performed with SMC as the dependent variable, along with PA, CA, SA, and HS as the independent variables. The study revealed that SMC was strongly related to PA, CA, and HS, while controlling covariates. Further, HS had the strongest link with SMC among these predictors while interaction effects (PA × HS, CA × HS, and SA × HS) were insignificant. In addition, different results were achieved in younger versus older groups. Participants with more CA, PA and perception of better health had lower frequency of memory complaints. PMID:24646046

  16. The Type IV Secretion System Effector Protein CirA Stimulates the GTPase Activity of RhoA and Is Required for Virulence in a Mouse Model of Coxiella burnetii Infection.

    PubMed

    Weber, Mary M; Faris, Robert; van Schaik, Erin J; McLachlan, Juanita Thrasher; Wright, William U; Tellez, Andres; Roman, Victor A; Rowin, Kristina; Case, Elizabeth Di Russo; Luo, Zhao-Qing; Samuel, James E

    2016-09-01

    Coxiella burnetii, the etiological agent of Q fever in humans, is an intracellular pathogen that replicates in an acidified parasitophorous vacuole derived from host lysosomes. Generation of this replicative compartment requires effectors delivered into the host cell by the Dot/Icm type IVb secretion system. Several effectors crucial for C. burnetii intracellular replication have been identified, but the host pathways coopted by these essential effectors are poorly defined, and very little is known about how spacious vacuoles are formed and maintained. Here we demonstrate that the essential type IVb effector, CirA, stimulates GTPase activity of RhoA. Overexpression of CirA in mammalian cells results in cell rounding and stress fiber disruption, a phenotype that is rescued by overexpression of wild-type or constitutively active RhoA. Unlike other effector proteins that subvert Rho GTPases to modulate uptake, CirA is the first effector identified that is dispensable for uptake and instead recruits Rho GTPase to promote biogenesis of the bacterial vacuole. Collectively our results highlight the importance of CirA in coopting host Rho GTPases for establishment of Coxiella burnetii infection and virulence in mammalian cell culture and mouse models of infection. PMID:27324482

  17. Shigella Effector OspB Activates mTORC1 in a Manner That Depends on IQGAP1 and Promotes Cell Proliferation

    PubMed Central

    Eshleman, Heather D.; Fu, Yang; Bloom, Alexander; Li, Zhigang; Sacks, David B.; Goldberg, Marcia B.

    2015-01-01

    The intracellular bacterial pathogen Shigella infects and spreads through the human intestinal epithelium. Effector proteins delivered by Shigella into cells promote infection by modulating diverse host functions. We demonstrate that the effector protein OspB interacts directly with the scaffolding protein IQGAP1, and that the absence of either OspB or IQGAP1 during infection leads to larger areas of S. flexneri spread through cell monolayers. We show that the effect on the area of bacterial spread is due to OspB triggering increased cell proliferation at the periphery of infected foci, thereby replacing some of the cells that die within infected foci and restricting the area of bacterial spread. We demonstrate that OspB enhancement of cell proliferation results from activation of mTORC1, a master regulator of cell growth, and is blocked by the mTORC1-specific inhibitor rapamycin. OspB activation of mTORC1, and its effects on cell proliferation and bacterial spread, depends on IQGAP1. Our results identify OspB as a regulator of mTORC1 and mTORC1-dependent cell proliferation early during S. flexneri infection and establish a role for IQGAP1 in mTORC1 signaling. They also raise the possibility that IQGAP1 serves as a scaffold for the assembly of an OspB-mTORC1 signaling complex. PMID:26473364

  18. Disrupting frontal eye-field activity impairs memory recall.

    PubMed

    Wantz, Andrea L; Martarelli, Corinna S; Cazzoli, Dario; Kalla, Roger; Müri, René; Mast, Fred W

    2016-04-13

    A large body of research demonstrated that participants preferably look back to the encoding location when retrieving visual information from memory. However, the role of this 'looking back to nothing' is still debated. The goal of the present study was to extend this line of research by examining whether an important area in the cortical representation of the oculomotor system, the frontal eye field (FEF), is involved in memory retrieval. To interfere with the activity of the FEF, we used inhibitory continuous theta burst stimulation (cTBS). Before stimulation was applied, participants encoded a complex scene and performed a short-term (immediately after encoding) or long-term (after 24 h) recall task, just after cTBS over the right FEF or sham stimulation. cTBS did not affect overall performance, but stimulation and statement type (object vs. location) interacted. cTBS over the right FEF tended to impair object recall sensitivity, whereas there was no effect on location recall sensitivity. These findings suggest that the FEF is involved in retrieving object information from scene memory, supporting the hypothesis that the oculomotor system contributes to memory recall. PMID:26901058

  19. Memory Systems Do Not Divide on Consciousness: Reinterpreting Memory in Terms of Activation and Binding

    ERIC Educational Resources Information Center

    Reder, Lynne M.; Park, Heekyeong; Kieffaber, Paul D.

    2009-01-01

    There is a popular hypothesis that performance on implicit and explicit memory tasks reflects 2 distinct memory systems. Explicit memory is said to store those experiences that can be consciously recollected, and implicit memory is said to store experiences and affect subsequent behavior but to be unavailable to conscious awareness. Although this…

  20. End-effector microprocessor

    NASA Technical Reports Server (NTRS)

    Doggett, William R.

    1992-01-01

    The topics are presented in viewgraph form and include: automated structures assembly facility current control hierarchy; automated structures assembly facility purposed control hierarchy; end-effector software state transition diagram; block diagram for ideal install composite; and conclusions.

  1. Advanced Aerodynamic Control Effectors

    NASA Technical Reports Server (NTRS)

    Wood, Richard M.; Bauer, Steven X. S.

    1999-01-01

    A 1990 research program that focused on the development of advanced aerodynamic control effectors (AACE) for military aircraft has been reviewed and summarized. Data are presented for advanced planform, flow control, and surface contouring technologies. The data show significant increases in lift, reductions in drag, and increased control power, compared to typical aerodynamic designs. The results presented also highlighted the importance of planform selection in the design of a control effector suite. Planform data showed that dramatic increases in lift (greater than 25%) can be achieved with multiple wings and a sawtooth forebody. Passive porosity and micro drag generator control effector data showed control power levels exceeding that available from typical effectors (moving surfaces). Application of an advanced planform to a tailless concept showed benefits of similar magnitude as those observed in the generic studies.

  2. Pomegranate Juice Augments Memory and fMRI Activity in Middle-Aged and Older Adults with Mild Memory Complaints

    PubMed Central

    Bookheimer, Susan Y.; Renner, Brian A.; Ekstrom, Arne; Henning, Susanne M.; Brown, Jesse A.; Jones, Mike; Moody, Teena; Small, Gary W.

    2013-01-01

    Despite increasing emphasis on the potential of dietary antioxidants in preventing memory loss and on diet as a precursor of neurological health, rigorous studies investigating the cognitive effects of foods and their components are rare. Recent animal studies have reported memory and other cognitive benefits of polyphenols, found abundantly in pomegranate juice. We performed a preliminary, placebo-controlled randomized trial of pomegranate juice in older subjects with age-associated memory complaints using memory testing and functional brain activation (fMRI) as outcome measures. Thirty-two subjects (28 completers) were randomly assigned to drink 8 ounces of either pomegranate juice or a flavor-matched placebo drink for 4 weeks. Subjects received memory testing, fMRI scans during cognitive tasks, and blood draws for peripheral biomarkers before and after the intervention. Investigators and subjects were all blind to group membership. After 4 weeks, only the pomegranate group showed a significant improvement in the Buschke selective reminding test of verbal memory and a significant increase in plasma trolox-equivalent antioxidant capacity (TEAC) and urolithin A-glucuronide. Furthermore, compared to the placebo group, the pomegranate group had increased fMRI activity during verbal and visual memory tasks. While preliminary, these results suggest a role for pomegranate juice in augmenting memory function through task-related increases in functional brain activity. PMID:23970941

  3. Activation of the human mirror neuron system during the observation of the manipulation of virtual tools in the absence of a visible effector limb.

    PubMed

    Modroño, Cristián; Navarrete, Gorka; Rodríguez-Hernández, Antonio F; González-Mora, José L

    2013-10-25

    This work explores the mirror neuron system activity produced by the observation of virtual tool manipulations in the absence of a visible effector limb. Functional MRI data was obtained from healthy right-handed participants who manipulated a virtual paddle in the context of a digital game and watched replays of their actions. The results show how action observation produced extended bilateral activations in the parietofrontal mirror neuron system. At the same time, three regions in the left hemisphere (in the primary motor and the primary somatosensory cortex, the supplementary motor area and the dorsolateral prefrontal cortex) showed a reduced BOLD, possibly related with the prevention of inappropriate motor execution. These results can be of interest for researchers and developers working in the field of action observation neurorehabilitation.

  4. Vaccinia virus entry is followed by core activation and proteasome-mediated release of the immunomodulatory effector VH1 from lateral bodies.

    PubMed

    Schmidt, Florian Ingo; Bleck, Christopher Karl Ernst; Reh, Lucia; Novy, Karel; Wollscheid, Bernd; Helenius, Ari; Stahlberg, Henning; Mercer, Jason

    2013-08-15

    Host cell entry of vaccinia virus, the prototypic poxvirus, involves a membrane fusion event delivering the viral core and two proteinaceous lateral bodies (LBs) into the cytosol. Uncoating of viral cores is poorly characterized, and the composition and function of LBs remains enigmatic. We found that cytosolic cores rapidly dissociated from LBs and expanded in volume, which coincided with reduction of disulfide-bonded core proteins. We identified the abundant phosphoprotein F17, the dual-specificity phosphatase VH1, and the oxidoreductase G4 as bona fide LB components. After reaching the cytosol, F17 was degraded in a proteasome-dependent manner. Proteasome activity, and presumably LB disassembly, was required for the immediate immunomodulatory activity of VH1: dephosphorylation of STAT1 to prevent interferon-γ-mediated antiviral responses. These results reveal a mechanism used by poxviruses to deliver viral enzymes to the host cell cytosol and are likely to facilitate the identification of additional LB-resident viral effectors.

  5. Memory Training in the ACTIVE study: How Much is Needed and Who Benefits?

    PubMed Central

    Rebok, George W.; Langbaum, Jessica B.S.; Jones, Richard N.; Gross, Alden L.; Parisi, Jeanine M.; Spira, Adam P.; Kueider, Alexandra; Petras, Hanno; Brandt, Jason

    2013-01-01

    Objectives and Methods Data from the memory training arm (n = 629) of the Advanced Cognitive Training for Independent and Vital Elderly (ACTIVE) trial were examined to characterize change in memory performance through five years of follow-up as a function of memory training, booster training, adherence to training, and other characteristics. Results Latent growth model analyses revealed that memory training was associated with improved memory performance through year five, but that neither booster training nor training adherence significantly influenced this effect. Baseline age was associated with change in memory performance attributable to the passage of time alone (i.e., to aging). Higher education and better self-rated health were associated with greater change in memory performance after training. Discussion These findings confirm that memory training can aid in maintaining long-term improvements in memory performance. Booster training and adherence to training do not appear to attenuate rates of normal age-related memory decline. PMID:23103452

  6. Dissociation of Active Working Memory and Passive Recognition in Rhesus Monkeys

    ERIC Educational Resources Information Center

    Basile, Benjamin M.; Hampton, Robert R.

    2013-01-01

    Active cognitive control of working memory is central in most human memory models, but behavioral evidence for such control in nonhuman primates is absent and neurophysiological evidence, while suggestive, is indirect. We present behavioral evidence that monkey memory for familiar images is under active cognitive control. Concurrent cognitive…

  7. Long-term exposure to superantigen induces p27Kip1 and Bcl-2 expression in effector memory CD4+ T cells.

    PubMed

    Koyanagi, Madoka; Fukada, Kenji; Uchiyama, Takehiko; Yagi, Junji; Arimura, Yutaka

    2007-08-01

    The long-term exposure of mice to superantigen SEA using a mini-osmotic pump (SEA pump) induced a long-lasting expansion of Vbeta3+ CD4+ T cells with T helper (Th) 2 cell-type properties. Removal of the SEA pump 10 days after pump implantation did not significantly alter the level of Vbeta3+ CD4+ T cell expansion/maintenance. Furthermore, CFSE-labeled CD4+ T cells failed to divide when transferred to post-implantation day 15 mice. Thus, CD4+ T cells appeared to survive for at least 30 days in the absence of a sufficient amount of antigen to trigger cell division. STAT6 deficient mice, in which Th2 cell development is largely impaired, also exhibited a protracted cell expansion, similar to that observed in normal mice, suggesting that the Th2 cell property is dispensable for the maintenance of Vbeta3+ CD4+ T cell expansion. The expanded CD4+ T cells on post-implantation day 26 were arrested in the G0/G1 phase of the cell cycle and showed a lower level of cell division upon restimulation. The Cdk inhibitor p27(Kip1) was highly expressed, and Cdk2 was downregulated. Moreover, the CD4+ T cells were resistant to in vitro apoptosis induction in parallel with their level of Bcl-2 expression. Collectively, the Vbeta3+ CD4+ T cells appeared to develop into long-lived memory T cells with cell cycle arrest upon long-term exposure to SEA.

  8. Roadmap for future research on plant pathogen effectors

    PubMed Central

    Alfano, James R.

    2009-01-01

    SUMMARY Bacterial and eukaryotic plant pathogens deliver effector proteins into plant cells to promote pathogenesis. Bacterial pathogens containing type III protein secretion systems are known to inject many of these effectors into plant cells. More recently, oomycete pathogens have been shown to possess a large family of effectors containing the RXLR motif, and many effectors are also being discovered in fungal pathogens. Although effector activities are largely unknown, at least a subset suppress plant immunity. A plethora of new plant pathogen genomes that will soon be available thanks to next-generation sequencing technologies will allow the identification of many more effectors. This article summarizes the key approaches used to identify plant pathogen effectors, many of which will continue to be useful for future effector discovery. Thus, it can be viewed as a ‘roadmap’ for effector and effector target identification. Because effectors can be used as tools to elucidate components of innate immunity, advances in our understanding of effectors and their targets should lead to improvements in agriculture. PMID:19849786

  9. Transcription activator-like effector nucleases efficiently disrupt the target gene in Iberian ribbed newts (Pleurodeles waltl), an experimental model animal for regeneration.

    PubMed

    Hayashi, Toshinori; Sakamoto, Kousuke; Sakuma, Tetsushi; Yokotani, Naoki; Inoue, Takeshi; Kawaguchi, Eri; Agata, Kiyokazu; Yamamoto, Takashi; Takeuchi, Takashi

    2014-01-01

    Regeneration of a lost tissue in an animal is an important issue. Although regenerative studies have a history of research spanning more than a century, the gene functions underlying regulation of the regeneration are mostly unclear. Analysis of knockout animals is a very powerful tool with which to elucidate gene function. Recently, transcription activator-like effector nucleases (TALENs) have been developed as an effective technique for genome editing. This technique enables gene targeting in amphibians such as newts that were previously impossible. Here we show that newts microinjected with TALEN mRNAs designed for targeting the tyrosinase gene in single-cell stage embryos revealed an albino phenotype. Sequence analysis revealed that the tyrosinase genes were effectively disrupted in these albino newts. Moreover, precise genome alteration was achieved using TALENs and single strand oligodeoxyribonucleotides. Our results suggest that TALENs are powerful tools for genome editing for regenerative research in newts.

  10. TRANSCRIPTION ACTIVATOR-LIKE EFFECTOR NUCLEASE-Mediated Generation and Metabolic Analysis of Camalexin-Deficient cyp71a12 cyp71a13 Double Knockout Lines1

    PubMed Central

    Müller, Teresa M.; Böttcher, Christoph; Morbitzer, Robert; Götz, Cornelia C.; Lehmann, Johannes; Lahaye, Thomas; Glawischnig, Erich

    2015-01-01

    In Arabidopsis (Arabidopsis thaliana), a number of defense-related metabolites are synthesized via indole-3-acetonitrile (IAN), including camalexin and indole-3-carboxylic acid (ICOOH) derivatives. Cytochrome P450 71A13 (CYP71A13) is a key enzyme for camalexin biosynthesis and catalyzes the conversion of indole-3-acetaldoxime (IAOx) to IAN. The CYP71A13 gene is located in tandem with its close homolog CYP71A12, also encoding an IAOx dehydratase. However, for CYP71A12, indole-3-carbaldehyde and cyanide were identified as major reaction products. To clarify CYP71A12 function in vivo and to better understand IAN metabolism, we generated two cyp71a12 cyp71a13 double knockout mutant lines. CYP71A12-specific transcription activator-like effector nucleases were introduced into the cyp71a13 background, and very efficient somatic mutagenesis was achieved. We observed stable transmission of the cyp71a12 mutation to the following generations, which is a major challenge for targeted mutagenesis in Arabidopsis. In contrast to cyp71a13 plants, in which camalexin accumulation is partially reduced, double mutants synthesized only traces of camalexin, demonstrating that CYP71A12 contributes to camalexin biosynthesis in leaf tissue. A major role of CYP71A12 was identified for the inducible biosynthesis of ICOOH. Specifically, the ICOOH methyl ester was reduced to 12% of the wild-type level in AgNO3-challenged cyp71a12 leaves. In contrast, indole-3-carbaldehyde derivatives apparently are synthesized via alternative pathways, such as the degradation of indole glucosinolates. Based on these results, we present a model for this surprisingly complex metabolic network with multiple IAN sources and channeling of IAOx-derived IAN into camalexin biosynthesis. In conclusion, transcription activator-like effector nuclease-mediated mutation is a powerful tool for functional analysis of tandem genes in secondary metabolism. PMID:25953104

  11. Human intracranial high-frequency activity during memory processing: neural oscillations or stochastic volatility?

    PubMed Central

    Burke, John F.; Ramayya, Ashwin G.; Kahana, Michael J.

    2015-01-01

    Intracranial high-frequency activity (HFA), which refers to fast fluctuations in electrophysiological recordings, increases during memory processing. Two views have emerged to explain this effect: (1) HFA reflects a synchronous signal, related to underlying gamma oscillations, that plays a mechanistic role in human memory and (2) HFA reflects an asynchronous signal that is a nonspecific marker of brain activation. Here, we review recent data supporting each of these views and conclude that HFA during memory processing is more consistent with an asynchronous signal. Memory-related HFA is therefore best conceptualized as a biomarker of neural activation that can functionally map memory with unprecedented spatial and temporal precision. PMID:25279772

  12. Mycobacterium tuberculosis culture filtrate protein 10-specific effector/memory CD4⁺ and CD8⁺ T cells in tubercular pleural fluid, with biased usage of T cell receptor Vβ chains.

    PubMed

    Qiao, Dan; Li, Li; Guo, Jian; Lao, Suihua; Zhang, Xianlan; Zhang, Jianping; Wu, Changyou

    2011-08-01

    T cell-mediated immunity is critical for the control of Mycobacterium tuberculosis infection. Identifying the precise immune mechanisms that lead to control of initial M. tuberculosis infection and preventing reactivation of latent infection are crucial for combating tuberculosis. However, a detailed understanding of the role of T cells in the immune response to infection has been hindered. In addition, there are few flow cytometry studies characterizing the Vβ repertoires of T cell receptors (TCRs) at local sites of M. tuberculosis infection in adult tuberculosis. In this study, we used culture filtrate protein 10 (CFP-10) from M. tuberculosis to characterize T cells at local sites of infection. We simultaneously analyzed the correlation of the production of cytokines with TCR Vβ repertoires in CFP-10-specific CD4(+) and CD8(+) T cell subsets. For the first time, we demonstrate that CFP-10-specific CD4(+) or CD8(+) T cells from tubercular pleural fluid can produce high levels of gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) and upregulate the expression of CD107a/b on the cell surface. The CFP-10-specific cells were effector/memory cells with a CD45RO(+) CD62L(-) CCR7(-) CD27(-) expression profile. In addition, we found CFP-10-specific CD4(+) and CD8(+) T cells in tubercular pleural fluid, with biased usage of TCR Vβ9, Vβ12, or Vβ7.2. Our findings of CFP-10-specific CD4(+) and CD8(+) T cells in tubercular pleural fluid are critical for understanding the mechanisms of the local cellular immune response and developing more effective therapeutic interventions in cases of M. tuberculosis infection.

  13. Activation of human NKCC by moderate exercise: increased frequency of NK cells with enhanced capability of effector--target lytic interactions.

    PubMed Central

    Targan, S; Britvan, L; Dorey, F

    1981-01-01

    In the present study we examined the mechanism of human natural killer cellular cytotoxicity (NKCC) augmentation by 5 min of moderate exercise and its interrelationship to in vitro interferon (IFN) activation. Cytotoxicity was measured by employing both a single-cell cytotoxic assay and a standard 3-hr chromium-51 (51Cr) release assay. The former was used to assess changes at the single NK cell--target cell level and the latter to assess changes in overall lytic capacity of a given population of NK cells. Several findings were obtained: (1) moderate exercise augmented NKCC in vivo by recruiting a 'new' population of active cytotoxic NK cells. (2) This 'new' population of active cells probably was derived from cells which can bind targets but are non-cytotoxic. (3) In a standard 51Cr-release assay, additional augmentation of these exercise-activated cells occurred in vitro following exposure to interferon. (4) This additional increase in cytotoxicity produced no alteration in the frequency of killer cells as viewed at the single cell level. (5) Thus interferon's capacity to increase further the overall lytic ability of exercise-activated NK cells was not due to its activation of an additional subset of pre-NK cells, but due to its increasing the capacity of effector--target lytic interactions (recycling) of the same set of NK and pre-NK cells. PMID:6172225

  14. Enhanced resting-state functional connectivity between core memory-task activation peaks is associated with memory impairment in MCI.

    PubMed

    Zhang, Yifei; Simon-Vermot, Lee; Araque Caballero, Miguel Á; Gesierich, Benno; Taylor, Alexander N W; Duering, Marco; Dichgans, Martin; Ewers, Michael

    2016-09-01

    Resting-state functional connectivity (FC) is altered in Alzheimer's disease (AD) but its predictive value for episodic memory impairment is debated. Here, we aimed to assess whether resting-state FC in core brain regions activated during memory-task functional magnetic resonance imaging is altered and predictive of memory performance in AD and amnestic mild cognitive impairment (aMCI). Twenty-three elderly cognitively healthy controls (HC), 76 aMCI subjects, and 19 AD dementia patients were included. We computed resting-state FC between 18 meta-analytically determined peak coordinates of brain activation during successful memory retrieval. Higher FC between the parahippocampus, parietal cortex, and the middle frontal gyrus was observed in both AD and mild cognitive impairment compared to HC (false-discovery rate-corrected p < 0.05). The increase in FC between the parahippocampus and middle frontal gyrus was associated with reduced episodic memory in aMCI, independent of amyloid-beta positron emission tomography binding and apolipoprotein E ε4-carrier status. In conclusion, increased parahippocampal-prefrontal FC is predictive of impaired episodic memory in aMCI and may reflect a dysfunctional change within the episodic memory-related neural network. PMID:27459924

  15. Acute up-regulation of the rat brain somatostatin receptor-effector system by leptin is related to activation of insulin signaling and may counteract central leptin actions.

    PubMed

    Perianes-Cachero, A; Burgos-Ramos, E; Puebla-Jiménez, L; Canelles, S; Frago, L M; Hervás-Aguilar, A; de Frutos, S; Toledo-Lobo, M V; Mela, V; Viveros, M P; Argente, J; Chowen, J A; Arilla-Ferreiro, E; Barrios, V

    2013-11-12

    Leptin and somatostatin (SRIF) have opposite effects on food seeking and ingestive behaviors, functions partially regulated by the frontoparietal cortex and hippocampus. Although it is known that the acute suppression of food intake mediated by leptin decreases with time, the counter-regulatory mechanisms remain unclear. Our aims were to analyze the effect of acute central leptin infusion on the SRIF receptor-effector system in these areas and the implication of related intracellular signaling mechanisms in this response. We studied 20 adult male Wister rats including controls and those treated intracerebroventricularly with a single dose of 5 μg of leptin and sacrificed 1 or 6h later. Density of SRIF receptors was unchanged at 1h, whereas leptin increased the density of SRIF receptors at 6h, which was correlated with an elevated capacity of SRIF to inhibit forskolin-stimulated adenylyl cyclase activity in both areas. The functional capacity of SRIF receptors was unaltered as cell membrane levels of αi1 and αi2 subunits of G inhibitory proteins were unaffected in both brain areas. The increased density of SRIF receptors was due to enhanced SRIF receptor subtype 2 (sst2) protein levels that correlated with higher mRNA levels for this receptor. These changes in sst2 mRNA levels were concomitant with increased activation of the insulin signaling, c-Jun and cyclic AMP response element-binding protein (CREB); however, activation of signal transducer and activator of transcription 3 was reduced in the cortex and unchanged in the hippocampus and suppressor of cytokine signaling 3 remained unchanged in these areas. In addition, the leptin antagonist L39A/D40A/F41A blocked the leptin-induced changes in SRIF receptors, leptin signaling and CREB activation. In conclusion, increased activation of insulin signaling after leptin infusion is related to acute up-regulation of the SRIF receptor-effector system that may antagonize short-term leptin actions in the rat brain.

  16. Differential Neural Activity during Search of Specific and General Autobiographical Memories elicited by Musical Cues

    PubMed Central

    Ford, Jaclyn Hennessey; Addis, Donna Rose; Giovanello, Kelly S.

    2011-01-01

    Previous neuroimaging studies that have examined autobiographical memory specificity have utilized retrieval cues associated with prior searches of the event, potentially changing the retrieval processes being investigated. In the current study, musical cues were used to naturally elicit memories from multiple levels of specificity (i.e., lifetime period, general event, and event-specific). Sixteen young adults participated in a neuroimaging study in which they retrieved autobiographical memories associated with musical cues. These musical cues led to the retrieval of highly emotional memories that had low levels of prior retrieval. Retrieval of all autobiographical memory levels was associated with activity in regions in the autobiographical memory network, specifically the ventromedial prefrontal cortex, posterior cingulate, and right medial temporal lobe. Owing to the use of music, memories from varying levels of specificity were retrieved, allowing for comparison of event memory and abstract personal knowledge, as well as comparison of specific and general event memory. Dorsolateral and dorsomedial prefrontal regions were engaged during event retrieval relative to personal knowledge retrieval, and retrieval of specific event memories was associated with increased activity in the bilateral medial temporal lobe and dorsomedial prefrontal cortex relative to retrieval of general event memories. These results suggest that the initial search processes for memories of different specificity levels preferentially engage different components of the autobiographical memory network. The potential underlying causes of these neural differences are discussed. PMID:21600227

  17. The NO-cGMP-PKG Signaling Pathway Regulates Synaptic Plasticity and Fear Memory Consolidation in the Lateral Amygdala via Activation of ERK/MAP Kinase

    ERIC Educational Resources Information Center

    Ota, Kristie T.; Pierre, Vicki J.; Ploski, Jonathan E.; Queen, Kaila; Schafe, Glenn E.

    2008-01-01

    Recent studies have shown that nitric oxide (NO) signaling plays a crucial role in memory consolidation of Pavlovian fear conditioning and in synaptic plasticity in the lateral amygdala (LA). In the present experiments, we examined the role of the cGMP-dependent protein kinase (PKG), a downstream effector of NO, in fear memory consolidation and…

  18. Systematic Analysis of Bacterial Effector-Postsynaptic Density 95/Disc Large/Zonula Occludens-1 (PDZ) Domain Interactions Demonstrates Shigella OspE Protein Promotes Protein Kinase C Activation via PDLIM Proteins*

    PubMed Central

    Yi, Chae-ryun; Allen, John E.; Russo, Brian; Lee, Soo Young; Heindl, Jason E.; Baxt, Leigh A.; Herrera, Bobby Brooke; Kahoud, Emily; MacBeath, Gavin; Goldberg, Marcia B.

    2014-01-01

    Diseases caused by many Gram-negative bacterial pathogens depend on the activities of bacterial effector proteins that are delivered into eukaryotic cells via specialized secretion systems. Effector protein function largely depends on specific subcellular targeting and specific interactions with cellular ligands. PDZ domains are common domains that serve to provide specificity in protein-protein interactions in eukaryotic systems. We show that putative PDZ-binding motifs are significantly enriched among effector proteins delivered into mammalian cells by certain bacterial pathogens. We use PDZ domain microarrays to identify candidate interaction partners of the Shigella flexneri effector proteins OspE1 and OspE2, which contain putative PDZ-binding motifs. We demonstrate in vitro and in cells that OspE proteins interact with PDLIM7, a member of the PDLIM family of proteins, which contain a PDZ domain and one or more LIM domains, protein interaction domains that participate in a wide variety of functions, including activation of isoforms of protein kinase C (PKC). We demonstrate that activation of PKC during S. flexneri infection is attenuated in the absence of PDLIM7 or OspE proteins and that the OspE PDZ-binding motif is required for wild-type levels of PKC activation. These results are consistent with a model in which binding of OspE to PDLIM7 during infection regulates the activity of PKC isoforms that bind to the PDLIM7 LIM domain. PMID:25124035

  19. Systematic analysis of bacterial effector-postsynaptic density 95/disc large/zonula occludens-1 (PDZ) domain interactions demonstrates Shigella OspE protein promotes protein kinase C activation via PDLIM proteins.

    PubMed

    Yi, Chae-ryun; Allen, John E; Russo, Brian; Lee, Soo Young; Heindl, Jason E; Baxt, Leigh A; Herrera, Bobby Brooke; Kahoud, Emily; MacBeath, Gavin; Goldberg, Marcia B

    2014-10-24

    Diseases caused by many Gram-negative bacterial pathogens depend on the activities of bacterial effector proteins that are delivered into eukaryotic cells via specialized secretion systems. Effector protein function largely depends on specific subcellular targeting and specific interactions with cellular ligands. PDZ domains are common domains that serve to provide specificity in protein-protein interactions in eukaryotic systems. We show that putative PDZ-binding motifs are significantly enriched among effector proteins delivered into mammalian cells by certain bacterial pathogens. We use PDZ domain microarrays to identify candidate interaction partners of the Shigella flexneri effector proteins OspE1 and OspE2, which contain putative PDZ-binding motifs. We demonstrate in vitro and in cells that OspE proteins interact with PDLIM7, a member of the PDLIM family of proteins, which contain a PDZ domain and one or more LIM domains, protein interaction domains that participate in a wide variety of functions, including activation of isoforms of protein kinase C (PKC). We demonstrate that activation of PKC during S. flexneri infection is attenuated in the absence of PDLIM7 or OspE proteins and that the OspE PDZ-binding motif is required for wild-type levels of PKC activation. These results are consistent with a model in which binding of OspE to PDLIM7 during infection regulates the activity of PKC isoforms that bind to the PDLIM7 LIM domain.

  20. Mycobacterium tuberculosis PstS1 amplifies IFN-γ and induces IL-17/IL-22 responses by unrelated memory CD4+ T cells via dendritic cell activation.

    PubMed

    Palma, Carla; Schiavoni, Giovanna; Abalsamo, Laura; Mattei, Fabrizio; Piccaro, Giovanni; Sanchez, Massimo; Fernandez, Carmen; Singh, Mahavir; Gabriele, Lucia

    2013-09-01

    The immunological mechanisms that modulate protection during Mycobacterium tuberculosis (Mtb) infection or vaccination are not fully understood. Secretion of IFN-γ and, to a lesser extent, of IL-17 by CD4(+) T cells plays a major role both in protection and immunopathology. Few Mtb Ags interacting with DCs affect priming, activation, and regulation of Ag-unrelated CD4(+) T-cell responses. Here we demonstrate that PstS1, a 38 kDa-lipoprotein of Mtb, promotes Ag-independent activation of memory T lymphocytes specific for Ag85B or Ag85A, two immunodominant protective Ags of Mtb. PstS1 expands CD4(+) and CD8(+) memory T cells, amplifies secretion of IFN-γ and IL-22 and induces IL-17 production by effector memory cells in an Ag-unrelated manner in vitro and in vivo. These effects were mediated through the stimulation of DCs, particularly of the CD8α(-) subtype, which respond to PstS1 by undergoing phenotypic maturation and by secreting IL-6, IL-1β and, to a lower extent, IL-23. IL-6 secretion by PstS1-stimulated DCs was required for IFN-γ, and to a lesser extent for IL-22 responses by Ag85B-specific memory T cells. These results may open new perspectives for immunotherapeutic strategies to control Th1/Th17 immune responses in Mtb infections and in vaccinations against tuberculosis. PMID:23719937

  1. The last half-repeat of transcription activator-like effector (TALE) is dispensable and thereby TALE-based technology can be simplified.

    PubMed

    Zheng, Chong-Ke; Wang, Chun-Lian; Zhang, Xiao-Ping; Wang, Fu-Jun; Qin, Teng-Fei; Zhao, Kai-Jun

    2014-09-01

    To activate the expression of host genes that contribute to pathogen growth, pathogenic Xanthomonas bacteria inject their transcription activator-like effectors (TALEs) into plant cells and the TALEs bind to target gene promoters by the central repeat region consisting of near-perfect 34-amino-acid repeats (34-aa repeats). Based on the recognition codes between the 34-aa repeats and the targeted nucleotides, TALE-based technologies, such as designer TALEs (dTALEs) and TALE nucleases (TALENs), have been developed. Amazingly, every natural TALE invariantly has a truncated last half-repeat (LHR) at the end of the 34-aa repeats. Consequently, all the reported dTALEs and TALENs also harbour their LHRs. Here, we show that the LHRs in dTALEs are dispensable for the function of gene activation by both transient expression assays in Nicotiana benthamiana and gene-specific targeting in the rice genome, indicating that TALEs might originate from a single progenitor. In the light of this finding, we demonstrate that dTALEs can be constructed through two simple steps. Moreover, the activation strengths of dTALEs lacking the LHR are comparable with those of dTALEs harbouring the LHR. Our results provide new insights into the origin of natural TALEs, and will facilitate the simplification of the design and assembly of TALE-based tools, such as dTALEs and TALENs, in the near future.

  2. Adaptive top-down suppression of hippocampal activity and the purging of intrusive memories from consciousness.

    PubMed

    Benoit, Roland G; Hulbert, Justin C; Huddleston, Ean; Anderson, Michael C

    2015-01-01

    When reminded of unwanted memories, people often attempt to suppress these experiences from awareness. Prior work indicates that control processes mediated by the dorsolateral prefrontal cortex (DLPFC) modulate hippocampal activity during such retrieval suppression. It remains unknown whether this modulation plays a role in purging an intrusive memory from consciousness. Here, we combined fMRI and effective connectivity analyses with phenomenological reports to scrutinize a role for adaptive top-down suppression of hippocampal retrieval processes in terminating mnemonic awareness of intrusive memories. Participants either suppressed or recalled memories of pictures depicting faces or places. After each trial, they reported their success at regulating awareness of the memory. DLPFC activation was greatest when unwanted memories intruded into consciousness and needed to be purged, and this increased engagement predicted superior control of intrusive memories over time. However, hippocampal activity was decreased during the suppression of place memories only. Importantly, the inhibitory influence of the DLPFC on the hippocampus was linked to the ensuing reduction in intrusions of the suppressed memories. Individuals who exhibited negative top-down coupling during early suppression attempts experienced fewer involuntary memory intrusions later on. Over repeated suppressions, the DLPFC-hippocampus connectivity grew less negative with the degree that they no longer had to purge unwanted memories from awareness. These findings support a role of DLPFC in countermanding the unfolding recollection of an unwanted memory via the suppression of hippocampal processing, a mechanism that may contribute to adaptation in the aftermath of traumatic experiences.

  3. Chronic parasitic infection maintains high frequencies of short-lived Ly6C+CD4+ effector T cells that are required for protection against re-infection.

    PubMed

    Peters, Nathan C; Pagán, Antonio J; Lawyer, Phillip G; Hand, Timothy W; Henrique Roma, Eric; Stamper, Lisa W; Romano, Audrey; Sacks, David L

    2014-12-01

    In contrast to the ability of long-lived CD8(+) memory T cells to mediate protection against systemic viral infections, the relationship between CD4(+) T cell memory and acquired resistance against infectious pathogens remains poorly defined. This is especially true for T helper 1 (Th1) concomitant immunity, in which protection against reinfection coincides with a persisting primary infection. In these situations, pre-existing effector CD4 T cells generated by ongoing chronic infection, not memory cells, may be essential for protection against reinfection. We present a systematic study of the tissue homing properties, functionality, and life span of subsets of memory and effector CD4 T cells activated in the setting of chronic Leishmania major infection in resistant C57Bl/6 mice. We found that pre-existing, CD44(+)CD62L(-)T-bet(+)Ly6C+ effector (T(EFF)) cells that are short-lived in the absence of infection and are not derived from memory cells reactivated by secondary challenge, mediate concomitant immunity. Upon adoptive transfer and challenge, non-dividing Ly6C(+) T(EFF) cells preferentially homed to the skin, released IFN-γ, and conferred protection as compared to CD44(+)CD62L(-)Ly6C(-) effector memory or CD44(+)CD62L(+)Ly6C(-) central memory cells. During chronic infection, Ly6C(+) T(EFF) cells were maintained at high frequencies via reactivation of T(CM) and the T(EFF) themselves. The lack of effective vaccines for many chronic diseases may be because protection against infectious challenge requires the maintenance of pre-existing T(EFF) cells, and is therefore not amenable to conventional, memory inducing, vaccination strategies.

  4. The NO-cGMP-PKG signaling pathway regulates synaptic plasticity and fear memory consolidation in the lateral amygdala via activation of ERK/MAP kinase.

    PubMed

    Ota, Kristie T; Pierre, Vicki J; Ploski, Jonathan E; Queen, Kaila; Schafe, Glenn E

    2008-10-01

    Recent studies have shown that nitric oxide (NO) signaling plays a crucial role in memory consolidation of Pavlovian fear conditioning and in synaptic plasticity in the lateral amygdala (LA). In the present experiments, we examined the role of the cGMP-dependent protein kinase (PKG), a downstream effector of NO, in fear memory consolidation and long-term potentiation (LTP) at thalamic and cortical input pathways to the LA. In behavioral experiments, rats given intra-LA infusions of either the PKG inhibitor Rp-8-Br-PET-cGMPS or the PKG activator 8-Br-cGMP exhibited dose-dependent impairments or enhancements of fear memory consolidation, respectively. In slice electrophysiology experiments, bath application of Rp-8-Br-PET-cGMPS or the guanylyl cyclase inhibitor LY83583 impaired LTP at thalamic, but not cortical inputs to the LA, while bath application of 8-Br-cGMP or the guanylyl cyclase activator YC-1 resulted in enhanced LTP at thalamic inputs to the LA. Interestingly, YC-1-induced enhancement of LTP in the LA was reversed by concurrent application of the MEK inhibitor U0126, suggesting that the NO-cGMP-PKG signaling pathway may promote synaptic plasticity and fear memory formation in the LA, in part by activating the ERK/MAPK signaling cascade. As a test of this hypothesis, we next showed that rats given intra-LA infusion of the PKG inhibitor Rp-8-Br-PET-cGMPS or the PKG activator 8-Br-cGMP exhibit impaired or enhanced activation, respectively, of ERK/MAPK in the LA after fear conditioning. Collectively, our findings suggest that an NO-cGMP-PKG-dependent form of synaptic plasticity at thalamic input synapses to the LA may underlie memory consolidation of Pavlovian fear conditioning, in part, via activation of the ERK/MAPK signaling cascade.

  5. Visualizing TCR-induced POLKADOTS formation and NF-κB activation in the D10 T-cell clone and mouse primary effector T cells.

    PubMed

    Paul, Suman; Schaefer, Brian C

    2015-01-01

    T cells are an immune cell lineage that play a central role in protection against pathogen infection. Antigen, in the form of pathogen-derived peptides, stimulates the T-cell receptor (TCR), leading to activation of the transcription factor, nuclear factor kappa B (NF-κB). The subsequent NF-κB-dependent gene expression program drives expansion and effector differentiation of antigen-specific T cells, leading to the adaptive anti-pathogen immune response. The cell surface TCR transmits activating signals to cytosolic NF-κB by a complex signaling cascade, in which the adapter protein Bcl10 plays a key role. We have previously demonstrated that TCR engagement leads to the formation of cytosolic Bcl10 clusters, called POLKADOTS, that provide a platform for the assembly of the terminal signaling complex that ultimately mediates NF-κB activation. In this chapter, we describe the methods utilized to visualize the formation of TCR-induced POLKADOTS and to study the temporal association between POLKADOTS formation and nuclear translocation of the NF-κB subunit, RelA/p65.

  6. Memories.

    ERIC Educational Resources Information Center

    Brand, Judith, Ed.

    1998-01-01

    This theme issue of the journal "Exploring" covers the topic of "memories" and describes an exhibition at San Francisco's Exploratorium that ran from May 22, 1998 through January 1999 and that contained over 40 hands-on exhibits, demonstrations, artworks, images, sounds, smells, and tastes that demonstrated and depicted the biological,…

  7. Exploitation of Eukaryotic Subcellular Targeting Mechanisms by Bacterial Effectors

    PubMed Central

    Hicks, Stuart W.; Galán, Jorge E.

    2013-01-01

    Several bacteria have evolved specialized secretion systems to deliver bacterial effector proteins into eukaryotic cells with the capacity to modulate cellular pathways to promote bacterial survival and replication. The spatial and temporal context in which effectors exert their biochemical activities is critical for their function. Understanding the mechanisms that lead to their precise subcellular localization following delivery into host cells is essential for understanding effector function in the context of infection. Recent studies have shown that bacterial effectors exploit host cellular machinery to accurately target their biochemical activities within the host cell. PMID:23588250

  8. Estrogen-dependent transcriptional activation and vitellogenin gene memory.

    PubMed

    Edinger, R S; Mambo, E; Evans, M I

    1997-12-01

    The concept of hepatic memory suggests that a gene responds more rapidly to a second exposure of an inducer than it does during the initial activation. To determine how soon estrogen-dependent DNA/protein interactions occur during the primary response, in vivo dimethylsulfate footprinting was carried out using genomic DNA amplified by ligation-mediated PCR. When estrogen was added to disrupted cells from a hormone-naive liver, changes within and around the estrogen response elements occurred within seconds, indicating a direct and rapid effect on this estrogen-responsive promoter that had never before been activated. Because this effect was so rapid relative to the delayed onset of mRNA accumulation during the primary response, run-on transcription assays were used to determine the transcription profiles for four of the yolk protein genes during the primary and secondary responses to estrogen. As with the accumulation of mRNA, the onset of transcription was delayed for all of these genes after a primary exposure to estrogen. Interestingly, after the secondary exposure to estrogen, the vitellogenin I, vitellogenin II, and very low density apolipoprotein II genes displayed a more rapid onset of transcription, whereas the primary and secondary profiles of apolipoprotein B transcription in response to estrogen were identical. Because the apoB gene is constitutively expressed in the absence of estrogen, and the vitellogenins are quiescent before the administration of the hormone, hepatic memory most likely represents a relatively stable event in the transition to an active state of a gene that is committed for tissue-specific expression.

  9. Structural requirements for the activation of Escherichia coli CTP synthase by the allosteric effector GTP are stringent, but requirements for inhibition are lax.

    PubMed

    Lunn, Faylene A; MacDonnell, Jennifer E; Bearne, Stephen L

    2008-01-25

    Cytidine 5'-triphosphate synthase catalyzes the ATP-dependent formation of CTP from UTP using either NH(3) or l-glutamine (Gln) as the source of nitrogen. GTP acts as an allosteric effector promoting Gln hydrolysis but inhibiting Gln-dependent CTP formation at concentrations of >0.15 mM and NH(3)-dependent CTP formation at all concentrations. A structure-activity study using a variety of GTP and guanosine analogues revealed that only a few GTP analogues were capable of activating Gln-dependent CTP formation to varying degrees: GTP approximately 6-thio-GTP > ITP approximately guanosine 5'-tetraphosphate > O(6)-methyl-GTP > 2'-deoxy-GTP. No activation was observed with guanosine, GMP, GDP, 2',3'-dideoxy-GTP, acycloguanosine, and acycloguanosine monophosphate, indicating that the 5'-triphosphate, 2'-OH, and 3'-OH are required for full activation. The 2-NH(2) group plays an important role in binding recognition, whereas substituents at the 6-position play an important role in activation. The presence of a 6-NH(2) group obviates activation, consistent with the inability of ATP to substitute for GTP. Nucleotide and nucleoside analogues of GTP and guanosine, respectively, all inhibited NH(3)- and Gln-dependent CTP formation (often in a cooperative manner) to a similar extent (IC(50) approximately 0.2-0.5 mM). This inhibition appeared to be due solely to the purine base and was relatively insensitive to the identity of the purine with the exception of inosine, ITP, and adenosine (IC(50) approximately 4-12 mM). 8-Oxoguanosine was the best inhibitor identified (IC(50) = 80 microM). Our findings suggest that modifying 2-aminopurine or 2-aminopurine riboside may serve as an effective strategy for developing cytidine 5'-triphosphate synthase inhibitors. PMID:18003612

  10. Vaccinia virus entry is followed by core activation and proteasome-mediated release of the immunomodulatory effector VH1 from lateral bodies.

    PubMed

    Schmidt, Florian Ingo; Bleck, Christopher Karl Ernst; Reh, Lucia; Novy, Karel; Wollscheid, Bernd; Helenius, Ari; Stahlberg, Henning; Mercer, Jason

    2013-08-15

    Host cell entry of vaccinia virus, the prototypic poxvirus, involves a membrane fusion event delivering the viral core and two proteinaceous lateral bodies (LBs) into the cytosol. Uncoating of viral cores is poorly characterized, and the composition and function of LBs remains enigmatic. We found that cytosolic cores rapidly dissociated from LBs and expanded in volume, which coincided with reduction of disulfide-bonded core proteins. We identified the abundant phosphoprotein F17, the dual-specificity phosphatase VH1, and the oxidoreductase G4 as bona fide LB components. After reaching the cytosol, F17 was degraded in a proteasome-dependent manner. Proteasome activity, and presumably LB disassembly, was required for the immediate immunomodulatory activity of VH1: dephosphorylation of STAT1 to prevent interferon-γ-mediated antiviral responses. These results reveal a mechanism used by poxviruses to deliver viral enzymes to the host cell cytosol and are likely to facilitate the identification of additional LB-resident viral effectors. PMID:23891003

  11. Activation and polar sequestration of PopA, a c-di-GMP effector protein involved in Caulobacter crescentus cell cycle control.

    PubMed

    Ozaki, Shogo; Schalch-Moser, Annina; Zumthor, Ludwig; Manfredi, Pablo; Ebbensgaard, Anna; Schirmer, Tilman; Jenal, Urs

    2014-11-01

    When Caulobacter crescentus enters S-phase the replication initiation inhibitor CtrA dynamically positions to the old cell pole to be degraded by the polar ClpXP protease. Polar delivery of CtrA requires PopA and the diguanylate cyclase PleD that positions to the same pole. Here we present evidence that PopA originated through gene duplication from its paralogue response regulator PleD and subsequent co-option as c-di-GMP effector protein. While the C-terminal catalytic domain (GGDEF) of PleD is activated by phosphorylation of the N-terminal receiver domain, functional adaptation has reversed signal transduction in PopA with the GGDEF domain adopting input function and the receiver domain serving as regulatory output. We show that the N-terminal receiver domain of PopA specifically interacts with RcdA, a component required for CtrA degradation. In contrast, the GGDEF domain serves to target PopA to the cell pole in response to c-di-GMP binding. In agreement with the divergent activation and targeting mechanisms, distinct markers sequester PleD and PopA to the old cell pole upon S-phase entry. Together these data indicate that PopA adopted a novel role as topology specificity factor to help recruit components of the CtrA degradation pathway to the protease specific old cell pole of C. crescentus.

  12. The Invertebrate Lysozyme Effector ILYS-3 Is Systemically Activated in Response to Danger Signals and Confers Antimicrobial Protection in C. elegans

    PubMed Central

    Gravato-Nobre, Maria João; Vaz, Filipa; Filipe, Sergio; Chalmers, Ronald; Hodgkin, Jonathan

    2016-01-01

    Little is known about the relative contributions and importance of antibacterial effectors in the nematode C. elegans, despite extensive work on the innate immune responses in this organism. We report an investigation of the expression, function and regulation of the six ilys (invertebrate-type lysozyme) genes of C. elegans. These genes exhibited a surprising variety of tissue-specific expression patterns and responses to starvation or bacterial infection. The most strongly expressed, ilys-3, was investigated in detail. ILYS-3 protein was expressed constitutively in the pharynx and coelomocytes, and dynamically in the intestine. Analysis of mutants showed that ILYS-3 was required for pharyngeal grinding (disruption of bacterial cells) during normal growth and consequently it contributes to longevity, as well as being protective against bacterial pathogens. Both starvation and challenge with Gram-positive pathogens resulted in ERK-MAPK-dependent up-regulation of ilys-3 in the intestine. The intestinal induction by pathogens, but not starvation, was found to be dependent on MPK-1 activity in the pharynx rather than in the intestine, demonstrating unexpected communication between these two tissues. The coelomocyte expression appeared to contribute little to normal growth or immunity. Recombinant ILYS-3 protein was found to exhibit appropriate lytic activity against Gram-positive cell wall material. PMID:27525822

  13. Control of pathogenic effector T-cell activities in situ by PD-L1 expression on respiratory inflammatory dendritic cells during respiratory syncytial virus infection

    PubMed Central

    Yao, S; Jiang, L; Moser, EK; Jewett, LB; Wright, J; Du, J; Zhou, B; Davis, SD; Krupp, NL; Braciale, TJ; Sun, J

    2015-01-01

    Respiratory syncytial virus (RSV) infection is a leading cause of severe lower respiratory tract illness in young infants, the elderly and immunocompromised individuals. We demonstrate here that the co-inhibitory molecule programmed cell death 1 (PD-1) is selectively upregulated on T cells within the respiratory tract during both murine and human RSV infection. Importantly, the interaction of PD-1 with its ligand PD-L1 is vital to restrict the pro-inflammatory activities of lung effector T cells in situ, thereby inhibiting the development of excessive pulmonary inflammation and injury during RSV infection. We further identify that PD-L1 expression on lung inflammatory dendritic cells is critical to suppress inflammatory T-cell activities, and an interferon–STAT1–IRF1 axis is responsible for increased PD-L1 expression on lung inflammatory dendritic cells. Our findings suggest a potentially critical role of PD-L1 and PD-1 interactions in the lung for controlling host inflammatory responses and disease progression in clinical RSV infection. PMID:25465101

  14. The Invertebrate Lysozyme Effector ILYS-3 Is Systemically Activated in Response to Danger Signals and Confers Antimicrobial Protection in C. elegans.

    PubMed

    Gravato-Nobre, Maria João; Vaz, Filipa; Filipe, Sergio; Chalmers, Ronald; Hodgkin, Jonathan

    2016-08-01

    Little is known about the relative contributions and importance of antibacterial effectors in the nematode C. elegans, despite extensive work on the innate immune responses in this organism. We report an investigation of the expression, function and regulation of the six ilys (invertebrate-type lysozyme) genes of C. elegans. These genes exhibited a surprising variety of tissue-specific expression patterns and responses to starvation or bacterial infection. The most strongly expressed, ilys-3, was investigated in detail. ILYS-3 protein was expressed constitutively in the pharynx and coelomocytes, and dynamically in the intestine. Analysis of mutants showed that ILYS-3 was required for pharyngeal grinding (disruption of bacterial cells) during normal growth and consequently it contributes to longevity, as well as being protective against bacterial pathogens. Both starvation and challenge with Gram-positive pathogens resulted in ERK-MAPK-dependent up-regulation of ilys-3 in the intestine. The intestinal induction by pathogens, but not starvation, was found to be dependent on MPK-1 activity in the pharynx rather than in the intestine, demonstrating unexpected communication between these two tissues. The coelomocyte expression appeared to contribute little to normal growth or immunity. Recombinant ILYS-3 protein was found to exhibit appropriate lytic activity against Gram-positive cell wall material. PMID:27525822

  15. The Sensory Nature of Episodic Memory: Sensory Priming Effects Due to Memory Trace Activation

    ERIC Educational Resources Information Center

    Brunel, Lionel; Labeye, Elodie; Lesourd, Mathieu; Versace, Remy

    2009-01-01

    The aim of this study was to provide evidence that memory and perceptual processing are underpinned by the same mechanisms. Specifically, the authors conducted 3 experiments that emphasized the sensory aspect of memory traces. They examined their predictions with a short-term priming paradigm based on 2 distinct phases: a learning phase consisting…

  16. Colostrum-derived B and T cells as an extra-lymphoid compartment of effector cell populations in humans.

    PubMed

    Peroni, Diego G; Chirumbolo, Salvatore; Veneri, Dino; Piacentini, Giorgio L; Tenero, Laura; Vella, Antonio; Ortolani, Riccardo; Raffaelli, Ricciarda; Boner, Attilio L

    2013-01-01

    Colostrum contains cellular components that convey immunological protection to offspring. In the present study the main subsets of lymphocytes present in colostrum and in peripheral blood of healthy screened mothers were compared through the evaluation of >15 different flow cytometry markers. Colostrum and peripheral blood samples were collected within 3 days after full-term delivery. Flow cytometry assays and laboratory tests were performed soon after collection. Among B cells, percentages of CD19(+)CD5(+) cells, pertaining to natural immunity system, were significantly higher in colostrum than in peripheral blood (33 vs. 5%, p = 0.047). CD4(+) T cells, effector cells (CD45RA(+)/CD27(-)) and effector memory cells (CD45RA(-)/CD27(-)) were significantly higher in colostrum (p < 0.001) than in peripheral blood, as well as activated CD4(+) T cells (HLA(-)DR(+)) (36% vs. 6% p = 0.0022) and CD4(+) terminally differentiated effector T cells (CD57(+)) (p < 0.001). With regards to CD8(+) T cells, a comparable significant increase in effector (p < 0.02) and effector memory cells (p < 0.001) was also observed. Moreover, an increased surface expression of HLA-DR and CD57 (p < 0.001) on CD8(+) T cells in colostrum was detected. Colostrum contains a different distribution of lymphocyte subsets with respect to peripheral blood from mothers, confirming the observation that lymphocytes probably migrate in milk in a selective way. Colostrum T and B lymphocytes appear to be enriched with subsets possessing effector functions or belonging to the innate immune system, what could transfer a prompt line of defence to offspring.

  17. Robotic end effector

    DOEpatents

    Minichan, R.L.

    1993-10-05

    An end effector is described for use in probing a surface with a robotic arm. The end effector has a first portion that carries a gimbal with a probe, the gimbal holding the probe normal to the surface, and a second portion with a set of three shafts within a housing for urging the gimbal and probe against the surface. The second portion contains a potentiometer connected by another shaft to the first portion to measure the position of the first portion with respect to the second so that the second portion can be moved to place and maintain the shafts at the midpoint of their travel. Then, as irregularities in the surface are encountered, the first portion can respond by moving closer to or farther from the second portion. 7 figures.

  18. Robotic end effector

    DOEpatents

    Minichan, Richard L.

    1993-01-01

    An end effector for use in probing a surface with a robotic arm. The end effector has a first portion that carries a gimbal with a probe, the gimbal holding the probe normal to the surface, and a second portion with a set of three shafts within a housing for urging the gimbal and probe against the surface. The second portion contains a potentiometer connected by another shaft to the first portion to measure the position of the first portion with respect to the second so that the second portion can be moved to place and maintain the shafts at the midpoint of their travel. Then, as irregularities in the surface are encountered, the first portion can respond by moving closer to or farther from the second portion.

  19. A single binding site mediates resistance- and disease-associated activities of the effector protein NIP1 from the barley pathogen Rhynchosporium secalis.

    PubMed

    van't Slot, Klaas A E; Gierlich, Angela; Knogge, Wolfgang

    2007-07-01

    The effector protein NIP1 from the barley (Hordeum vulgare) pathogen Rhynchosporium secalis specifically induces the synthesis of defense-related proteins in cultivars of barley expressing the complementary resistance gene, Rrs1. In addition, it stimulates the activity of the barley plasma membrane H(+)-ATPase in a genotype-unspecific manner and it induces necrotic lesions in leaf tissues of barley and other cereal plant species. NIP1 variants type I and II, which display quantitative differences in their activities as elicitor and H(+)-ATPase stimulator, and the inactive mutant variants type III* and type IV*, were produced in Escherichia coli. Binding studies using (125)I-NIP1 type I revealed a single class of binding sites with identical binding characteristics in microsomes from near-isogenic resistant (Rrs1) and susceptible (rrs1) barley. Binding was specific, reversible, and saturable, and saturation ligand-binding experiments yielded a K(d) of 5.6 nm. A binding site was also found in rye (Secale cereale) and the nonhost species wheat (Triticum aestivum), oat (Avena sativa), and maize (Zea mays), but not in Arabidopsis (Arabidopsis thaliana). For NIP1 types I and II, equilibrium competition-binding experiments revealed a correlation between the difference in their affinities to the binding site and the differences in their elicitor activity and H(+)-ATPase stimulation, indicating a single target molecule to mediate both activities. In contrast, the inactive proteins type III* and type IV* are both characterized by high affinities similar to type I, suggesting that binding of NIP1 to this target is not sufficient for its activities. PMID:17478637

  20. Unscheduled Akt-Triggered Activation of Cyclin-Dependent Kinase 2 as a Key Effector Mechanism of Apoptin's Anticancer Toxicity▿

    PubMed Central

    Maddika, Subbareddy; Panigrahi, Soumya; Wiechec, Emilia; Wesselborg, Sebastian; Fischer, Ute; Schulze-Osthoff, Klaus; Los, Marek

    2009-01-01

    Apoptin, a protein from the chicken anemia virus, has attracted attention because it specifically kills tumor cells while leaving normal cells unharmed. The reason for this tumor selectivity is unclear and depends on subcellular localization, as apoptin resides in the cytoplasm of normal cells but in the nuclei of transformed cells. It was shown that nuclear localization and tumor-specific killing crucially require apoptin's phosphorylation by an as yet unknown kinase. Here we elucidate the pathway of apoptin-induced apoptosis and show that it essentially depends on abnormal phosphatidylinositol 3-kinase (PI3-kinase)/Akt activation, resulting in the activation of the cyclin-dependent kinase CDK2. Inhibitors as well as dominant-negative mutants of PI3-kinase and Akt not only inhibited CDK2 activation but also protected cells from apoptin-induced cell death. Akt activated CDK2 by direct phosphorylation as well as by the phosphorylation-induced degradation of the inhibitor p27Kip1. Importantly, we also identified CDK2 as the principal kinase that phosphorylates apoptin and is crucially required for apoptin-induced cell death. Immortalized CDK2-deficient fibroblasts and CDK2 knockdown cells were markedly protected against apoptin. Thus, our results not only decipher the pathway of apoptin-induced cell death but also provide mechanistic insights for the selective killing of tumor cells. PMID:19103742

  1. Modification of Bacterial Effector Proteins Inside Eukaryotic Host Cells

    PubMed Central

    Popa, Crina M.; Tabuchi, Mitsuaki; Valls, Marc

    2016-01-01

    Pathogenic bacteria manipulate their hosts by delivering a number of virulence proteins -called effectors- directly into the plant or animal cells. Recent findings have shown that such effectors can suffer covalent modifications inside the eukaryotic cells. Here, we summarize the recent reports where effector modifications by the eukaryotic machinery have been described. We restrict our focus on proteins secreted by the type III or type IV systems, excluding other bacterial toxins. We describe the known examples of effectors whose enzymatic activity is triggered by interaction with plant and animal cell factors, including GTPases, E2-Ubiquitin conjugates, cyclophilin and thioredoxins. We focus on the structural interactions with these factors and their influence on effector function. We also review the described examples of host-mediated post-translational effector modifications which are required for proper subcellular location and function. These host-specific covalent modifications include phosphorylation, ubiquitination, SUMOylation, and lipidations such as prenylation, fatty acylation and phospholipid binding. PMID:27489796

  2. Modification of Bacterial Effector Proteins Inside Eukaryotic Host Cells.

    PubMed

    Popa, Crina M; Tabuchi, Mitsuaki; Valls, Marc

    2016-01-01

    Pathogenic bacteria manipulate their hosts by delivering a number of virulence proteins -called effectors- directly into the plant or animal cells. Recent findings have shown that such effectors can suffer covalent modifications inside the eukaryotic cells. Here, we summarize the recent reports where effector modifications by the eukaryotic machinery have been described. We restrict our focus on proteins secreted by the type III or type IV systems, excluding other bacterial toxins. We describe the known examples of effectors whose enzymatic activity is triggered by interaction with plant and animal cell factors, including GTPases, E2-Ubiquitin conjugates, cyclophilin and thioredoxins. We focus on the structural interactions with these factors and their influence on effector function. We also review the described examples of host-mediated post-translational effector modifications which are required for proper subcellular location and function. These host-specific covalent modifications include phosphorylation, ubiquitination, SUMOylation, and lipidations such as prenylation, fatty acylation and phospholipid binding.

  3. The presence and preferential activation of Tregs diminishes adoptive transfer of autoimmune diabetes by polyclonal NOD T cell effectors into NSG versus NOD-scid mice1

    PubMed Central

    Presa, Maximiliano; Chen, Yi-Guang; Grier, Alexandra E.; Leiter, Edward H.; Brehm, Michael A.; Greiner, Dale L.; Shultz, Leonard D.; Serreze, David V.

    2015-01-01

    NOD-scid.Il2rgnull (NSG) mice are currently being used as recipients to screen for pathogenic autoreactive T-cells in Type 1 Diabetes (T1D) patients. We questioned whether the restriction of IL-2 receptor gamma chain (Il-2rγ) dependent cytokine signaling only to donor cells in NSG recipients differently influenced the activities of transferred diabetogenic T-cells when they were introduced as a monoclonal/oligoclonal population versus being part of a polyclonal repertoire. Unexpectedly, a significantly decreased T1D transfer by splenocytes from prediabetic NOD donors was observed in Il2rγnull -NSG versus Il2rγ-intact standard NOD-scid recipients. In contrast, NOD-derived monoclonal/oligoclonal TCR transgenic ß-cell autoreactive T-cells in either the CD8 (AI4, NY8.3) or CD4 (BDC2.5) compartments transferred disease significantly more rapidly to NSG than to NOD-scid recipients. The reduced diabetes transfer efficiency by polyclonal T cells in NSG recipients was associated with enhanced activation of regulatory T-cells (Tregs) mediated by NSG myeloid APC. This enhanced suppressor activity was associated with higher levels of Treg GITR expression in the presence of NSG than NOD-scid APC. These collective results indicate NSG recipients might be efficiently employed to test the activity of T1D patient-derived ß-cell autoreactive T-cell clones and lines, but when screening for pathogenic effectors within polyclonal populations, Tregs should be removed from the transfer inoculum to avoid false negative results. PMID:26283479

  4. Structure and activity of the Cas3 HD nuclease MJ0384, an effector enzyme of the CRISPR interference

    SciTech Connect

    Beloglazova, Natalia; Petit, Pierre; Flick, Robert; Brown, Greg; Savchenko, Alexei; Yakunin, Alexander F.

    2012-03-15

    Clustered regularly interspaced short palindromic repeats (CRISPRs) and Cas proteins represent an adaptive microbial immunity system against viruses and plasmids. Cas3 proteins have been proposed to play a key role in the CRISPR mechanism through the direct cleavage of invasive DNA. Here, we show that the Cas3 HD domain protein MJ0384 from Methanocaldococcus jannaschii cleaves endonucleolytically and exonucleolytically (3'-5') single-stranded DNAs and RNAs, as well as 3'-flaps, splayed arms, and R-loops. The degradation of branched DNA substrates by MJ0384 is stimulated by the Cas3 helicase MJ0383 and ATP. The crystal structure of MJ0384 revealed the active site with two bound metal cations and together with site-directed mutagenesis suggested a catalytic mechanism. Our studies suggest that the Cas3 HD nucleases working together with the Cas3 helicases can completely degrade invasive DNAs through the combination of endo- and exonuclease activities.

  5. Major histocompatibility complex-unrestricted cytolytic activity of human T cells: analysis of precursor frequency and effector phenotype

    SciTech Connect

    Patel, S.S.; Thiele, D.L.; Lipsky, P.E.

    1987-12-01

    The frequency and phenotype of human T cells that mediate major histocompatibility complex (MHC)-unrestricted cytolysis were analyzed. T cell clones were generated by culturing adherent cell-depleted peripheral blood mononuclear cells at a density of 0.3 cell/well with phytohemagglutinin, recombinant interleukin 2 (rIL-2), and irradiated autologous peripheral blood mononuclear cells and/or Epstein-Barr virus-transformed lymphoblastoid cell lines. All of the 198 clones generated by this method were T cells (CD2/sup +/, CD3/sup +/, CD4/sup +/ or CD2/sup +/, CD3/sup +/, CD8/sup +/) that possessed potent lytic activity against K562, an erythroleukemia line sensitive to lysis by human natural killer cells, and Cur, a renal carcinoma cell line resistant to human natural killer activity. Cytolysis, measured by /sup 51/Cr release, was MHC-unrestricted, since the clones were able to lyse MHC class I or class II negative targets, as well as MHC class I and class II negative targets. Although the clones produced tissue necrosis factor/lymphotoxin-like molecules, lysis of Cur of K562 was not mediated by a soluble factor secreted by the clones. These data indicate that the capacity for MHC-unrestricted tumoricidal activity and expression of NKH1 and CD11b, but not CD 16, are properties common to all or nearly all human peripheral blood-derived T cell clones regardless of CD4 or CD8 phenotype.

  6. A novel genetic system to isolate a dominant negative effector on DNA-binding activity of Oct-2.

    PubMed Central

    Terunuma, A; Shiba, K; Noda, T

    1997-01-01

    Recent studies have revealed that interactions between transcription factors play an important role in regulation of gene expression in eukaryotic cells. To isolate cDNA clones that dominantly inhibit the DNA-binding activity of Oct-2, chosen as a representative factor, we have developed a novel screening system. This employs an Escherichia coli tester strain carrying a modified lac operon as a reporter gene, with the lac operator sequence replaced by an octamer sequence. Oct-2 expressed in this tester strain represses the expression of the reporter gene and changes the phenotype of the cell from Lac+to Lac-. Introduction of a cDNA expression library prepared from a human T-cell line into the Oct-2-harboring tester strain allowed selection of three Lac+clones out of 1 x 10(5) transformants. One of them, hT86, encoding a putative zinc finger protein was found to derepress beta-galactosidase activity in the Oct-2-harboring tester strain at the transcriptional level. In gel mobility shift assays, hT86 attenuated the intensity of the retarded band composed of the octamer probe and Oct-2, suggesting a dominant negative effect on the DNA-binding activity of Oct-2. The strategy described here provides a new approach for studying protein-protein interactions that govern the complex regulation of gene expression. PMID:9115366

  7. De novo-engineered transcription activator-like effector (TALE) hybrid nuclease with novel DNA binding specificity creates double-strand breaks.

    PubMed

    Mahfouz, Magdy M; Li, Lixin; Shamimuzzaman, Md; Wibowo, Anjar; Fang, Xiaoyun; Zhu, Jian-Kang

    2011-02-01

    Site-specific and rare cutting nucleases are valuable tools for genome engineering. The generation of double-strand DNA breaks (DSBs) promotes homologous recombination in eukaryotes and can facilitate gene targeting, additions, deletions, and inactivation. Zinc finger nucleases have been used to generate DSBs and subsequently, for genome editing but with low efficiency and reproducibility. The transcription activator-like family of type III effectors (TALEs) contains a central domain of tandem repeats that could be engineered to bind specific DNA targets. Here, we report the generation of a Hax3-based hybrid TALE nuclease with a user-selected DNA binding specificity. We show that the engineered TALE nuclease can bind to its target sequence in vitro and that the homodimeric TALE nuclease can cleave double-stranded DNA in vitro if the DNA binding sites have the proper spacing and orientation. Transient expression assays in tobacco leaves suggest that the hybrid nuclease creates DSB in its target sequence, which is subsequently repaired by nonhomologous end-joining repair. Taken together, our data show the feasibility of engineering TALE-based hybrid nucleases capable of generating site-specific DSBs and the great potential for site-specific genome modification in plants and eukaryotes in general.

  8. Production of α1,3-galactosyltransferase targeted pigs using transcription activator-like effector nuclease-mediated genome editing technology.

    PubMed

    Kang, Jung-Taek; Kwon, Dae-Kee; Park, A-Rum; Lee, Eun-Jin; Yun, Yun-Jin; Ji, Dal-Young; Lee, Kiho; Park, Kwang-Wook

    2016-03-01

    Recent developments in genome editing technology using meganucleases demonstrate an efficient method of producing gene edited pigs. In this study, we examined the effectiveness of the transcription activator-like effector nuclease (TALEN) system in generating specific mutations on the pig genome. Specific TALEN was designed to induce a double-strand break on exon 9 of the porcine α1,3-galactosyltransferase (GGTA1) gene as it is the main cause of hyperacute rejection after xenotransplantation. Human decay-accelerating factor (hDAF) gene, which can produce a complement inhibitor to protect cells from complement attack after xenotransplantation, was also integrated into the genome simultaneously. Plasmids coding for the TALEN pair and hDAF gene were transfected into porcine cells by electroporation to disrupt the porcine GGTA1 gene and express hDAF. The transfected cells were then sorted using a biotin-labeled IB4 lectin attached to magnetic beads to obtain GGTA1 deficient cells. As a result, we established GGTA1 knockout (KO) cell lines with biallelic modification (35.0%) and GGTA1 KO cell lines expressing hDAF (13.0%). When these cells were used for somatic cell nuclear transfer, we successfully obtained live GGTA1 KO pigs expressing hDAF. Our results demonstrate that TALEN-mediated genome editing is efficient and can be successfully used to generate gene edited pigs. PMID:27051344

  9. Knockout of a transgene by transcription activator-like effector nucleases (TALENs) in the sawfly, Athalia rosae (Hymenoptera) and the ladybird beetle, Harmonia axyridis (Coleoptera).

    PubMed

    Hatakeyama, M; Yatomi, J; Sumitani, M; Takasu, Y; Sekiné, K; Niimi, T; Sezutsu, H

    2016-02-01

    Transcription activator-like effector nucleases (TALENs) are efficient tools for targeted genome editing and have been utilized in a number of insects. Here, we demonstrate the gene disruption (knockout) caused by TALENs targeting a transgene, 3xP3-driven enhanced green fluorescence protein (EGFP), that is integrated in the genome of two species, the sawfly Athalia rosae (Hymenoptera) and the ladybird beetle Harmonia axyridis (Coleoptera). Messenger RNAs of TALENs targeting the sequences adjacent to the chromophore region were microinjected into the eggs/embryos of each species. In At. rosae, when microinjection was performed at the posterior end of eggs, 15% of G(0) individuals showed a somatic mosaic phenotype for eye EGFP fluorescence. Three-quarters of the somatic mosaics produced EGFP-negative G(1) progeny. When eggs were injected at the anterior end, 63% of the G(0) individuals showed somatic mosaicism, and 17% of them produced EGFP-negative G(1) progeny. In H. axyridis, 25% of posterior-injected and 8% of anterior-injected G(0) individuals produced EGFP-negative G(1) progeny. In both species, the EGFP-negative progeny retained the EGFP gene, and various deletions were detected in the target sequences, indicating that gene disruption was successfully induced. Finally, for both species, 18-21% of G(0) founders produced gene knockout progeny sufficient for establishing knockout strains. PMID:26496859

  10. Gadd45α activity is the principal effector of Shigella mitochondria-dependent epithelial cell death in vitro and ex vivo.

    PubMed

    Lembo-Fazio, L; Nigro, G; Noël, G; Rossi, G; Chiara, F; Tsilingiri, K; Rescigno, M; Rasola, A; Bernardini, M L

    2011-01-01

    Modulation of death is a pathogen strategy to establish residence and promote survival in host cells and tissues. Shigella spp. are human pathogens that invade colonic mucosa, where they provoke lesions caused by their ability to manipulate the host cell responses. Shigella spp. induce various types of cell death in different cell populations. However, they are equally able to protect host cells from death. Here, we have investigated on the molecular mechanisms and cell effectors governing the balance between survival and death in epithelial cells infected with Shigella. To explore these aspects, we have exploited both, the HeLa cell invasion assay and a novel ex vivo human colon organ culture model of infection that mimics natural conditions of shigellosis. Our results definitely show that Shigella induces a rapid intrinsic apoptosis of infected cells, via mitochondrial depolarization and the ensuing caspase-9 activation. Moreover, for the first time we identify the eukaryotic stress-response factor growth arrest and DNA damage 45α as a key player in the induction of the apoptotic process elicited by Shigella in epithelial cells, revealing an unexplored role of this molecule in the course of infections sustained by invasive pathogens.

  11. Knockout of a transgene by transcription activator-like effector nucleases (TALENs) in the sawfly, Athalia rosae (Hymenoptera) and the ladybird beetle, Harmonia axyridis (Coleoptera).

    PubMed

    Hatakeyama, M; Yatomi, J; Sumitani, M; Takasu, Y; Sekiné, K; Niimi, T; Sezutsu, H

    2016-02-01

    Transcription activator-like effector nucleases (TALENs) are efficient tools for targeted genome editing and have been utilized in a number of insects. Here, we demonstrate the gene disruption (knockout) caused by TALENs targeting a transgene, 3xP3-driven enhanced green fluorescence protein (EGFP), that is integrated in the genome of two species, the sawfly Athalia rosae (Hymenoptera) and the ladybird beetle Harmonia axyridis (Coleoptera). Messenger RNAs of TALENs targeting the sequences adjacent to the chromophore region were microinjected into the eggs/embryos of each species. In At. rosae, when microinjection was performed at the posterior end of eggs, 15% of G(0) individuals showed a somatic mosaic phenotype for eye EGFP fluorescence. Three-quarters of the somatic mosaics produced EGFP-negative G(1) progeny. When eggs were injected at the anterior end, 63% of the G(0) individuals showed somatic mosaicism, and 17% of them produced EGFP-negative G(1) progeny. In H. axyridis, 25% of posterior-injected and 8% of anterior-injected G(0) individuals produced EGFP-negative G(1) progeny. In both species, the EGFP-negative progeny retained the EGFP gene, and various deletions were detected in the target sequences, indicating that gene disruption was successfully induced. Finally, for both species, 18-21% of G(0) founders produced gene knockout progeny sufficient for establishing knockout strains.

  12. Visual short-term memory: activity supporting encoding and maintenance in retinotopic visual cortex.

    PubMed

    Sneve, Markus H; Alnæs, Dag; Endestad, Tor; Greenlee, Mark W; Magnussen, Svein

    2012-10-15

    Recent studies have demonstrated that retinotopic cortex maintains information about visual stimuli during retention intervals. However, the process by which transient stimulus-evoked sensory responses are transformed into enduring memory representations is unknown. Here, using fMRI and short-term visual memory tasks optimized for univariate and multivariate analysis approaches, we report differential involvement of human retinotopic areas during memory encoding of the low-level visual feature orientation. All visual areas show weaker responses when memory encoding processes are interrupted, possibly due to effects in orientation-sensitive primary visual cortex (V1) propagating across extrastriate areas. Furthermore, intermediate areas in both dorsal (V3a/b) and ventral (LO1/2) streams are significantly more active during memory encoding compared with non-memory (active and passive) processing of the same stimulus material. These effects in intermediate visual cortex are also observed during memory encoding of a different stimulus feature (spatial frequency), suggesting that these areas are involved in encoding processes on a higher level of representation. Using pattern-classification techniques to probe the representational content in visual cortex during delay periods, we further demonstrate that simply initiating memory encoding is not sufficient to produce long-lasting memory traces. Rather, active maintenance appears to underlie the observed memory-specific patterns of information in retinotopic cortex.

  13. Nck adaptors, besides promoting N-WASP mediated actin-nucleation activity at pedestals, influence the cellular levels of enteropathogenic Escherichia coli Tir effector.

    PubMed

    Nieto-Pelegrin, Elvira; Kenny, Brendan; Martinez-Quiles, Narcisa

    2014-01-01

    Enteropathogenic Escherichia coli (EPEC) binding to human intestinal cells triggers the formation of disease-associated actin rich structures called pedestals. The latter process requires the delivery, via a Type 3 secretion system, of the translocated Intimin receptor (Tir) protein into the host plasma membrane where binding of a host kinase-modified form to the bacterial surface protein Intimin triggers pedestal formation. Tir-Intimin interaction recruits the Nck adaptor to a Tir tyrosine phosphorylated residue where it activates neural Wiskott-Aldrich syndrome protein (N-WASP); initiating the major pathway to actin polymerization mediated by the actin-related protein (Arp) 2/3 complex. Previous studies with Nck-deficient mouse embryonic fibroblasts (MEFs) identified a key role for Nck in pedestal formation, presumed to reflect a lack of N-WASP activation. Here, we show the defect relates to reduced amounts of Tir within Nck-deficient cells. Indeed, Tir delivery and, thus, pedestal formation defects were much greater for MEFs than HeLa (human epithelial) cells. Crucially, the levels of two other effectors (EspB/EspF) within Nck-deficient MEFs were not reduced unlike that of Map (Mitochondrial associated protein) which, like Tir, requires CesT chaperone function for efficient delivery. Interestingly, drugs blocking various host protein degradation pathways failed to increase Tir cellular levels unlike an inhibitor of deacetylase activity (Trichostatin A; TSA). Treatments with TSA resulted in significant recovery of Tir levels, potentiation of actin polymerization and improvement in bacterial attachment to cells. Our findings have important implications for the current model of Tir-mediated actin polymerization and opens new lines of research in this area. PMID:25482634

  14. A Novel Two-Component Signaling System That Activates Transcription of an Enterohemorrhagic Escherichia coli Effector Involved in Remodeling of Host Actin▿

    PubMed Central

    Reading, Nicola C.; Torres, Alfredo G.; Kendall, Melissa M.; Hughes, David T.; Yamamoto, Kaneyoshi; Sperandio, Vanessa

    2007-01-01

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for worldwide outbreaks of bloody diarrhea, hemorrhagic colitis, and life-threatening hemolytic uremic syndrome. After colonizing the large intestine, EHEC forms attaching and effacing (AE) lesions on intestinal epithelial cells. These lesions cause destruction of the microvilli and elicit actin rearrangement to form pedestals that cup each bacterium individually. EHEC responds to a signal produced by the intestinal microbial flora, autoinducer-3 (AI-3), and the host hormones epinephrine and norepinephrine to activate transcription of the genes involved in AE lesion formation. These three signals, involved in interkingdom communication, are sensed by bacterial sensor kinases. Here we describe a novel two-component system, QseEF (quorum-sensing E. coli regulators E and F), which is part of the AI-3/epinephrine/norepinephrine signaling system. QseE is the sensor kinase and QseF the response regulator. The qseEF genes are cotranscribed, and transcription of qseEF is activated by epinephrine through the QseC sensor. A qseF mutant does not form AE lesions. QseF activates transcription of the gene encoding EspFu, an effector protein translocated to the host cell by the EHEC, which mimics a eukaryotic SH2/SH3 adapter protein to engender actin polymerization during pedestal formation. Expression of the espFu gene from a plasmid restored AE lesion formation to the qseF mutant, suggesting that lack of espFu expression in this mutant was responsible for the loss of pedestal formation. These findings suggest the QseEF is a two-component system involved in the regulation of AE lesion formation by EHEC. PMID:17220220

  15. Age differences in brain activity related to unsuccessful declarative memory retrieval.

    PubMed

    Grady, Cheryl L; St-Laurent, Marie; Burianová, Hana

    2015-07-01

    Although memory recall is known to be reduced with normal aging, little is known about the patterns of brain activity that accompany these recall failures. By assessing faulty memory, we can identify the brain regions engaged during retrieval attempts in the absence of successful memory and determine the impact of aging on this functional activity. We used functional magnetic resonance imaging to examine age differences in brain activity associated with memory failure in three memory retrieval tasks: autobiographical (AM), episodic (EM) and semantic (SM). Compared to successful memory retrieval, both age groups showed more activity when they failed to recall a memory in regions consistent with the salience network (SLN), a brain network also associated with non-memory errors. Both groups also showed strong functional coupling among SLN regions during incorrect trials and in intrinsic patterns of functional connectivity. In comparison to young adults, older adults demonstrated (1) less activity within the SLN during unsuccessful AM trials; (2) weaker intrinsic functional connectivity between SLN nodes and dorsolateral prefrontal cortex; and (3) less differentiation of SLN functional connectivity during incorrect trials across memory conditions. These results suggest that the SLN is engaged during recall failures, as it is for non-memory errors, which may be because errors in general have particular salience for adapting behavior. In older adults, the dedifferentiation of functional connectivity within the SLN across memory conditions and the reduction of functional coupling between it and prefrontal cortex may indicate poorer inter-network communication and less flexible use of cognitive control processes, either while retrieval is attempted or when monitoring takes place after retrieval has failed. This article is part of a Special Issue entitled SI: Memory & Aging.

  16. TAL effectors and the executor R genes

    PubMed Central

    Zhang, Junli; Yin, Zhongchao; White, Frank

    2015-01-01

    Transcription activator-like (TAL) effectors are bacterial type III secretion proteins that function as transcription factors in plants during Xanthomonas/plant interactions, conditioning either host susceptibility and/or host resistance. Three types of TAL effector associated resistance (R) genes have been characterized—recessive, dominant non-transcriptional, and dominant TAL effector-dependent transcriptional based resistance. Here, we discuss the last type of R genes, whose functions are dependent on direct TAL effector binding to discrete effector binding elements in the promoters. Only five of the so-called executor R genes have been cloned, and commonalities are not clear. We have placed the protein products in two groups for conceptual purposes. Group 1 consists solely of the protein from pepper, BS3, which is predicted to have catalytic function on the basis of homology to a large conserved protein family. Group 2 consists of BS4C-R, XA27, XA10, and XA23, all of which are relatively short proteins from pepper or rice with multiple potential transmembrane domains. Group 2 members have low sequence similarity to proteins of unknown function in closely related species. Firm predictions await further experimentation on these interesting new members to the R gene repertoire, which have potential broad application in new strategies for disease resistance. PMID:26347759

  17. Hippocampal Activation of Rac1 Regulates the Forgetting of Object Recognition Memory.

    PubMed

    Liu, Yunlong; Du, Shuwen; Lv, Li; Lei, Bo; Shi, Wei; Tang, Yikai; Wang, Lianzhang; Zhong, Yi

    2016-09-12

    Forgetting is a universal feature for most types of memories. The best-defined and extensively characterized behaviors that depict forgetting are natural memory decay and interference-based forgetting [1, 2]. Molecular mechanisms underlying the active forgetting remain to be determined for memories in vertebrates. Recent progress has begun to unravel such mechanisms underlying the active forgetting [3-11] that is induced through the behavior-dependent activation of intracellular signaling pathways. In Drosophila, training-induced activation of the small G protein Rac1 mediates natural memory decay and interference-based forgetting of aversive conditioning memory [3]. In mice, the activation of photoactivable-Rac1 in recently potentiated spines in a motor learning task erases the motor memory [12]. These lines of evidence prompted us to investigate a role for Rac1 in time-based natural memory decay and interference-based forgetting in mice. The inhibition of Rac1 activity in hippocampal neurons through targeted expression of a dominant-negative Rac1 form extended object recognition memory from less than 72 hr to over 72 hr, whereas Rac1 activation accelerated memory decay within 24 hr. Interference-induced forgetting of this memory was correlated with Rac1 activation and was completely blocked by inhibition of Rac1 activity. Electrophysiological recordings of long-term potentiation provided independent evidence that further supported a role for Rac1 activation in forgetting. Thus, Rac1-dependent forgetting is evolutionarily conserved from invertebrates to vertebrates.

  18. Hippocampal Activation of Rac1 Regulates the Forgetting of Object Recognition Memory.

    PubMed

    Liu, Yunlong; Du, Shuwen; Lv, Li; Lei, Bo; Shi, Wei; Tang, Yikai; Wang, Lianzhang; Zhong, Yi

    2016-09-12

    Forgetting is a universal feature for most types of memories. The best-defined and extensively characterized behaviors that depict forgetting are natural memory decay and interference-based forgetting [1, 2]. Molecular mechanisms underlying the active forgetting remain to be determined for memories in vertebrates. Recent progress has begun to unravel such mechanisms underlying the active forgetting [3-11] that is induced through the behavior-dependent activation of intracellular signaling pathways. In Drosophila, training-induced activation of the small G protein Rac1 mediates natural memory decay and interference-based forgetting of aversive conditioning memory [3]. In mice, the activation of photoactivable-Rac1 in recently potentiated spines in a motor learning task erases the motor memory [12]. These lines of evidence prompted us to investigate a role for Rac1 in time-based natural memory decay and interference-based forgetting in mice. The inhibition of Rac1 activity in hippocampal neurons through targeted expression of a dominant-negative Rac1 form extended object recognition memory from less than 72 hr to over 72 hr, whereas Rac1 activation accelerated memory decay within 24 hr. Interference-induced forgetting of this memory was correlated with Rac1 activation and was completely blocked by inhibition of Rac1 activity. Electrophysiological recordings of long-term potentiation provided independent evidence that further supported a role for Rac1 activation in forgetting. Thus, Rac1-dependent forgetting is evolutionarily conserved from invertebrates to vertebrates. PMID:27593377

  19. Hippocampal activation during retrieval of spatial context from episodic and semantic memory.

    PubMed

    Hoscheidt, Siobhan M; Nadel, Lynn; Payne, Jessica; Ryan, Lee

    2010-10-15

    The hippocampus, a region implicated in the processing of spatial information and episodic memory, is central to the debate concerning the relationship between episodic and semantic memory. Studies of medial temporal lobe amnesic patients provide evidence that the hippocampus is critical for the retrieval of episodic but not semantic memory. On the other hand, recent neuroimaging studies of intact individuals report hippocampal activation during retrieval of both autobiographical memories and semantic information that includes historical facts, famous faces, and categorical information, suggesting that episodic and semantic memory may engage the hippocampus during memory retrieval in similar ways. Few studies have matched episodic and semantic tasks for the degree to which they include spatial content, even though spatial content may be what drives hippocampal activation during semantic retrieval. To examine this issue, we conducted a functional magnetic resonance imaging (fMRI) study in which retrieval of spatial and nonspatial information was compared during an episodic and semantic recognition task. Results show that the hippocampus (1) participates preferentially in the retrieval of episodic memories; (2) is also engaged by retrieval of semantic memories, particularly those that include spatial information. These data suggest that sharp dissociations between episodic and semantic memory may be overly simplistic and that the hippocampus plays a role in the retrieval of spatial content whether drawn from a memory of one's own life experiences or real-world semantic knowledge. PMID:20385169

  20. Working Memory-Related Neural Activity Predicts Future Smoking Relapse

    PubMed Central

    Loughead, James; Wileyto, E Paul; Ruparel, Kosha; Falcone, Mary; Hopson, Ryan; Gur, Ruben; Lerman, Caryn

    2015-01-01

    Brief abstinence from smoking impairs cognition, particularly executive function, and this has a role in relapse to smoking. This study examined whether working memory-related brain activity predicts subsequent smoking relapse above and beyond standard clinical and behavioral measures. Eighty treatment-seeking smokers completed two functional magnetic resonance imaging sessions (smoking satiety vs 24 h abstinence challenge) during performance of a visual N-back task. Brief counseling and a short-term quit attempt followed. Relapse during the first 7 days was biochemically confirmed by the presence of the nicotine metabolite cotinine. Mean percent blood oxygen level-dependent (BOLD) signal change was extracted from a priori regions of interest: bilateral dorsolateral prefrontal cortex (DLPFC), medial frontal/cingulate gyrus, posterior cingulate cortex (PCC), and ventromedial prefrontal cortex. Signal from these brain regions and additional clinical measures were used to model outcome status, which was then validated with resampling techniques. Relapse to smoking was predicted by increased withdrawal symptoms, decreased left DLPFC and increased PCC BOLD percent signal change (abstinence vs smoking satiety). Receiver operating characteristic analysis demonstrated 81% area under the curve using these predictors, a significant improvement over the model with clinical variables only. The combination of abstinence-induced decreases in left DLPFC activation and reduced suppression of PCC may be a prognostic marker for poor outcome, specifically early smoking relapse. PMID:25469682

  1. Rho GTPases and their effector proteins.

    PubMed Central

    Bishop, A L; Hall, A

    2000-01-01

    Rho GTPases are molecular switches that regulate many essential cellular processes, including actin dynamics, gene transcription, cell-cycle progression and cell adhesion. About 30 potential effector proteins have been identified that interact with members of the Rho family, but it is still unclear which of these are responsible for the diverse biological effects of Rho GTPases. This review will discuss how Rho GTPases physically interact with, and regulate the activity of, multiple effector proteins and how specific effector proteins contribute to cellular responses. To date most progress has been made in the cytoskeleton field, and several biochemical links have now been established between GTPases and the assembly of filamentous actin. The main focus of this review will be Rho, Rac and Cdc42, the three best characterized mammalian Rho GTPases, though the genetic analysis of Rho GTPases in lower eukaryotes is making increasingly important contributions to this field. PMID:10816416

  2. Silymarin inhibits ultraviolet radiation-induced immune suppression through DNA repair-dependent activation of dendritic cells and stimulation of effector T cells.

    PubMed

    Vaid, Mudit; Prasad, Ram; Singh, Tripti; Elmets, Craig A; Xu, Hui; Katiyar, Santosh K

    2013-04-15

    Silymarin inhibits UVB-induced immunosuppression in mouse skin. To identify the molecular mechanisms underlying this effect, we used an adoptive transfer approach in which dendritic cells (DCs) from the draining lymph nodes of donor mice that had been UVB-exposed and sensitized to 2,4,-dinitrofluorobenzene (DNFB) were transferred into naïve recipient mice. The contact hypersensitivity (CHS) response of the recipient mice to DNFB was then measured. When DCs were obtained from UVB-exposed donor mice that were not treated with silymarin, the CHS response was suppressed confirming the role of DCs in the UVB-induced immunosuppression. Silymarin treatment of UVB-exposed donor mice relieved this suppression of the CHS response in the recipients. Silymarin treatment was associated with rapid repair of UVB-induced cyclobutane pyrimidine dimers (CPDs) in DCs and silymarin treatment did not prevent UV-induced immunosuppression in XPA-deficient mice which are unable to repair UV-induced DNA damage. The CHS response in mice receiving DCs from silymarin-treated UV-exposed donor mice also was associated with enhanced secretion of Th1-type cytokines and stimulation of T cells. Adoptive transfer of T cells revealed that transfer of either CD8(+) or CD4(+) cells from silymarin-treated, UVB-exposed donors resulted in enhancement of the CHS response. Cell culture study showed enhanced secretion of IL-2 and IFNγ by CD8(+) T cells, and reduced secretion of Th2 cytokines by CD4(+) T cells, obtained from silymarin-treated UVB-exposed mice. These data suggest that DNA repair-dependent functional activation of DCs, a reduction in CD4(+) regulatory T-cell activity, and stimulation of CD8(+) effector T cells contribute to silymarin-mediated inhibition of UVB-induced immunosuppression. PMID:23395695

  3. Insights into the TOR-S6K signaling pathway in maize (Zea mays L.). pathway activation by effector-receptor interaction.

    PubMed

    Garrocho-Villegas, Verónica; Aguilar C, Raúl; Sánchez de Jiménez, Estela

    2013-12-23

    The primordial TOR pathway, known to control growth and cell proliferation, has still not been fully described for plants. Nevertheless, in maize, an insulin-like growth factor (ZmIGF) peptide has been reported to stimulate this pathway. This research provides further insight into the TOR pathway in maize, using a biochemical approach in cultures of fast-growing (FG) and slow-growing (SG) calli, as a model system. Our results revealed that addition of either ZmIGF or insulin to SG calli stimulated DNA synthesis and increased the growth rate through cell proliferation and increased the rate of ribosomal protein (RP) synthesis by the selective mobilization of RP mRNAs into polysomes. Furthermore, analysis of the phosphorylation status of the main TOR and S6K kinases from the TOR pathway revealed stimulation by ZmIGF or insulin, whereas rapamycin inhibited its activation. Remarkably, a putative maize insulin-like receptor was recognized by a human insulin receptor antibody, as demonstrated by immunoprecipitation from membrane protein extracts of maize callus. Furthermore, competition experiments between ZmIGF and insulin for the receptor site on maize protoplasts suggested structural recognition of the putative receptor by either effector. These data were confirmed by confocal immunolocalization within the cell membrane of callus cells. Taken together, these data indicate that cell growth and cell proliferation in maize depend on the activation of the TOR-S6K pathway through the interaction of an insulin-like growth factor and its receptor. This evidence suggests that higher plants as well as metazoans have conserved this biochemical pathway to regulate their growth, supporting the conclusion that it is a highly evolved conserved pathway.

  4. Hippocampal noradrenergic activation is necessary for object recognition memory consolidation and can promote BDNF increase and memory persistence.

    PubMed

    Mello-Carpes, Pâmela B; da Silva de Vargas, Liane; Gayer, Mateus Cristofari; Roehrs, Rafael; Izquierdo, Ivan

    2016-01-01

    Previously we showed that activation of the Nucleus of the Solitary Tract (NTS)-Nucleus Paragigantocellularis (PGi)-Locus coeruleus (LC) pathway, which theoretically culminates with norepinephrine (NE) release in dorsal hippocampus (CA1 region) and basolateral amygdala (BLA) is necessary for the consolidation of object recognition (OR) memory. Here we show that, while the microinjection of the beta-noradrenergic receptor blocker timolol into CA1 impairs OR memory consolidation, the microinjection of norepinephrine (NE) promotes the persistence of this type of memory. Further, we show that OR consolidation is attended by an increase of norepinephrine (NE) levels and of the expression of brain derived neurotrophic factor (BDNF) in hippocampus, which are impaired by inactivation of the NTS-PGi-LC pathway by the infusion of muscimol into the NTS. PMID:26691781

  5. Hippocampal noradrenergic activation is necessary for object recognition memory consolidation and can promote BDNF increase and memory persistence.

    PubMed

    Mello-Carpes, Pâmela B; da Silva de Vargas, Liane; Gayer, Mateus Cristofari; Roehrs, Rafael; Izquierdo, Ivan

    2016-01-01

    Previously we showed that activation of the Nucleus of the Solitary Tract (NTS)-Nucleus Paragigantocellularis (PGi)-Locus coeruleus (LC) pathway, which theoretically culminates with norepinephrine (NE) release in dorsal hippocampus (CA1 region) and basolateral amygdala (BLA) is necessary for the consolidation of object recognition (OR) memory. Here we show that, while the microinjection of the beta-noradrenergic receptor blocker timolol into CA1 impairs OR memory consolidation, the microinjection of norepinephrine (NE) promotes the persistence of this type of memory. Further, we show that OR consolidation is attended by an increase of norepinephrine (NE) levels and of the expression of brain derived neurotrophic factor (BDNF) in hippocampus, which are impaired by inactivation of the NTS-PGi-LC pathway by the infusion of muscimol into the NTS.

  6. CD69-mediated pathway of lymphocyte activation: anti-CD69 monoclonal antibodies trigger the cytolytic activity of different lymphoid effector cells with the exception of cytolytic T lymphocytes expressing T cell receptor alpha/beta

    PubMed Central

    1991-01-01

    The effect of anti-CD69 monoclonal antibodies (mAbs) on the induction of the cytolytic activity in different types of lymphoid effector cells has been investigated. Three anti-CD69 mAbs, including the reference mAb MLR3 and two new mAbs (c227 and 31C4), have been used. All cloned CD3-CD16+ natural killer (NK) cells belonging to different subsets (as defined by the surface expression of GL183 and/or EB6 antigens) were efficiently triggered by anti-CD69 mAbs and lysed P815 mastocytoma cells in a redirected killing assay. Triggering of the cytolytic activity could also be induced in CD3-CD16- NK clones, which fail to respond to other stimuli (including anti-CD16, anti-CD2 mAbs, or phytohemagglutinin). A similar triggering effect was detected in T cell receptor (TCR) gamma/delta+ clones belonging to different subsets. On the other hand, anti-CD69 mAbs could not induce triggering of the cytolytic activity in TCR alpha/beta+ cytolytic clones. Since all thymocytes are known to express CD69 antigen after cell activation, we analyzed a series of phenotypically different cytolytic thymocyte populations and clones for their responsiveness to anti-CD69 mAb in a redirected killing assay. Again, anti-CD69 mAb triggered TCR gamma/delta+ but not TCR alpha/beta+ thymocytes. Anti-CD69 mAb efficiently triggered the cytolytic activity of "early" thymocytes lines or clones (CD3-4-8-7+), which lack all other known pathways of cell activation. Thus, it appears that CD69 molecules may initiate a pathway of activation of cytolytic functions common to a number of activated effector lymphocytes with the remarkable exception of TCR alpha/beta+ cytolytic cells. PMID:1720808

  7. Virus-induced polyclonal B cell activation improves protective CTL memory via retained CD27 expression on memory CTL.

    PubMed

    Matter, Matthias; Mumprecht, Sabine; Pinschewer, Daniel D; Pavelic, Viktor; Yagita, Hideo; Krautwald, Stefan; Borst, Jannie; Ochsenbein, Adrian F

    2005-11-01

    Different viruses elicit distinct phenotypes of memory cytotoxic T lymphocytes (CTL). This is reflected in differential expression of homing receptors and costimulatory molecules like CD27. Memory CTL retained CD27 following lymphocytic choriomeningitis virus (LCMV) infection, but not after immunization with recombinant vaccinia virus or tumor cells expressing LCMV glycoprotein. Stable CD27 expression on memory CTL required ligation by CD70 expressed on polyclonally activated B cells during the contraction phase. The functional consequence of CD27 expressed on virus-specific CTL was analyzed in CD27-deficient mice. LCMV infection of CD27(-/-) mice revealed that primary CTL activation and expansion as well as elimination of the virus were independent of CD27 expression. In contrast, ligation of CD27 on memory CTL upon secondary antigen encounter increased clonal expansion and improved protection against re-infection. This points to novel B cell-CTL interactions during viral infection and to a beneficial role of polyclonal B cell activation that represents a characteristic of murine LCMV, human immunodeficiency virus and human hepatitis B and C virus infection. PMID:16231287

  8. The hippocampus remains activated over the long term for the retrieval of truly episodic memories.

    PubMed

    Harand, Caroline; Bertran, Françoise; La Joie, Renaud; Landeau, Brigitte; Mézenge, Florence; Desgranges, Béatrice; Peigneux, Philippe; Eustache, Francis; Rauchs, Géraldine

    2012-01-01

    The role of the hippocampus in declarative memory consolidation is a matter of intense debate. We investigated the neural substrates of memory retrieval for recent and remote information using functional magnetic resonance imaging (fMRI). 18 young, healthy participants learned a series of pictures. Then, during two fMRI recognition sessions, 3 days and 3 months later, they had to determine whether they recognized or not each picture using the "Remember/Know" procedure. Presentation of the same learned images at both delays allowed us to track the evolution of memories and distinguish consistently episodic memories from those that were initially episodic and then became familiar or semantic over time and were retrieved without any contextual detail. Hippocampal activation decreased over time for initially episodic, later semantic memories, but remained stable for consistently episodic ones, at least in its posterior part. For both types of memories, neocortical activations were observed at both delays, notably in the ventromedial prefrontal and anterior cingulate cortices. These activations may reflect a gradual reorganization of memory traces within neural networks. Our data indicate maintenance and strengthening of hippocampal and cortico-cortical connections in the consolidation and retrieval of episodic memories over time, in line with the Multiple Trace theory (Nadel and Moscovitch, 1997). At variance, memories becoming semantic over time consolidate through strengthening of cortico-cortical connections and progressive disengagement of the hippocampus. PMID:22937055

  9. EFFECTOR CELL BLOCKADE

    PubMed Central

    Schrader, John W.; Nossal, G. J. V.

    1974-01-01

    This study describes the effects of incubating antibody-forming cells (AFC), either as mass cell suspensions, or as single AFC in microdroplets, with antigens against which the cells display specificity. Most of the work was done with hapten-specific anti-DNP-AFC, but AFC with specificity against flagellar antigens or fowl gamma globulin (FGG) were also included. It was noted that 30-min incubation of AFC with highly multivalent forms of antigen caused a substantial partial suppression of the antibody-forming performance of the AFC as measured by a hemolytic plaque test. Thus, when cell suspensions containing anti-DNP plaque-forming cells (PFC), were incubated for 30 min at 37°C with 100 µg of DNP-polymerized flagellin (DNP-POL), the number of plaques appearing after washing of the cells and placing them in plaque-revealing erythrocyte monolayers was reduced to 50% or less compared with the number of plaques observed with control portions preincubated with medium alone. Preincubation with DNP-lysine, with oligovalent DNP-protein conjugates, or with irrelevant antigens produced no such inhibition. Studies where preinhibited PFC suspensions were mixed with control suspensions before assay showed that a nonspecific carryover of antigen into the assay system was not involved. The inhibitory effect could also be initiated by holding cells at 0°C with DNP-POL, but in that case, inhibition only became manifest after cells were incubated for 30 min at 37°C before being placed in plaque-revealing monolayers. This suggested that inhibition was initiated by adsorption of multivalent antigen onto PFC-surface Ig, but required some active process before secretion actually slowed down. The effect was dose- and time-dependent, antigen-specific, and generalized for all antigens studied. As well as yielding reduced plaque numbers, the preinhibited cells also gave smaller, more turbid plaques, suggesting a reduction in antibody-forming rate by each PFC rather than the elimination

  10. Lateral posterior parietal activity during reality monitoring discriminations of memories of high and low perceptual vividness.

    PubMed

    King, Danielle R; Schubert, Misty L; Miller, Michael B

    2015-09-01

    Regions of the lateral posterior parietal cortex (PPC) tend to be more active during recognition of previously studied items compared to correct rejection of unstudied items. Previously, we demonstrated that this effect is source-specific. While items that were encoded through visual perception elicited robust successful retrieval activity in the lateral PPC during a subsequent source memory test, items that were visually imagined did not elicit this effect. Memories of perceived events typically contain more perceptually-based contextual details than memories of imagined events. Therefore, source-based differences in lateral parietal activity might be due to a difference in the perceptual vividness of memories of perceived and imagined events. The goal of the present study was to test this hypothesis. Participants perceived and imagined items in both high and low perceptual vividness conditions. Experiment 1 demonstrated that memories for items encoded in the high vividness conditions contained significantly greater visual detail than memories encoded in the low vividness conditions. In Experiment 2, participants were scanned while they made source memory judgments about items that were previously perceived and imagined in high and low vividness conditions. Consistent with previous findings, the left lateral PPC was more active during retrieval of perceived compared to imagined events. However, lateral PPC activity did not vary according to vividness, suggesting that source effects in this region cannot be explained by a difference in the perceptual vividness of memories encoded through perception versus imagination.

  11. Age Differences in Brain Activity Related to Unsuccessful Declarative Memory Retrieval

    PubMed Central

    Grady, Cheryl L.; St-Laurent, Marie; Burianová, Hana

    2016-01-01

    Although memory recall is known to be reduced with normal aging, little is known about the patterns of brain activity that accompany these recall failures. By assessing faulty memory, we can identify the brain regions engaged during retrieval attempts in the absence of successful memory and determine the impact of aging on this functional activity. We used functional magnetic resonance imaging to examine age differences in brain activity associated with memory failure in three memory retrieval tasks: autobiographical (AM), episodic (EM) and semantic (SM). Compared to successful memory retrieval, both age groups showed more activity when they failed to recall a memory in regions consistent with the salience network (SLN), a brain network also associated with non-memory errors. Both groups also showed strong functional coupling among SLN regions during incorrect trials and in intrinsic patterns of functional connectivity. In comparison to young adults, older adults demonstrated (1) less activity within the SLN during unsuccessful AM trials; (2) weaker intrinsic functional connectivity between SLN nodes and dorsolateral prefrontal cortex; and (3) less differentiation of SLN functional connectivity during incorrect trials across memory conditions. These results suggest that the SLN is engaged during recall failures, as it is for non-memory errors, which may be because errors in general have particular salience for adapting behavior. In older adults, the dedifferentiation of functional connectivity within the SLN across memory conditions and the reduction of functional coupling between it and prefrontal cortex may indicate poorer internetwork communication and less flexible use of cognitive control processes, either while retrieval is attempted or when monitoring takes place after retrieval has failed. PMID:25541365

  12. Brain region-specific activity patterns after recent or remote memory retrieval of auditory conditioned fear.

    PubMed

    Kwon, Jeong-Tae; Jhang, Jinho; Kim, Hyung-Su; Lee, Sujin; Han, Jin-Hee

    2012-01-01

    Memory is thought to be sparsely encoded throughout multiple brain regions forming unique memory trace. Although evidence has established that the amygdala is a key brain site for memory storage and retrieval of auditory conditioned fear memory, it remains elusive whether the auditory brain regions may be involved in fear memory storage or retrieval. To investigate this possibility, we systematically imaged the brain activity patterns in the lateral amygdala, MGm/PIN, and AuV/TeA using activity-dependent induction of immediate early gene zif268 after recent and remote memory retrieval of auditory conditioned fear. Consistent with the critical role of the amygdala in fear memory, the zif268 activity in the lateral amygdala was significantly increased after both recent and remote memory retrieval. Interesting, however, the density of zif268 (+) neurons in both MGm/PIN and AuV/TeA, particularly in layers IV and VI, was increased only after remote but not recent fear memory retrieval compared to control groups. Further analysis of zif268 signals in AuV/TeA revealed that conditioned tone induced stronger zif268 induction compared to familiar tone in each individual zif268 (+) neuron after recent memory retrieval. Taken together, our results support that the lateral amygdala is a key brain site for permanent fear memory storage and suggest that MGm/PIN and AuV/TeA might play a role for remote memory storage or retrieval of auditory conditioned fear, or, alternatively, that these auditory brain regions might have a different way of processing for familiar or conditioned tone information at recent and remote time phases. PMID:22993170

  13. Memory retrieval by activating engram cells in mouse models of early Alzheimer’s disease

    PubMed Central

    Roy, Dheeraj S.; Arons, Autumn; Mitchell, Teryn I.; Pignatelli, Michele; Ryan, Tomás J.; Tonegawa, Susumu

    2016-01-01

    Summary Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by progressive memory decline and subsequent loss of broader cognitive functions1. Memory decline in early stages of Alzheimer’s is mostly limited to episodic memory, for which the hippocampus (HPC) plays a crucial role2. However, it has been uncertain whether the observed amnesia in early stages of Alzheimer’s is due to disrupted encoding and consolidation of episodic information, or an impairment in the retrieval of stored memory information. Here we show that in transgenic mouse models of early Alzheimer’s, direct optogenetic activation of hippocampal memory engram cells results in memory retrieval despite the fact that these mice are amnesic in long-term memory tests when natural recall cues are utilized, revealing a retrieval, rather than a storage impairment. Prior to amyloid plaque deposition, the amnesia in these mice is age-dependent3–5, which correlates with a progressive reduction of spine density of hippocampal dentate gyrus (DG) engram cells. We show that optogenetic induction of long-term potentiation (LTP) at perforant path (PP) synapses of DG engram cells restores both spine density and long-term memory. We also demonstrate that an ablation of DG engram cells containing restored spine density prevents the rescue of long-term memory. Thus, selective rescue of spine density in engram cells may lead to an effective strategy for treating memory loss in early stages of Alzheimer’s disease. PMID:26982728

  14. Memory retrieval by activating engram cells in mouse models of early Alzheimer's disease.

    PubMed

    Roy, Dheeraj S; Arons, Autumn; Mitchell, Teryn I; Pignatelli, Michele; Ryan, Tomás J; Tonegawa, Susumu

    2016-03-24

    Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive memory decline and subsequent loss of broader cognitive functions. Memory decline in the early stages of AD is mostly limited to episodic memory, for which the hippocampus has a crucial role. However, it has been uncertain whether the observed amnesia in the early stages of AD is due to disrupted encoding and consolidation of episodic information, or an impairment in the retrieval of stored memory information. Here we show that in transgenic mouse models of early AD, direct optogenetic activation of hippocampal memory engram cells results in memory retrieval despite the fact that these mice are amnesic in long-term memory tests when natural recall cues are used, revealing a retrieval, rather than a storage impairment. Before amyloid plaque deposition, the amnesia in these mice is age-dependent, which correlates with a progressive reduction in spine density of hippocampal dentate gyrus engram cells. We show that optogenetic induction of long-term potentiation at perforant path synapses of dentate gyrus engram cells restores both spine density and long-term memory. We also demonstrate that an ablation of dentate gyrus engram cells containing restored spine density prevents the rescue of long-term memory. Thus, selective rescue of spine density in engram cells may lead to an effective strategy for treating memory loss in the early stages of AD. PMID:26982728

  15. Shape memory alloy-based active chiral composite cells

    NASA Astrophysics Data System (ADS)

    Prajapati, Maulik; Roy Mahapatra, D.

    2014-04-01

    Wing morphing is one of the emerging methodology towards improving aerodynamic efficiency of flight vehicle structures. In this paper a morphing structural element is designed and studied which has its origin in the well known chiral structures. The new aspect of design and functionality explored in this paper is that the chiral cell is actuated using thermal Shape Memory Alloy (SMA) actuator wires to provide directional motion. Such structure utilizes the potential of different actuations concepts based on actuator embedded in the chiral structure skin. This paper describes a new class of chiral cell structure with integrated SMA wire for actuation. Chiral topological constructs are obtained by considering passive and active load path decoupling and sub-optimal shape changes. Single cell of chiral honeycomb with actuators are analyzed using finite element simulation results and experiments. To this end, a multi-cell plan-form is characterized showing interesting possibilities in structural morphing applications. The applicability of the developed chiral cell to flexible wing skin, variable stiffness based design and controlling longitudinal-to-transverse stiffness ratio are discussed.

  16. SHADE: A Shape-Memory-Activated Device Promoting Ankle Dorsiflexion

    NASA Astrophysics Data System (ADS)

    Pittaccio, S.; Viscuso, S.; Rossini, M.; Magoni, L.; Pirovano, S.; Villa, E.; Besseghini, S.; Molteni, F.

    2009-08-01

    Acute post-stroke rehabilitation protocols include passive mobilization as a means to prevent contractures. A device (SHADE) that provides repetitive passive motion to a flaccid ankle by using shape memory alloy actuators could be of great help in providing this treatment. A suitable actuator was designed as a cartridge of approximately 150 × 20 × 15 mm, containing 2.5 m of 0.25 mm diameter NiTi wire. This actuator was activated by Joule’s effect employing a 7 s current input at 0.7 A, which provided 10 N through 76 mm displacement. Cooling and reset by natural convection took 30 s. A prototype of SHADE was assembled with two thermoplastic shells hinged together at the ankle and strapped on the shin and foot. Two actuators were fixed on the upper shell while an inextensible thread connected each NiTi wire to the foot shell. The passive ankle motion (passive range of motion, PROM) generated by SHADE was evaluated optoelectronically on three flaccid patients (58 ± 5 years old); acceptability was assessed by a questionnaire presented to further three flaccid patients (44 ± 11.5 years old) who used SHADE for 5 days, 30 min a day. SHADE was well accepted by all patients, produced good PROM, and caused no pain. The results prove that suitable limb mobilization can be produced by SMA actuators.

  17. Spatial Patterns of Persistent Neural Activity Vary with the Behavioral Context of Short-Term Memory

    PubMed Central

    Daie, Kayvon

    2015-01-01

    Summary A short-term memory can be evoked by different inputs and control separate targets in different behavioral contexts. To address the circuit mechanisms underlying context-dependent memory function, we determined through optical imaging how memory is encoded at the whole-network level in two behavioral settings. Persistent neural activity maintaining a memory of desired eye position was imaged throughout the oculomotor integrator after saccadic or optokinetic stimulation. While eye position was encoded by the amplitude of network activity, the spatial patterns of firing were context-dependent: cells located caudally generally were most persistent following saccadic input, whereas cells located rostrally were most persistent following optokinetic input. To explain these data, we computationally identified four independent modes of network activity and found these were differentially accessed by saccadic and optokinetic inputs. These results show how a circuit can simultaneously encode memory value and behavioral context, respectively, in its amplitude and spatial pattern of persistent firing. PMID:25661184

  18. Rab27a regulates epithelial sodium channel (ENaC) activity through synaptotagmin-like protein (SLP-5) and Munc13-4 effector mechanism

    SciTech Connect

    Saxena, Sunil K. . E-mail: ssaxena@stevens.edu; Horiuchi, Hisanori; Fukuda, Mitsunori

    2006-06-02

    Liddle's syndrome (excessive absorption of sodium ions) and PHA-1 (pseudohypoaldosteronism type 1) with decreased sodium absorption are caused by the mutations in the amiloride-sensitive epithelial sodium channel ENaC. Rab proteins are small GTPases involved in vesicle transport, docking, and fusion. Earlier, we reported that Rab27a inhibits ENaC-mediated currents through protein-protein interaction in HT-29 cells. We hereby report that Rab27a-dependent inhibition is associated with the GTP/GDP status as constitutively active or GTPase-deficient mutant Q78L inhibits amiloride-sensitive currents whereas GDP-locked inactive mutant T23N showed no effect. In order to further explore the molecular mechanism of this regulation, we performed competitive assays with two Rab27a-binding proteins: synaptotagmin-like protein (SLP-5) and Munc13-4 (a putative priming factor for exocytosis). Both proteins eliminate negative modulation of Rab27a on ENaC function. The SLP-5 reversal of Rab27a effect was restricted to C-terminal C2A/C2B domains assigned for putative phospholipids-binding function while the Rab27a-binding SHD motif imparted higher inhibition. The ENaC-mediated currents remain unaffected by Rab27a though SLP-5 appears to strongly bind it. The immunoprecipitation experiments suggest that in the presence of excessive Munc13-4 and SLP-5 proteins, Rab27a interaction with ENaC is diminished. Munc13-4 and SLP-5 limit the Rab27a availability to ENaC, thus minimizing its effect on channel function. These observations decisively prove that Rab27a inhibits ENaC function through a complex mechanism that involves GTP/GDP status, and protein-protein interactions involving Munc13-4 and SLP-5 effector proteins.

  19. Targeting of the Plzf Gene in the Rat by Transcription Activator-Like Effector Nuclease Results in Caudal Regression Syndrome in Spontaneously Hypertensive Rats

    PubMed Central

    Liška, František; Peterková, Renata; Peterka, Miroslav; Landa, Vladimír; Zídek, Václav; Mlejnek, Petr; Šilhavý, Jan; Šimáková, Miroslava; Křen, Vladimír; Starker, Colby G.; Voytas, Daniel F.; Izsvák, Zsuzsanna; Pravenec, Michal

    2016-01-01

    Recently, it has been found that spontaneous mutation Lx (polydactyly-luxate syndrome) in the rat is determined by deletion of a conserved intronic sequence of the Plzf (Promyelocytic leukemia zinc finger protein) gene. In addition, Plzf is a prominent candidate gene for quantitative trait loci (QTLs) associated with cardiac hypertrophy and fibrosis in the spontaneously hypertensive rat (SHR). In the current study, we tested the effects of Plzf gene targeting in the SHR using TALENs (transcription activator-like effector nucleases). SHR ova were microinjected with constructs pTAL438/439 coding for a sequence-specific endonuclease that binds to target sequence in the first coding exon of the Plzf gene. Out of 43 animals born after microinjection, we detected a single male founder. Sequence analysis revealed a deletion of G that resulted in frame shift mutation starting in codon 31 and causing a premature stop codon at position of amino acid 58. The Plzftm1Ipcv allele is semi-lethal since approximately 95% of newborn homozygous animals died perinatally. All homozygous animals exhibited manifestations of a caudal regression syndrome including tail anomalies and serious size reduction and deformities of long bones, and oligo- or polydactyly on the hindlimbs. The heterozygous animals only exhibited the tail anomalies. Impaired development of the urinary tract was also revealed: one homozygous and one heterozygous rat exhibited a vesico-ureteric reflux with enormous dilatation of ureters and renal pelvis. In the homozygote, this was combined with a hypoplastic kidney. These results provide evidence for the important role of Plzf gene during development of the caudal part of a body—column vertebrae, hindlimbs and urinary system in the rat. PMID:27727328

  20. Effect of activated antigen-specific B cells on ES-62-mediated modulation of effector function of heterologous antigen-specific T cells in vivo

    PubMed Central

    Marshall, Fraser A; Watson, Katherine A; Garside, Paul; Harnett, Margaret M; Harnett, William

    2008-01-01

    There is currently great interest in the idea of using helminth-derived molecules for therapeutic purposes and indeed we have shown that ES-62, a filarial nematode-derived phosphorylcholine-containing glycoprotein, significantly reduces the severity of arthritis in a murine model. Clearly, knowledge of mechanism of action is important when considering molecules for use in treating disease and although much is known regarding how ES-62 interacts with the immune system, gaps in our understanding remain. A feature of filarial nematode infection is a defective, T helper 2 (Th2)-polarized antigen-specific T-cell response and in relation to this we have recently shown that ES-62 inhibits clonal expansion and modulates effector function towards a Th2 phenotype, of antigen-specific T cells in vivo. ES-62 is also known to directly modulate B-cell behaviour and hence to determine whether it was mediating these effects on T cells by disrupting B–T-cell co-operation, we have investigated antigen-specific responses using an adoptive transfer system in which traceable numbers of tg ovalbumin (OVA)-specific T cells and hen egg lysozyme (HEL)-specific B cells respond to a chemically coupled form of OVA–HEL that contains linked epitopes that promote cognate T- and B-cell interactions. Surprisingly, these studies indicate that activated B cells restore T-cell expansion and prevent Th2-like polarization. However, ES-62-treated double cell transfer mice demonstrate a more generalized immunosuppression with reduced levels of Th1 and -2 type cytokines and antibody subclasses. Collectively, these results suggest that whilst ES-62 can target B–T-cell co-operation, this does not promote polarizing of T-cell responses towards a Th2-type phenotype. PMID:17961164

  1. Default Mode Network Activity Predicts Early Memory Decline in Healthy Young Adults Aged 18-31.

    PubMed

    Nelson, Steven M; Savalia, Neil K; Fishell, Andrew K; Gilmore, Adrian W; Zou, Fan; Balota, David A; McDermott, Kathleen B

    2016-08-01

    Functional magnetic resonance imaging (fMRI) research conducted in healthy young adults is typically done with the assumption that this sample is largely homogeneous. However, studies from cognitive psychology suggest that long-term memory and attentional control begin to diminish in the third decade of life. Here, 100 participants between the ages of 18 and 31 learned Lithuanian translations of English words in an individual differences study using fMRI. Long-term memory ability was operationalized for each participant by deriving a memory score from 3 convergent measures. Age of participant predicted memory score in this cohort. In addition, degree of deactivation during initial encoding in a set of regions occurring largely in the default mode network (DMN) predicted both age and memory score. The current study demonstrates that early memory decline may partially be accounted for by failure to modulate activity in the DMN.

  2. Two-axis angular effector

    DOEpatents

    Vaughn, Mark R.; Robinett, III, Rush D.; Phelan, John R.; Van Zuiden, Don M.

    1997-01-21

    A new class of coplanar two-axis angular effectors. These effectors combine a two-axis rotational joint analogous to a Cardan joint with linear actuators in a manner to produce a wider range of rotational motion about both axes defined by the joint. This new class of effectors also allows design of robotic manipulators having very high strength and efficiency. These effectors are particularly suited for remote operation in unknown surroundings, because of their extraordinary versatility. An immediate application is to the problems which arise in nuclear waste remediation.

  3. Mapping Neuronal Activation and the Influence of Adrenergic Signaling during Contextual Memory Retrieval

    ERIC Educational Resources Information Center

    Zhang, Wei-Ping; Guzowski, John F.; Thomas, Steven A.

    2005-01-01

    We recently described a critical role for adrenergic signaling in the hippocampus during contextual and spatial memory retrieval. To determine which neurons are activated by contextual memory retrieval and its sequelae in the presence and absence of adrenergic signaling, transcriptional imaging for the immediate-early gene "Arc" was used in…

  4. Memory Activation and the Availability of Explanations in Sequential Diagnostic Reasoning

    ERIC Educational Resources Information Center

    Mehlhorn, Katja; Taatgen, Niels A.; Lebiere, Christian; Krems, Josef F.

    2011-01-01

    In the field of diagnostic reasoning, it has been argued that memory activation can provide the reasoner with a subset of possible explanations from memory that are highly adaptive for the task at hand. However, few studies have experimentally tested this assumption. Even less empirical and theoretical work has investigated how newly incoming…

  5. The Effects of an Afterschool Physical Activity Program on Working Memory in Preadolescent Children

    ERIC Educational Resources Information Center

    Kamijo, Keita; Pontifex, Matthew B.; O'Leary, Kevin C.; Scudder, Mark R.; Wu, Chien-Ting; Castelli, Darla M.; Hillman, Charles H.

    2011-01-01

    The present study examined the effects of a 9-month randomized control physical activity intervention aimed at improving cardiorespiratory fitness on changes in working memory performance in preadolescent children relative to a waitlist control group. Participants performed a modified Sternberg task, which manipulated working memory demands based…

  6. Activation of Midbrain Structures by Associative Novelty and the Formation of Explicit Memory in Humans

    ERIC Educational Resources Information Center

    Schott, Bjorn H.; Sellner, Daniela B.; Lauer, Corinna-J.; Habib, Reza; Frey, Julietta U.; Guderian, Sebastian; Heinze, Hans-Jochen; Duzel, Emrah

    2004-01-01

    Recent evidence suggests a close functional relationship between memory formation in the hippocampus and dopaminergic neuromodulation originating in the ventral tegmental area and medial substantia nigra of the midbrain. Here we report midbrain activation in two functional MRI studies of visual memory in healthy young adults. In the first study,…

  7. The Role of Memory Activation in Creating False Memories of Encoding Context

    ERIC Educational Resources Information Center

    Arndt, Jason

    2010-01-01

    Using 3 experiments, I examined false memory for encoding context by presenting Deese-Roediger-McDermott themes (Deese, 1959; Roediger & McDermott, 1995) in usual-looking fonts and by testing related, but unstudied, lure items in a font that was shown during encoding. In 2 of the experiments, testing lure items in the font used to study their…

  8. Memory-Based Decision-Making with Heuristics: Evidence for a Controlled Activation of Memory Representations

    ERIC Educational Resources Information Center

    Khader, Patrick H.; Pachur, Thorsten; Meier, Stefanie; Bien, Siegfried; Jost, Kerstin; Rosler, Frank

    2011-01-01

    Many of our daily decisions are memory based, that is, the attribute information about the decision alternatives has to be recalled. Behavioral studies suggest that for such decisions we often use simple strategies (heuristics) that rely on controlled and limited information search. It is assumed that these heuristics simplify decision-making by…

  9. Overlapping Parietal Activity in Memory and Perception: Evidence for the Attention to Memory Model

    ERIC Educational Resources Information Center

    Cabeza, Roberto; Mazuz, Yonatan S.; Stokes, Jared; Kragel, James E.; Woldorff, Marty G.; Ciaramelli, Elisa; Olson, Ingrid R.; Moscovitch, Morris

    2011-01-01

    The specific role of different parietal regions to episodic retrieval is a topic of intense debate. According to the Attention to Memory (AtoM) model, dorsal parietal cortex (DPC) mediates top-down attention processes guided by retrieval goals, whereas ventral parietal cortex (VPC) mediates bottom-up attention processes captured by the retrieval…

  10. Fear Extinction Memory Consolidation Requires Potentiation of Pontine-Wave Activity during REM Sleep

    PubMed Central

    Datta, Subimal; O'Malley, Matthew W .

    2013-01-01

    Sleep plays an important role in memory consolidation within multiple memory systems including contextual fear extinction memory, but little is known about the mechanisms that underlie this process. Here, we show that fear extinction training in rats, which extinguished conditioned fear, increased both slow-wave sleep and rapid-eye movement (REM) sleep. Surprisingly, 24 h later, during memory testing, only 57% of the fear-extinguished animals retained fear extinction memory. We found that these animals exhibited an increase in phasic pontine-wave (P-wave) activity during post-training REM sleep, which was absent in the 43% of animals that failed to retain fear extinction memory. The results of this study provide evidence that brainstem activation, specifically potentiation of phasic P-wave activity, during post-training REM sleep is critical for consolidation of fear extinction memory. The results of this study also suggest that, contrary to the popular hypothesis of sleep and memory, increased sleep after training alone does not guarantee consolidation and/or retention of fear extinction memory. Rather, the potentiation of specific sleep-dependent physiological events may be a more accurate predictor for successful consolidation of fear extinction memory. Identification of this unique mechanism will significantly improve our present understanding of the cellular and molecular mechanisms that underlie the sleep-dependent regulation of emotional memory. Additionally, this discovery may also initiate development of a new, more targeted treatment method for clinical disorders of fear and anxiety in humans that is more efficacious than existing methods such as exposure therapy that incorporate only fear extinction. PMID:23467372

  11. Tumoricidal effector mechanisms of murine Bacillus Calmette-Guérin-activated macrophages: mediation of cytolysis, mitochondrial respiration inhibition, and release of intracellular iron by distinct mechanisms.

    PubMed

    Klostergaard, J; Leroux, M E; Ezell, S M; Kull, F C

    1987-04-15

    Murine Bacillus Calmette-Guérin-activated macrophages mediate discrete cytotoxic effects in cocultured tumor target cells in vitro. These effects include: the loss of intracellular iron, in part associated with reversible inhibition of the Kreb's cycle enzyme, aconitase; cytostasis, associated with reversible lesions inflicted in the electron transport chain (ETC) of the mitochondria resulting in reversible loss of proliferative capacity; and cytolysis, manifested by eventual gross perturbation of the integrity of the plasma membrane. We demonstrate that these manifestations of cytotoxicity are the result of three independent mechanisms employing apparently distinct macromolecules for their commission. Analysis of target cells that are highly susceptible (L-929), highly resistant (L-1210), or have incomplete resistance (EMT-6) to the cytolytic effects of cocultured activated macrophages indicates that there is no consistent relationship between the release of intracellular 59Fe and 51Cr. Thus, perturbation of intracellular iron pools did not appear to be an obligatory step on the pathway to cytolysis. Further evidence for this dissociation was obtained by employing a specific heteroantiserum reactive with cytolytic molecule(s). This antiserum could block the cytolytic response (51Cr release of cocultured L-929 and EMT-6 targets) but had no effect on the extent of iron release from viable EMT-6 or L-1210 targets. Furthermore, the cytolytic factor itself was incapable of mediating effects on the ETC or in causing release of intracellular iron. Two lines of evidence suggested that effects on the ETC are not linked with loss of intracellular iron. First, the monokine respiration inhibitory factor was incapable of causing release of intracellular iron from target cells in which the mitochondria were strongly suppressed. Second, the kinetics of release of respiration inhibitory factor from endotoxin-triggered Bacillus Calmette-Guérin-activated macrophages indicate a

  12. ERK Pathway Activation Bidirectionally Affects Visual Recognition Memory and Synaptic Plasticity in the Perirhinal Cortex

    PubMed Central

    Silingardi, Davide; Angelucci, Andrea; De Pasquale, Roberto; Borsotti, Marco; Squitieri, Giovanni; Brambilla, Riccardo; Putignano, Elena; Pizzorusso, Tommaso; Berardi, Nicoletta

    2011-01-01

    ERK 1,2 pathway mediates experience-dependent gene transcription in neurons and several studies have identified its pivotal role in experience-dependent synaptic plasticity and in forms of long term memory involving hippocampus, amygdala, or striatum. The perirhinal cortex (PRHC) plays an essential role in familiarity-based object recognition memory. It is still unknown whether ERK activation in PRHC is necessary for recognition memory consolidation. Most important, it is unknown whether by modulating the gain of the ERK pathway it is possible to bidirectionally affect visual recognition memory and PRHC synaptic plasticity. We have first pharmacologically blocked ERK activation in the PRHC of adult mice and found that this was sufficient to impair long term recognition memory in a familiarity-based task, the object recognition task (ORT). We have then tested performance in the ORT in Ras-GRF1 knock-out (KO) mice, which exhibit a reduced activation of ERK by neuronal activity, and in ERK1 KO mice, which have an increased activation of ERK2 and exhibit enhanced striatal plasticity and striatal mediated memory. We found that Ras-GRF1 KO mice have normal short term memory but display a long term memory deficit; memory reconsolidation is also impaired. On the contrary, ERK1 KO mice exhibit a better performance than WT mice at 72 h retention interval, suggesting a longer lasting recognition memory. In parallel with behavioral data, LTD was strongly reduced and LTP was significantly smaller in PRHC slices from Ras-GRF1 KO than in WT mice while enhanced LTP and LTD were found in PRHC slices from ERK1 KO mice. PMID:22232579

  13. Specific marker of feigned memory impairment: The activation of left superior frontal gyrus.

    PubMed

    Chen, Zi-Xiang; Xue, Li; Liang, Chun-Yu; Wang, Li-Li; Mei, Wei; Zhang, Qiang; Zhao, Hu

    2015-11-01

    Faking memory impairment means normal people complain lots of memory problems without organic damage in forensic assessments. Using alternative forced-choice paradigm, containing digital or autobiographical information, previous neuroimaging studies have indicated that faking memory impairment could cause the activation in the prefrontal and parietal regions, and might involve a fronto-parietal-subcortical circuit. However, it is still unclear whether different memory types have influence on faking or not. Since different memory types, such as long-term memory (LTM) and short-term memory (STM), were found supported by different brain areas, we hypothesized that feigned STM or LTM impairment had distinct neural activation mapping. Besides that, some common neural correlates may act as the general characteristic of feigned memory impairment. To verify this hypothesis, the functional magnetic resonance imaging (fMRI) combined with an alternative word forced-choice paradigm were used in this study. A total of 10 right-handed participants, in this study, had to perform both STW and LTM tasks respectively under answering correctly, answering randomly and feigned memory impairment conditions. Our results indicated that the activation of the left superior frontal gyrus and the left medial frontal gyrus was associated with feigned LTM impairment, whereas the left superior frontal gyrus, the left precuneus and the right anterior cingulate cortex (ACC) were highly activated while feigning STM impairment. Furthermore, an overlapping was found in the left superior frontal gyrus, and it suggested that the activity of the left superior frontal gyrus might be acting as a specific marker of feigned memory impairment. PMID:26479324

  14. Quantitative assessment of the functional plasticity of memory CD8(+) T cells.

    PubMed

    Baz, Adriana; Groves, Penny; Buttigieg, Kathy; Apte, Simon H; Kienzle, Norbert; Kelso, Anne

    2016-04-01

    While the functional plasticity of memory CD4(+) T cells has been studied extensively, less is known about this property in memory CD8(+) T cells. Here, we report the direct measurement of plasticity by paired daughter analysis of effector and memory OT-I CD8(+) T cells primed in vivo with ovalbumin. Naïve, effector, and memory OT-I cells were isolated and activated in single-cell culture; then, after the first division, their daughter cells were transferred to new cultures with and without IL-4; expression of IFN-γ and IL-4 mRNAs was measured 5 days later in the resultant subclones. Approximately 40% of clonogenic memory CD8(+) T cells were bipotential in this assay, giving rise to an IL-4(-) subclone in the absence of IL-4 and an IL-4(+) subclone in the presence of IL-4. The frequency of bipotential cells was lower among memory cells than naïve cells but markedly higher than among 8-day effectors. Separation based on high or low expression of CD62L, CD122, CD127, or Ly6C did not identify a phenotypic marker of the bipotential cells. Functional plasticity in memory CD8(+) T-cell populations can therefore reflect modulation at the level of a single memory cell and its progeny.

  15. Quantitative assessment of the functional plasticity of memory CD8(+) T cells.

    PubMed

    Baz, Adriana; Groves, Penny; Buttigieg, Kathy; Apte, Simon H; Kienzle, Norbert; Kelso, Anne

    2016-04-01

    While the functional plasticity of memory CD4(+) T cells has been studied extensively, less is known about this property in memory CD8(+) T cells. Here, we report the direct measurement of plasticity by paired daughter analysis of effector and memory OT-I CD8(+) T cells primed in vivo with ovalbumin. Naïve, effector, and memory OT-I cells were isolated and activated in single-cell culture; then, after the first division, their daughter cells were transferred to new cultures with and without IL-4; expression of IFN-γ and IL-4 mRNAs was measured 5 days later in the resultant subclones. Approximately 40% of clonogenic memory CD8(+) T cells were bipotential in this assay, giving rise to an IL-4(-) subclone in the absence of IL-4 and an IL-4(+) subclone in the presence of IL-4. The frequency of bipotential cells was lower among memory cells than naïve cells but markedly higher than among 8-day effectors. Separation based on high or low expression of CD62L, CD122, CD127, or Ly6C did not identify a phenotypic marker of the bipotential cells. Functional plasticity in memory CD8(+) T-cell populations can therefore reflect modulation at the level of a single memory cell and its progeny. PMID:26799367

  16. The Conserved MAPK Site in E(spl)-M8, an Effector of Drosophila Notch Signaling, Controls Repressor Activity during Eye Development

    PubMed Central

    Bandyopadhyay, Mohna; Bishop, Clifton P.

    2016-01-01

    The specification of patterned R8 photoreceptors at the onset of eye development depends on timely inhibition of Atonal (Ato) by the Enhancer of split (E(spl) repressors. Repression of Ato by E(spl)-M8 requires the kinase CK2 and is inhibited by the phosphatase PP2A. The region targeted by CK2 harbors additional conserved Ser residues, raising the prospect of regulation via multi-site phosphorylation. Here we investigate one such motif that meets the consensus for modification by MAPK, a well-known effector of Epidermal Growth Factor Receptor (EGFR) signaling. Our studies reveal an important role for the predicted MAPK site of M8 during R8 birth. Ala/Asp mutations reveal that the CK2 and MAPK sites ensure that M8 repression of Ato and the R8 fate occurs in a timely manner and at a specific stage (stage-2/3) of the morphogenetic furrow (MF). M8 repression of Ato is mitigated by halved EGFR dosage, and this effect requires an intact MAPK site. Accordingly, variants with a phosphomimetic Asp at the MAPK site exhibit earlier (inappropriate) activity against Ato even at stage-1 of the MF, where a positive feedback-loop is necessary to raise Ato levels to a threshold sufficient for the R8 fate. Analysis of deletion variants reveals that both kinase sites (CK2 and MAPK) contribute to ‘cis’-inhibition of M8. This key regulation by CK2 and MAPK is bypassed by the E(spl)D mutation encoding the truncated protein M8*, which potently inhibits Ato at stage-1 of R8 birth. We also provide evidence that PP2A likely targets the MAPK site. Thus multi-site phosphorylation controls timely onset of M8 repressor activity in the eye, a regulation that appears to be dispensable in the bristle. The high conservation of the CK2 and MAPK sites in the insect E(spl) proteins M7, M5 and Mγ, and their mammalian homologue HES6, suggest that this mode of regulation may enable E(spl)/HES proteins to orchestrate repression by distinct tissue-specific mechanisms, and is likely to have broader

  17. Resident memory T cells in human health and disease

    PubMed Central

    Clark, Rachael A.

    2015-01-01

    Resident memory T cells are non-recirculating memory T cells that persist long term in epithelial barrier tissues, including the gastrointestinal tract, lung, skin and reproductive tract. Resident memory T cells persist in the absence of antigens, have impressive effector functions and provide rapid on-site immune protection against known pathogens in peripheral tissues. A fundamentally distinct gene expression program differentiates resident memory T cells from circulating T cells. Although these cells likely evolved to provide rapid immune protection against pathogens, autoreactive, aberrantly activated and malignant resident memory cells contribute to numerous human inflammatory diseases including mycosis fungoides and psoriasis. This review will discuss both the science and medicine of resident memory T cells, exploring how these cells contribute to healthy immune function and discussing what is known about how these cells contribute to human inflammatory and autoimmune diseases. PMID:25568072

  18. Phenotypic analysis of nylon-wool-adherent suppressor cells that inhibit the effector process of tumour cell lysis by lymphokine-activated killer cells in patients with advanced gastric carcinoma.

    PubMed

    Koyama, S; Fukao, K

    1994-01-01

    The causes of down-regulation of cytotoxic immune responses in cancer patients have not been fully evaluated. We previously demonstrated that T-cell-growth-factor-activated peripheral blood lymphocytes (PBL) with the surface phenotype CD8+ CD11b-, from patients with widespread metastasis of gastric carcinoma, inhibited the effector process of lymphokine-activated-killer(LAK)-cell-mediated cytolysis. In this study, we examined suppressor cell activity in freshly prepared PBL from 18 patients with advanced gastric carcinoma, and 10 normal healthy individuals. The suppressor cell activity was assayed by recording whether or not PBL inhibited directly the effector process of LAK cell cytotoxicity. Most of the PBL suspensions from cancer patients showed that they contained a population of cells that can directly inhibit the effector phase of tumor cell lysis of the cytotoxic cells. To analyze further the PBL responsible for the suppression, the cells were passed over a nylon-wool column. Nylon-wool-adherent cells significantly augmented the suppression, while the cells passing through abrogated the suppressive effect. Most nylon-wool-adherent cells from 10 normal healthy controls did not inhibit the cytotoxic reaction. To determine further the suppressor-effector population in nylon-wool-adherent cells, negative-selection studies using CD8-, CD4- or CD11b-coated magnetic beads, and positive-selection studies using CD8- or CD4-coated magnetic beads were performed. Finally the results suggest that the suppressor-effector cells comprise at least two different surface phenotypes: CD8+ T and CD8-CD11b+ cells. The possible role of CD4+ T cells and HLA-DR+ LeuM3+ macrophages as suppressor cells was ruled out in nylon-wool-adherent cells. CD8+ T and possibly CD8-CD11b+ cells apparently suppressed the efferent limb of the antitumor immunity. The selective immune suppression mediated by these cells may partly be concerned with escape mechanisms of gastric carcinoma from the host

  19. Lithium activates brain phospholipase A2 and improves memory in rats: implications for Alzheimer's disease.

    PubMed

    Mury, Fábio B; da Silva, Weber C; Barbosa, Nádia R; Mendes, Camila T; Bonini, Juliana S; Sarkis, Jorge Eduardo Souza; Cammarota, Martin; Izquierdo, Ivan; Gattaz, Wagner F; Dias-Neto, Emmanuel

    2016-10-01

    Phospholipase A2 (Pla2) is required for memory retrieval, and its inhibition in the hippocampus has been reported to impair memory acquisition in rats. Moreover, cognitive decline and memory deficits showed to be reduced in animal models after lithium treatment, prompting us to evaluate possible links between Pla2, lithium and memory. Here, we evaluated the possible modulation of Pla2 activity by a long-term treatment of rats with low doses of lithium and its impact in memory. Wistar rats were trained for the inhibitory avoidance task, treated with lithium for 100 days and tested for perdurability of long-term memory. Hippocampal samples were used for quantifying the expression of 19 brain-expressed Pla2 genes and for evaluating the enzymatic activity of Pla2 using group-specific radio-enzymatic assays. Our data pointed to a significant perdurability of long-term memory, which correlated with increased transcriptional and enzymatic activities of certain members of the Pla2 family (iPla2 and sPla2) after the chronic lithium treatment. Our data suggest new possible targets of lithium, add more information on its pharmacological activity and reinforce the possible use of low doses of lithium for the treatment of neurodegenerative conditions such as the Alzheimer's disease. PMID:26661385

  20. Lithium activates brain phospholipase A2 and improves memory in rats: implications for Alzheimer's disease.

    PubMed

    Mury, Fábio B; da Silva, Weber C; Barbosa, Nádia R; Mendes, Camila T; Bonini, Juliana S; Sarkis, Jorge Eduardo Souza; Cammarota, Martin; Izquierdo, Ivan; Gattaz, Wagner F; Dias-Neto, Emmanuel

    2016-10-01

    Phospholipase A2 (Pla2) is required for memory retrieval, and its inhibition in the hippocampus has been reported to impair memory acquisition in rats. Moreover, cognitive decline and memory deficits showed to be reduced in animal models after lithium treatment, prompting us to evaluate possible links between Pla2, lithium and memory. Here, we evaluated the possible modulation of Pla2 activity by a long-term treatment of rats with low doses of lithium and its impact in memory. Wistar rats were trained for the inhibitory avoidance task, treated with lithium for 100 days and tested for perdurability of long-term memory. Hippocampal samples were used for quantifying the expression of 19 brain-expressed Pla2 genes and for evaluating the enzymatic activity of Pla2 using group-specific radio-enzymatic assays. Our data pointed to a significant perdurability of long-term memory, which correlated with increased transcriptional and enzymatic activities of certain members of the Pla2 family (iPla2 and sPla2) after the chronic lithium treatment. Our data suggest new possible targets of lithium, add more information on its pharmacological activity and reinforce the possible use of low doses of lithium for the treatment of neurodegenerative conditions such as the Alzheimer's disease.

  1. [Activity of Ginkgo biloba extract on short-term memory].

    PubMed

    Hindmarch, I

    1986-09-25

    Eight healthy female volunteers were included in a double-blind, cross-over trial comparing Ginkgo biloba extract in acute and ascending doses (120, 240, 600 mg) with a placebo. One hour after treatment they were subjected to a battery of tests, including: critical flicker fusion, choice reaction time, subjective rating scale and Sternberg memory scanning test. No statistically significant differences with the placebo were observed in the first three tests. In contrast, short term memory, as assessed by the Sternberg technique, was very significantly improved following 600 mg of Ginkgo biloba extract, as compared with the placebo. These results differentiate Ginkgo biloba extract from sedative and stimulant drugs and suggest a specific effect on memory processes.

  2. Dynamic trajectory of multiple single-unit activity during working memory task in rats

    PubMed Central

    Zhang, Xiaofan; Yi, Hu; Bai, Wenwen; Tian, Xin

    2015-01-01

    Working memory plays an important role in complex cognitive tasks. A popular theoretical view is that transient properties of neuronal dynamics underlie cognitive processing. The question raised here as to how the transient dynamics evolve in working memory. To address this issue, we investigated the multiple single-unit activity dynamics in rat medial prefrontal cortex (mPFC) during a Y-maze working memory task. The approach worked by reconstructing state space from delays of the original single-unit firing rate variables, which were further analyzed using kernel principal component analysis (KPCA). Then the neural trajectories were obtained to visualize the multiple single-unit activity. Furthermore, the maximal Lyapunov exponent (MLE) was calculated to quantitatively evaluate the neural trajectories during the working memory task. The results showed that the neuronal activity produced stable and reproducible neural trajectories in the correct trials while showed irregular trajectories in the incorrect trials, which may establish a link between the neurocognitive process and behavioral performance in working memory. The MLEs significantly increased during working memory in the correctly performed trials, indicating an increased divergence of the neural trajectories. In the incorrect trials, the MLEs were nearly zero and remained unchanged during the task. Taken together, the trial-specific neural trajectory provides an effective way to track the instantaneous state of the neuronal population during the working memory task and offers valuable insights into working memory function. The MLE describes the changes of neural dynamics in working memory and may reflect different neuronal population states in working memory. PMID:26441626

  3. Biomedical Applications of Thermally Activated Shape Memory Polymers

    SciTech Connect

    Small IV, W; Singhal, P; Wilson, T S; Maitland, D J

    2009-04-10

    Shape memory polymers (SMPs) are smart materials that can remember a primary shape and can return to this primary shape from a deformed secondary shape when given an appropriate stimulus. This property allows them to be delivered in a compact form via minimally invasive surgeries in humans, and deployed to achieve complex final shapes. Here we review the various biomedical applications of SMPs and the challenges they face with respect to actuation and biocompatibility. While shape memory behavior has been demonstrated with heat, light and chemical environment, here we focus our discussion on thermally stimulated SMPs.

  4. Biomedical applications of thermally activated shape memory polymers†

    PubMed Central

    Small, Ward; Singhal, Pooja; Wilson, Thomas S.

    2011-01-01

    Shape memory polymers (SMPs) are smart materials that can remember a primary shape and can return to this primary shape from a deformed secondary shape when given an appropriate stimulus. This property allows them to be delivered in a compact form via minimally invasive surgeries in humans, and deployed to achieve complex final shapes. Here we review the various biomedical applications of SMPs and the challenges they face with respect to actuation and biocompatibility. While shape memory behavior has been demonstrated with heat, light and chemical environment, here we focus our discussion on thermally stimulated SMPs. PMID:21258605

  5. The Associative Memory Deficit in Aging Is Related to Reduced Selectivity of Brain Activity during Encoding.

    PubMed

    Saverino, Cristina; Fatima, Zainab; Sarraf, Saman; Oder, Anita; Strother, Stephen C; Grady, Cheryl L

    2016-09-01

    Human aging is characterized by reductions in the ability to remember associations between items, despite intact memory for single items. Older adults also show less selectivity in task-related brain activity, such that patterns of activation become less distinct across multiple experimental tasks. This reduced selectivity or dedifferentiation has been found for episodic memory, which is often reduced in older adults, but not for semantic memory, which is maintained with age. We used fMRI to investigate whether there is a specific reduction in selectivity of brain activity during associative encoding in older adults, but not during item encoding, and whether this reduction predicts associative memory performance. Healthy young and older adults were scanned while performing an incidental encoding task for pictures of objects and houses under item or associative instructions. An old/new recognition test was administered outside the scanner. We used agnostic canonical variates analysis and split-half resampling to detect whole-brain patterns of activation that predicted item versus associative encoding for stimuli that were later correctly recognized. Older adults had poorer memory for associations than did younger adults, whereas item memory was comparable across groups. Associative encoding trials, but not item encoding trials, were predicted less successfully in older compared with young adults, indicating less distinct patterns of associative-related activity in the older group. Importantly, higher probability of predicting associative encoding trials was related to better associative memory after accounting for age and performance on a battery of neuropsychological tests. These results provide evidence that neural distinctiveness at encoding supports associative memory and that a specific reduction of selectivity in neural recruitment underlies age differences in associative memory. PMID:27082043

  6. MARTX toxins as effector delivery platforms.

    PubMed

    Gavin, Hannah E; Satchell, Karla J F

    2015-12-01

    Bacteria frequently manipulate their host environment via delivery of microbial 'effector' proteins to the cytosol of eukaryotic cells. In the case of the multifunctional autoprocessing repeats-in-toxins (MARTX) toxin, this phenomenon is accomplished by a single, >3500 amino acid polypeptide that carries information for secretion, translocation, autoprocessing and effector activity. MARTX toxins are secreted from bacteria by dedicated Type I secretion systems. The released MARTX toxins form pores in target eukaryotic cell membranes for the delivery of up to five cytopathic effectors, each of which disrupts a key cellular process. Targeted cellular processes include modulation or modification of small GTPases, manipulation of host cell signaling and disruption of cytoskeletal integrity. More recently, MARTX toxins have been shown to be capable of heterologous protein translocation. Found across multiple bacterial species and genera--frequently in pathogens lacking Type 3 or Type 4 secretion systems--MARTX toxins in multiple cases function as virulence factors. Innovative research at the intersection of toxin biology and bacterial genetics continues to elucidate the intricacies of the toxin as well as the cytotoxic mechanisms of its diverse effector collection.

  7. Induction of an Olfactory Memory by the Activation of a Metabotropic Glutamate Receptor

    NASA Astrophysics Data System (ADS)

    Kaba, Hideto; Hayashi, Yasunori; Higuchi, Takashi; Nakanishi, Shigetada

    1994-07-01

    Female mice form an olfactory memory of male pheromones at mating; exposure to the pheromones of a strange male after that mating will block pregnancy. The formation of this memory is mediated by the accessory olfactory system, in which an increase in norepinephrine after mating reduces inhibitory transmission of γ-aminobutyric acid from the granule cells to the mitral cells. This study shows that the activation of mGluR2, a metabotropic glutamate receptor that suppresses the γ-aminobutyric acid inhibition of the mitral cells, permits the formation of a specific olfactory memory without the occurrence of mating by infusion of mGluR2 agonists into the female's accessory olfactory bulb. This memory faithfully reflects the memory formed at mating.

  8. Activity in Prelimbic Cortex Subserves Fear Memory Reconsolidation over Time

    ERIC Educational Resources Information Center

    Stern, Cristina A. J.; Gazarini, Lucas; Vanvossen, Ana C.; Hames, Mayara S.; Bertoglio, Leandro J.

    2014-01-01

    The prelimbic cortex has been implicated in the consolidation of previously learned fear. Herein, we report that temporarily inactivating this medial prefrontal cortex subregion with the GABA [subscript A] agonist muscimol (4.0 nmol in 0.2 µL per hemisphere) was able to equally disrupt 1-, 7-, and 21-d-old contextual fear memories after their…

  9. Opening the Ralstonia solanacearum type III effector tool box: insights into host cell subversion mechanisms.

    PubMed

    Deslandes, Laurent; Genin, Stephane

    2014-08-01

    Effectors delivered to host cells by the Type III secretion system are essential to Ralstonia solanacearum pathogenicity, as in several other plant pathogenic bacteria. The establishment of exhaustive effector repertoires in multiple R. solanacearum strains drew a first picture of the evolutionary dynamics of the pathogen effector suites. Effector repertoires are diversified, with a core of 20-30 effectors present in most of the strains and the obtention of mutants lacking one or more effector genes revealed the functional overlap among this effector network. Recent functional studies have provided insights into the ability of single effectors to manipulate the host proteasome, elicit cell death, trigger the expression of plant genes, and/or display biochemical activities on plant protein targets.

  10. KLRG+ invariant natural killer T cells are long-lived effectors.

    PubMed

    Shimizu, Kanako; Sato, Yusuke; Shinga, Jun; Watanabe, Takashi; Endo, Takaho; Asakura, Miki; Yamasaki, Satoru; Kawahara, Kazuyoshi; Kinjo, Yuki; Kitamura, Hiroshi; Watarai, Hiroshi; Ishii, Yasuyuki; Tsuji, Moriya; Taniguchi, Masaru; Ohara, Osamu; Fujii, Shin-ichiro

    2014-08-26

    Immunological memory has been regarded as a unique feature of the adaptive immune response mediated in an antigen-specific manner by T and B lymphocytes. However, natural killer (NK) cells and γδT cells, which traditionally are classified as innate immune cells, have been shown in recent studies to have hallmark features of memory cells. Invariant NKT cell (iNKT cell)-mediated antitumor effects indicate that iNKT cells are activated in vivo by vaccination with iNKT cell ligand-loaded CD1d(+) cells, but not by vaccination with unbound NKT cell ligand. In such models, it previously was thought that the numbers of IFN-γ-producing cells in the spleen returned to the basal level around 1 wk after the vaccination. In the current study, we demonstrate the surprising presence of effector memory-like iNKT cells in the lung. We found long-term antitumor activity in the lungs of mice was enhanced after vaccination with iNKT cell ligand-loaded dendritic cells. Further analyses showed that the KLRG1(+) (Killer cell lectin-like receptor subfamily G, member 1-positive) iNKT cells coexpressing CD49d and granzyme A persisted for several months and displayed a potent secondary response to cognate antigen. Finally, analyses of CDR3β by RNA deep sequencing demonstrated that some particular KLRG1(+) iNKT-cell clones accumulated, suggesting the selection of certain T-cell receptor repertoires by an antigen. The current findings identifying effector memory-like KLRG1(+) iNKT cells in the lung could result in a paradigm shift regarding the basis of newly developed extrathymic iNKT cells and could contribute to the future development of antitumor immunotherapy by uniquely energizing iNKT cells. PMID:25118276

  11. Emotion regulation modulates anticipatory brain activity that predicts emotional memory encoding in women.

    PubMed

    Galli, Giulia; Griffiths, Victoria A; Otten, Leun J

    2014-03-01

    It has been shown that the effectiveness with which unpleasant events are encoded into memory is related to brain activity set in train before the events. Here, we assessed whether encoding-related activity before an aversive event can be modulated by emotion regulation. Electrical brain activity was recorded from the scalps of healthy women while they performed an incidental encoding task on randomly intermixed unpleasant and neutral visual scenes. A cue presented 1.5 s before each picture indicated the upcoming valence. In half of the blocks of trials, the instructions emphasized to let emotions arise in a natural way. In the other half, participants were asked to decrease their emotional response by adopting the perspective of a detached observer. Memory for the scenes was probed 1 day later with a recognition memory test. Brain activity before unpleasant scenes predicted later memory of the scenes, but only when participants felt their emotions and did not detach from them. The findings indicate that emotion regulation can eliminate the influence of anticipatory brain activity on memory encoding. This may be relevant for the understanding and treatment of psychiatric diseases with a memory component.

  12. Disentangling the roles of arousal and amygdala activation in emotional declarative memory.

    PubMed

    de Voogd, Lycia D; Fernández, Guillén; Hermans, Erno J

    2016-09-01

    A large body of evidence in animals and humans implicates the amygdala in promoting memory for arousing experiences. Although the amygdala can trigger threat-related noradrenergic-sympathetic arousal, in humans amygdala activation and noradrenergic-sympathetic arousal do not always concur. This raises the question how these two processes play a role in enhancing emotional declarative memory. This study was designed to disentangle these processes in a combined subsequent-memory/fear-conditioning paradigm with neutral items belonging to two conceptual categories as conditioned stimuli. Functional MRI, skin conductance (index of sympathetic activity), and pupil dilation (indirect index of central noradrenergic activity) were acquired throughout procedures. Recognition memory for individual items was tested 24 h later. We found that pupil dilation and skin conductance responses were higher on CS+ (associated with a shock) compared with CS- trials, irrespective of later memory for those items. By contrast, amygdala activity was only higher for CS+ items that were later confidently remembered compared with CS+ items that were later forgotten. Thus, amygdala activity and not noradrenergic-sympathetic arousal, predicted enhanced declarative item memory. This dissociation is in line with animal models stating that the amygdala integrates arousal-related neuromodulatory changes to alter mnemonic processes elsewhere in the brain. PMID:27217115

  13. Disentangling the roles of arousal and amygdala activation in emotional declarative memory.

    PubMed

    de Voogd, Lycia D; Fernández, Guillén; Hermans, Erno J

    2016-09-01

    A large body of evidence in animals and humans implicates the amygdala in promoting memory for arousing experiences. Although the amygdala can trigger threat-related noradrenergic-sympathetic arousal, in humans amygdala activation and noradrenergic-sympathetic arousal do not always concur. This raises the question how these two processes play a role in enhancing emotional declarative memory. This study was designed to disentangle these processes in a combined subsequent-memory/fear-conditioning paradigm with neutral items belonging to two conceptual categories as conditioned stimuli. Functional MRI, skin conductance (index of sympathetic activity), and pupil dilation (indirect index of central noradrenergic activity) were acquired throughout procedures. Recognition memory for individual items was tested 24 h later. We found that pupil dilation and skin conductance responses were higher on CS+ (associated with a shock) compared with CS- trials, irrespective of later memory for those items. By contrast, amygdala activity was only higher for CS+ items that were later confidently remembered compared with CS+ items that were later forgotten. Thus, amygdala activity and not noradrenergic-sympathetic arousal, predicted enhanced declarative item memory. This dissociation is in line with animal models stating that the amygdala integrates arousal-related neuromodulatory changes to alter mnemonic processes elsewhere in the brain.

  14. Induction of memory cytotoxic T cells to influenza A virus and subsequent viral clearance is not modulated by PB1-F2-dependent inflammasome activation

    PubMed Central

    Lee, Patricia (Hoi Yee); Bird, Nicola; MacKenzie-Kludas, Charley; Mansell, Ashley; Kedzierska, Katherine; Brown, Lorena; McAuley, Julie

    2016-01-01

    Expression of the viral virulence protein PB1-F2 during infection has been linked to NLRP3 inflammasome complex activation in macrophages and induction of early inflammatory events enhancing immunopathology during influenza disease. We sought to determine whether PB1-F2-specific NLRP3 inflammasome activation influenced the magnitude and/or robustness of the CD8+ T-cell responses specific for conserved viral antigens and subsequent virus elimination. Using murine heterosubtypic viral infection models, we showed that mice infected with virus unable to produce PB1-F2 protein showed no deficit in the overall magnitude and functional memory responses of CD8+ T cells established during the effector phase compared with those infected with wild-type PB1-F2-expressing virus and were equally capable of mounting robust recall responses. These data indicate that while expression of PB1-F2 protein can induce inflammatory events, the capacity to generate memory CD8+ T cells specific for immunodominant viral epitopes remains uncompromised. PMID:26667784

  15. Stimulation over primary motor cortex during action observation impairs effector recognition.

    PubMed

    Naish, Katherine R; Barnes, Brittany; Obhi, Sukhvinder S

    2016-04-01

    Recent work suggests that motor cortical processing during action observation plays a role in later recognition of the object involved in the action. Here, we investigated whether recognition of the effector making an action is also impaired when transcranial magnetic stimulation (TMS) - thought to interfere with normal cortical activity - is applied over the primary motor cortex (M1) during action observation. In two experiments, single-pulse TMS was delivered over the hand area of M1 while participants watched short clips of hand actions. Participants were then asked whether an image (experiment 1) or a video (experiment 2) of a hand presented later in the trial was the same or different to the hand in the preceding video. In Experiment 1, we found that participants' ability to recognise static images of hands was significantly impaired when TMS was delivered over M1 during action observation, compared to when no TMS was delivered, or when stimulation was applied over the vertex. Conversely, stimulation over M1 did not affect recognition of dot configurations, or recognition of hands that were previously presented as static images (rather than action movie clips) with no object. In Experiment 2, we found that effector recognition was impaired when stimulation was applied part way through (300ms) and at the end (500ms) of the action observation period, indicating that 200ms of action-viewing following stimulation was not long enough to form a new representation that could be used for later recognition. The findings of both experiments suggest that interfering with cortical motor activity during action observation impairs subsequent recognition of the effector involved in the action, which complements previous findings of motor system involvement in object memory. This work provides some of the first evidence that motor processing during action observation is involved in forming representations of the effector that are useful beyond the action observation period

  16. Progressive CD4+ central–memory T cell decline results in CD4+ effector–memory insufficiency and overt disease in chronic SIV infection

    PubMed Central

    Okoye, Afam; Meier-Schellersheim, Martin; Brenchley, Jason M.; Hagen, Shoko I.; Walker, Joshua M.; Rohankhedkar, Mukta; Lum, Richard; Edgar, John B.; Planer, Shannon L.; Legasse, Alfred; Sylwester, Andrew W.; Piatak, Michael; Lifson, Jeffrey D.; Maino, Vernon C.; Sodora, Donald L.; Douek, Daniel C.; Axthelm, Michael K.; Grossman, Zvi; Picker, Louis J.

    2007-01-01

    Primary simian immunodeficiency virus (SIV) infections of rhesus macaques result in the dramatic depletion of CD4+ CCR5+ effector–memory T (TEM) cells from extra-lymphoid effector sites, but in most infections, an increased rate of CD4+ memory T cell proliferation appears to prevent collapse of effector site CD4+ TEM cell populations and acute-phase AIDS. Eventually, persistent SIV replication results in chronic-phase AIDS, but the responsible mechanisms remain controversial. Here, we demonstrate that in the chronic phase of progressive SIV infection, effector site CD4+ TEM cell populations manifest a slow, continuous decline, and that the degree of this depletion remains a highly significant correlate of late-onset AIDS. We further show that due to persistent immune activation, effector site CD4+ TEM cells are predominantly short-lived, and that their homeostasis is strikingly dependent on the production of new CD4+ TEM cells from central–memory T (TCM) cell precursors. The instability of effector site CD4+ TEM cell populations over time was not explained by increasing destruction of these cells, but rather was attributable to progressive reduction in their production, secondary to decreasing numbers of CCR5− CD4+ TCM cells. These data suggest that although CD4+ TEM cell depletion is a proximate mechanism of immunodeficiency, the tempo of this depletion and the timing of disease onset are largely determined by destruction, failing production, and gradual decline of CD4+ TCM cells. PMID:17724130

  17. Electrocorticography reveals the temporal dynamics of posterior parietal cortical activity during recognition memory decisions.

    PubMed

    Gonzalez, Alex; Hutchinson, J Benjamin; Uncapher, Melina R; Chen, Janice; LaRocque, Karen F; Foster, Brett L; Rangarajan, Vinitha; Parvizi, Josef; Wagner, Anthony D

    2015-09-01

    Theories of the neurobiology of episodic memory predominantly focus on the contributions of medial temporal lobe structures, based on extensive lesion, electrophysiological, and imaging evidence. Against this backdrop, functional neuroimaging data have unexpectedly implicated left posterior parietal cortex (PPC) in episodic retrieval, revealing distinct activation patterns in PPC subregions as humans make memory-related decisions. To date, theorizing about the functional contributions of PPC has been hampered by the absence of information about the temporal dynamics of PPC activity as retrieval unfolds. Here, we leveraged electrocorticography to examine the temporal profile of high gamma power (HGP) in dorsal PPC subregions as participants made old/new recognition memory decisions. A double dissociation in memory-related HGP was observed, with activity in left intraparietal sulcus (IPS) and left superior parietal lobule (SPL) differing in time and sign for recognized old items (Hits) and correctly rejected novel items (CRs). Specifically, HGP in left IPS increased for Hits 300-700 ms poststimulus onset, and decayed to baseline ∼200 ms preresponse. By contrast, HGP in left SPL increased for CRs early after stimulus onset (200-300 ms) and late in the memory decision (from 700 ms to response). These memory-related effects were unique to left PPC, as they were not observed in right PPC. Finally, memory-related HGP in left IPS and SPL was sufficiently reliable to enable brain-based decoding of the participant's memory state at the single-trial level, using multivariate pattern classification. Collectively, these data provide insights into left PPC temporal dynamics as humans make recognition memory decisions. PMID:26283375

  18. Activities, self-referent memory beliefs, and cognitive performance: evidence for direct and mediated relations.

    PubMed

    Jopp, Daniela; Hertzog, Christopher

    2007-12-01

    In this study, the authors investigated the role of activities and self-referent memory beliefs for cognitive performance in a life-span sample. A factor analysis identified 8 activity factors, including Developmental Activities, Experiential Activities, Social Activities, Physical Activities, Technology Use, Watching Television, Games, and Crafts. A second-order general activity factor was significantly related to a general factor of cognitive function as defined by ability tests. Structural regression models suggested that prediction of cognition by activity level was partially mediated by memory beliefs, controlling for age, education, health, and depressive affect. Models adding paths from general and specific activities to aspects of crystallized intelligence suggested additional unique predictive effects for some activities. In alternative models, nonsignificant effects of beliefs on activities were detected when cognition predicted both variables, consistent with the hypothesis that beliefs derive from monitoring cognition and have no influence on activity patterns. PMID:18179299

  19. Electrospun nanofiber membranes for electrically activated shape memory nanocomposites

    NASA Astrophysics Data System (ADS)

    Zhang, Fenghua; Zhang, Zhichun; Liu, Yanju; Leng, Jinsong

    2014-06-01

    A novel shape memory nanocomposite system, consisting of a thermoplastic Nafion polymer and ultrathin electrospun polyacrylonitrile (PAN)-based carbonization nanofiber membranes, is successfully synthesized. PAN-based carbonization nanofiber networks that offer responses to deformations are considered to be an excellent actuation source. Significant improvement in the electrical conductivity of carbon nanofiber membranes is found by adjusting the applied voltage power in the electrospinning PAN process varying from 7.85 to 12.30 S cm-1. The porous structure of the carbon nanofiber membranes provides a large specific surface area and interfacial contact area when combined with the polymer matrix. Shape memory Nafion nanocomposites filled with interpenetrating non-woven electrospun PAN carbonization membranes can be actuated by applying 14 V electrical voltage within 5 s. The results, as demonstrated through morphology, electrical and thermal measurements and a shape recovery test, suggest a valuable route to producing soft nanocomposites.

  20. Lethal giant larvae-1 deficiency enhances the CD8(+) effector T-cell response to antigen challenge in vivo.

    PubMed

    Ramsbottom, Kelly M; Sacirbegovic, Faruk; Hawkins, Edwin D; Kallies, Axel; Belz, Gabrielle T; Van Ham, Vanessa; Haynes, Nicole M; Durrant, Michael J; Humbert, Patrick O; Russell, Sarah M; Oliaro, Jane

    2016-03-01

    Lethal giant larvae-1 (Lgl-1) is an evolutionary conserved protein that regulates cell polarity in diverse lineages; however, the role of Lgl-1 in the polarity and function of immune cells remains to be elucidated. To assess the role of Lgl-1 in T cells, we generated chimeric mice with a hematopoietic system deficient for Lgl-1. Lgl-1 deficiency did not impair the activation or function of peripheral CD8(+) T cells in response to antigen presentation in vitro, but did skew effector and memory T-cell differentiation. When challenged with antigen-expressing virus or tumor, Lgl-1-deficient mice displayed altered T-cell responses. This manifested in a stronger antiviral and antitumor effector CD8(+) T-cell response, the latter resulting in enhanced control of MC38-OVA tumors. These results reveal a novel role for Lgl-1 in the regulation of virus-specific T-cell responses and antitumor immunity.

  1. Lethal giant larvae-1 deficiency enhances the CD8(+) effector T-cell response to antigen challenge in vivo.

    PubMed

    Ramsbottom, Kelly M; Sacirbegovic, Faruk; Hawkins, Edwin D; Kallies, Axel; Belz, Gabrielle T; Van Ham, Vanessa; Haynes, Nicole M; Durrant, Michael J; Humbert, Patrick O; Russell, Sarah M; Oliaro, Jane

    2016-03-01

    Lethal giant larvae-1 (Lgl-1) is an evolutionary conserved protein that regulates cell polarity in diverse lineages; however, the role of Lgl-1 in the polarity and function of immune cells remains to be elucidated. To assess the role of Lgl-1 in T cells, we generated chimeric mice with a hematopoietic system deficient for Lgl-1. Lgl-1 deficiency did not impair the activation or function of peripheral CD8(+) T cells in response to antigen presentation in vitro, but did skew effector and memory T-cell differentiation. When challenged with antigen-expressing virus or tumor, Lgl-1-deficient mice displayed altered T-cell responses. This manifested in a stronger antiviral and antitumor effector CD8(+) T-cell response, the latter resulting in enhanced control of MC38-OVA tumors. These results reveal a novel role for Lgl-1 in the regulation of virus-specific T-cell responses and antitumor immunity. PMID:26391810

  2. Spatial memory extinction differentially affects dorsal and ventral hippocampal metabolic activity and associated functional brain networks.

    PubMed

    Méndez-Couz, Marta; González-Pardo, Héctor; Vallejo, Guillermo; Arias, Jorge L; Conejo, Nélida M

    2016-10-01

    Previous studies showed the involvement of brain regions associated with both spatial learning and associative learning in spatial memory extinction, although the specific role of the dorsal and ventral hippocampus and the extended hippocampal system including the mammillary body in the process is still controversial. The present study aimed to identify the involvement of the dorsal and ventral hippocampus, together with cortical regions, the amygdaloid nuclei, and the mammillary bodies in the extinction of a spatial memory task. To address these issues, quantitative cytochrome c oxidase histochemistry was applied as a metabolic brain mapping method. Rats were trained in a reference memory task using the Morris water maze, followed by an extinction procedure of the previously acquired memory task. Results show that rats learned successfully the spatial memory task as shown by the progressive decrease in measured latencies to reach the escape platform and the results obtained in the probe test. Spatial memory was subsequently extinguished as shown by the descending preference for the previously reinforced location. A control naïve group was added to ensure that brain metabolic changes were specifically related with performance in the spatial memory extinction task. Extinction of the original spatial learning task significantly modified the metabolic activity in the dorsal and ventral hippocampus, the amygdala and the mammillary bodies. Moreover, the ventral hippocampus, the lateral mammillary body and the retrosplenial cortex were differentially recruited in the spatial memory extinction task, as shown by group differences in brain metabolic networks. These findings provide new insights on the brain regions and functional brain networks underlying spatial memory, and specifically spatial memory extinction. © 2016 Wiley Periodicals, Inc.

  3. Spatial memory extinction differentially affects dorsal and ventral hippocampal metabolic activity and associated functional brain networks.

    PubMed

    Méndez-Couz, Marta; González-Pardo, Héctor; Vallejo, Guillermo; Arias, Jorge L; Conejo, Nélida M

    2016-10-01

    Previous studies showed the involvement of brain regions associated with both spatial learning and associative learning in spatial memory extinction, although the specific role of the dorsal and ventral hippocampus and the extended hippocampal system including the mammillary body in the process is still controversial. The present study aimed to identify the involvement of the dorsal and ventral hippocampus, together with cortical regions, the amygdaloid nuclei, and the mammillary bodies in the extinction of a spatial memory task. To address these issues, quantitative cytochrome c oxidase histochemistry was applied as a metabolic brain mapping method. Rats were trained in a reference memory task using the Morris water maze, followed by an extinction procedure of the previously acquired memory task. Results show that rats learned successfully the spatial memory task as shown by the progressive decrease in measured latencies to reach the escape platform and the results obtained in the probe test. Spatial memory was subsequently extinguished as shown by the descending preference for the previously reinforced location. A control naïve group was added to ensure that brain metabolic changes were specifically related with performance in the spatial memory extinction task. Extinction of the original spatial learning task significantly modified the metabolic activity in the dorsal and ventral hippocampus, the amygdala and the mammillary bodies. Moreover, the ventral hippocampus, the lateral mammillary body and the retrosplenial cortex were differentially recruited in the spatial memory extinction task, as shown by group differences in brain metabolic networks. These findings provide new insights on the brain regions and functional brain networks underlying spatial memory, and specifically spatial memory extinction. © 2016 Wiley Periodicals, Inc. PMID:27102086

  4. Oxysterols and Their Cellular Effectors

    PubMed Central

    Olkkonen, Vesa M.; Béaslas, Olivier; Nissilä, Eija

    2012-01-01

    Oxysterols are oxidized 27-carbon cholesterol derivatives or by-products of cholesterol biosynthesis, with a spectrum of biologic activities. Several oxysterols have cytotoxic and pro-apoptotic activities, the ability to interfere with the lateral domain organization, and packing of membrane lipids. These properties may account for their suggested roles in the pathology of diseases such as atherosclerosis, age-onset macular degeneration and Alzheimer’s disease. Oxysterols also have the capacity to induce inflammatory responses and play roles in cell differentiation processes. The functions of oxysterols as intermediates in the synthesis of bile acids and steroid hormones, and as readily transportable forms of sterol, are well established. Furthermore, their actions as endogenous regulators of gene expression in lipid metabolism via liver X receptors and the Insig (insulin-induced gene) proteins have been investigated in detail. The cytoplasmic oxysterol-binding protein (OSBP) homologues form a group of oxysterol/cholesterol sensors that has recently attracted a lot of attention. However, their mode of action is, as yet, poorly understood. Retinoic acid receptor-related orphan receptors (ROR) α and γ, and Epstein-Barr virus induced gene 2 (EBI2) have been identified as novel oxysterol receptors, revealing new physiologic oxysterol effector mechanisms in development, metabolism, and immunity, and evoking enhanced interest in these compounds in the field of biomedicine. PMID:24970128

  5. Physical activity and memory functions: are neurotrophins and cerebral gray matter volume the missing link?

    PubMed

    Flöel, A; Ruscheweyh, R; Krüger, K; Willemer, C; Winter, B; Völker, K; Lohmann, H; Zitzmann, M; Mooren, F; Breitenstein, C; Knecht, S

    2010-02-01

    Epidemiological studies reveal better cognitive function in physically active individuals. Possible mediators for this effect are neurotrophins, which are up-regulated through physical exercise and induce neuronal growth and synaptogenesis in the animal model. Here we cross-sectionally assessed 75 healthy older individuals for levels of physical activity, aerobic fitness, and memory encoding, as well as neurotrophin levels and cerebral gray matter volume. We found that physical activity, but not cardiovascular fitness, was associated with better memory encoding after controlling for age, sex, education, depression, alcohol consumption, and smoking. Higher levels of physical activity were associated with higher levels of the neurotrophin granulocyte colony stimulating factor (G-CSF) and increased cerebral gray matter volume in prefrontal and cingulate cortex as assessed by magnetic resonance voxel-based morphometry. While mediating factors will need to be further elucidated, these findings indicate that even low-level physical activity exerts beneficial effects on memory functions in older individuals.

  6. Activation of hippocampal nuclear factor-kappa B by retrieval is required for memory reconsolidation.

    PubMed

    Boccia, Mariano; Freudenthal, Ramiro; Blake, Mariano; de la Fuente, Veronica; Acosta, Gabriela; Baratti, Carlos; Romano, Arturo

    2007-12-01

    Initially, memory is labile and requires consolidation to become stable. However, several studies support that consolidated memories can undergo a new period of lability after retrieval. The mechanistic differences of this process, termed reconsolidation, with the consolidation process are under debate, including the participation of hippocampus. Up to this point, few reports describe molecular changes and, in particular, transcription factor (TF) involvement in memory restabilization. Increasing evidence supports the participation of the TF nuclear factor-kappaB (NF-kappaB) in memory consolidation. Here, we demonstrate that the inhibition of NF-kappaB after memory reactivation impairs retention of a hippocampal-dependent inhibitory avoidance task in mice. We used two independent disruptive strategies to reach this conclusion. First, we administered intracerebroventricular or intrahippocampal sulfasalazine, an inhibitor of IKK (IkappaB kinase), the kinase that activates NF-kappaB. Second, we infused intracerebroventricular or intrahippocampal kappaB decoy, a direct inhibitor of NF-kappaB consisting of a double-stranded DNA oligonucleotide that contains the kappaB consensus sequence. When injected immediately after memory retrieval, sulfasalazine or kappaB decoy (Decoy) impaired long-term retention. In contrast, a one base mutated kappaB decoy (mDecoy) had no effect. Furthermore, we also found NF-kappaB activation in the hippocampus, with a peak 15 min after memory retrieval. This activation was earlier than that found during consolidation. Together, these results indicate that NF-kappaB is an important transcriptional regulator in memory consolidation and reconsolidation in hippocampus, although the temporal kinetics of activation differs between the two processes.

  7. Contralateral Cortical Organisation of Information in Visual Short-Term Memory: Evidence from Lateralized Brain Activity during Retrieval

    ERIC Educational Resources Information Center

    Fortier-Gauthier, Ulysse; Moffat, Nicolas; Dell'Acqua, Robert; McDonald, John J.; Jolicoeur, Pierre

    2012-01-01

    We studied brain activity during retention and retrieval phases of two visual short-term memory (VSTM) experiments. Experiment 1 used a balanced memory array, with one color stimulus in each hemifield, followed by a retention interval and a central probe, at the fixation point that designated the target stimulus in memory about which to make a…

  8. Watching TV news as a memory task -- brain activation and age effects

    PubMed Central

    2010-01-01

    Background Neuroimaging studies which investigate brain activity underlying declarative memory processes typically use artificial, unimodal laboratory stimuli. In contrast, we developed a paradigm which much more closely approximates real-life situations of information encoding. Methods In this study, we tested whether ecologically valid stimuli - clips of a TV news show - are apt to assess memory-related fMRI activation in healthy participants across a wide age range (22-70 years). We contrasted brain responses during natural stimulation (TV news video clips) with a control condition (scrambled versions of the same clips with reversed audio tracks). After scanning, free recall performance was assessed. Results The memory task evoked robust activation of a left-lateralized network, including primarily lateral temporal cortex, frontal cortex, as well as the left hippocampus. Further analyses revealed that - when controlling for performance effects - older age was associated with greater activation of left temporal and right frontal cortex. Conclusion We demonstrate the feasibility of assessing brain activity underlying declarative memory using a natural stimulation paradigm with high ecological validity. The preliminary result of greater brain activation with increasing age might reflect an attempt to compensate for decreasing episodic memory capacity associated with aging. PMID:20738888

  9. Music improves verbal memory encoding while decreasing prefrontal cortex activity: an fNIRS study.

    PubMed

    Ferreri, Laura; Aucouturier, Jean-Julien; Muthalib, Makii; Bigand, Emmanuel; Bugaiska, Aurelia

    2013-01-01

    Listening to music engages the whole brain, thus stimulating cognitive performance in a range of non-purely musical activities such as language and memory tasks. This article addresses an ongoing debate on the link between music and memory for words. While evidence on healthy and clinical populations suggests that music listening can improve verbal memory in a variety of situations, it is still unclear what specific memory process is affected and how. This study was designed to explore the hypothesis that music specifically benefits the encoding part of verbal memory tasks, by providing a richer context for encoding and therefore less demand on the dorsolateral prefrontal cortex (DLPFC). Twenty-two healthy young adults were subjected to functional near-infrared spectroscopy (fNIRS) imaging of their bilateral DLPFC while encoding words in the presence of either a music or a silent background. Behavioral data confirmed the facilitating effect of music background during encoding on subsequent item recognition. fNIRS results revealed significantly greater activation of the left hemisphere during encoding (in line with the HERA model of memory lateralization) and a sustained, bilateral decrease of activity in the DLPFC in the music condition compared to silence. These findings suggest that music modulates the role played by the DLPFC during verbal encoding, and open perspectives for applications to clinical populations with prefrontal impairments, such as elderly adults or Alzheimer's patients. PMID:24339807

  10. Music improves verbal memory encoding while decreasing prefrontal cortex activity: an fNIRS study.

    PubMed

    Ferreri, Laura; Aucouturier, Jean-Julien; Muthalib, Makii; Bigand, Emmanuel; Bugaiska, Aurelia

    2013-01-01

    Listening to music engages the whole brain, thus stimulating cognitive performance in a range of non-purely musical activities such as language and memory tasks. This article addresses an ongoing debate on the link between music and memory for words. While evidence on healthy and clinical populations suggests that music listening can improve verbal memory in a variety of situations, it is still unclear what specific memory process is affected and how. This study was designed to explore the hypothesis that music specifically benefits the encoding part of verbal memory tasks, by providing a richer context for encoding and therefore less demand on the dorsolateral prefrontal cortex (DLPFC). Twenty-two healthy young adults were subjected to functional near-infrared spectroscopy (fNIRS) imaging of their bilateral DLPFC while encoding words in the presence of either a music or a silent background. Behavioral data confirmed the facilitating effect of music background during encoding on subsequent item recognition. fNIRS results revealed significantly greater activation of the left hemisphere during encoding (in line with the HERA model of memory lateralization) and a sustained, bilateral decrease of activity in the DLPFC in the music condition compared to silence. These findings suggest that music modulates the role played by the DLPFC during verbal encoding, and open perspectives for applications to clinical populations with prefrontal impairments, such as elderly adults or Alzheimer's patients.

  11. Music improves verbal memory encoding while decreasing prefrontal cortex activity: an fNIRS study

    PubMed Central

    Ferreri, Laura; Aucouturier, Jean-Julien; Muthalib, Makii; Bigand, Emmanuel; Bugaiska, Aurelia

    2013-01-01

    Listening to music engages the whole brain, thus stimulating cognitive performance in a range of non-purely musical activities such as language and memory tasks. This article addresses an ongoing debate on the link between music and memory for words. While evidence on healthy and clinical populations suggests that music listening can improve verbal memory in a variety of situations, it is still unclear what specific memory process is affected and how. This study was designed to explore the hypothesis that music specifically benefits the encoding part of verbal memory tasks, by providing a richer context for encoding and therefore less demand on the dorsolateral prefrontal cortex (DLPFC). Twenty-two healthy young adults were subjected to functional near-infrared spectroscopy (fNIRS) imaging of their bilateral DLPFC while encoding words in the presence of either a music or a silent background. Behavioral data confirmed the facilitating effect of music background during encoding on subsequent item recognition. fNIRS results revealed significantly greater activation of the left hemisphere during encoding (in line with the HERA model of memory lateralization) and a sustained, bilateral decrease of activity in the DLPFC in the music condition compared to silence. These findings suggest that music modulates the role played by the DLPFC during verbal encoding, and open perspectives for applications to clinical populations with prefrontal impairments, such as elderly adults or Alzheimer’s patients. PMID:24339807

  12. Target selection biases from recent experience transfer across effectors.

    PubMed

    Moher, Jeff; Song, Joo-Hyun

    2016-02-01

    Target selection is often biased by an observer's recent experiences. However, not much is known about whether these selection biases influence behavior across different effectors. For example, does looking at a red object make it easier to subsequently reach towards another red object? In the current study, we asked observers to find the uniquely colored target object on each trial. Randomly intermixed pre-trial cues indicated the mode of action: either an eye movement or a visually guided reach movement to the target. In Experiment 1, we found that priming of popout, reflected in faster responses following repetition of the target color on consecutive trials, occurred regardless of whether the effector was repeated from the previous trial or not. In Experiment 2, we examined whether an inhibitory selection bias away from a feature could transfer across effectors. While priming of popout reflects both enhancement of the repeated target features and suppression of the repeated distractor features, the distractor previewing effect isolates a purely inhibitory component of target selection in which a previewed color is presented in a homogenous display and subsequently inhibited. Much like priming of popout, intertrial suppression biases in the distractor previewing effect transferred across effectors. Together, these results suggest that biases for target selection driven by recent trial history transfer across effectors. This indicates that representations in memory that bias attention towards or away from specific features are largely independent from their associated actions. PMID:26563393

  13. DNA methyltransferase activity is required for memory-related neural plasticity in the lateral amygdala.

    PubMed

    Maddox, Stephanie A; Watts, Casey S; Schafe, Glenn E

    2014-01-01

    We have previously shown that auditory Pavlovian fear conditioning is associated with an increase in DNA methyltransferase (DNMT) expression in the lateral amygdala (LA) and that intra-LA infusion or bath application of an inhibitor of DNMT activity impairs the consolidation of an auditory fear memory and long-term potentiation (LTP) at thalamic and cortical inputs to the LA, in vitro. In the present study, we use awake behaving neurophysiological techniques to examine the role of DNMT activity in memory-related neurophysiological changes accompanying fear memory consolidation and reconsolidation in the LA, in vivo. We show that auditory fear conditioning results in a training-related enhancement in the amplitude of short-latency auditory-evoked field potentials (AEFPs) in the LA. Intra-LA infusion of a DNMT inhibitor impairs both fear memory consolidation and, in parallel, the consolidation of training-related neural plasticity in the LA; that is, short-term memory (STM) and short-term training-related increases in AEFP amplitude in the LA are intact, while long-term memory (LTM) and long-term retention of training-related increases in AEFP amplitudes are impaired. In separate experiments, we show that intra-LA infusion of a DNMT inhibitor following retrieval of an auditory fear memory has no effect on post-retrieval STM or short-term retention of training-related changes in AEFP amplitude in the LA, but significantly impairs both post-retrieval LTM and long-term retention of AEFP amplitude changes in the LA. These findings are the first to demonstrate the necessity of DNMT activity in the consolidation and reconsolidation of memory-associated neural plasticity, in vivo.

  14. Sound-Induced Activity in Voice-Sensitive Cortex Predicts Voice Memory Ability

    PubMed Central

    Watson, Rebecca; Latinus, Marianne; Bestelmeyer, Patricia E. G.; Crabbe, Frances; Belin, Pascal

    2012-01-01

    The “temporal voice areas” (TVAs; Belin et al., 2000) of the human brain show greater neuronal activity in response to human voices than to other categories of non-vocal sounds. However, a direct link between TVA activity and voice perception behavior has not yet been established. Here we show that a functional magnetic resonance imaging measure of activity in the TVAs predicts individual performance at a separately administered voice memory test. This relation holds when general sound memory ability is taken into account. These findings provide the first evidence that the TVAs are specifically involved in voice cognition. PMID:22485101

  15. The YopB protein of Yersinia pseudotuberculosis is essential for the translocation of Yop effector proteins across the target cell plasma membrane and displays a contact-dependent membrane disrupting activity.

    PubMed Central

    Håkansson, S; Schesser, K; Persson, C; Galyov, E E; Rosqvist, R; Homblé, F; Wolf-Watz, H

    1996-01-01

    During infection of cultured epithelial cells, surface-located Yersinia pseudotuberculosis deliver Yop (Yersinia outer protein) virulence factors into the cytoplasm of the target cell. A non-polar yopB mutant strain displays a wild-type phenotype with respect to in vitro Yop regulation and secretion but fails to elicit a cytotoxic response in cultured HeLa cells and is unable to inhibit phagocytosis by macrophage-like J774 cells. Additionally, the yopB mutant strain was avirulent in the mouse model. No YopE or YopH protein were observed within HeLa cells infected with the yopB mutant strain, suggesting that the loss of virulence of the mutant strain was due to its inability to translocate Yop effector proteins through the target cell plasma membrane. Expression of YopB is necessary for Yersinia-induced lysis of sheep erythrocytes. Purified YopB was shown to have membrane disruptive activity in vitro. YopB-dependent haemolytic activity required cell contact between the bacteria and the erythrocytes and could be inhibited by high, but not low, molecular weight carbohydrates. Similarly, expression of YopE reduced haemolytic activity. Therefore, we propose that YopB is essential for the formation of a pore in the target cell membrane that is required for the cell-to-cell transfer of Yop effector proteins. Images PMID:8918459

  16. PPARγ activation prevents impairments in spatial memory and neurogenesis following transient illness.

    PubMed

    Ormerod, Brandi K; Hanft, Simon J; Asokan, Aditya; Haditsch, Ursula; Lee, Star W; Palmer, Theo D

    2013-03-01

    The detrimental effects of illness on cognition are familiar to virtually everyone. Some effects resolve quickly while others may linger after the illness resolves. We found that a transient immune response stimulated by lipopolysaccharide (LPS) compromised hippocampal neurogenesis and impaired hippocampus-dependent spatial memory. The immune event caused an ∼50% reduction in the number of neurons generated during the illness and the onset of the memory impairment was delayed and coincided with the time when neurons generated during the illness would have become functional within the hippocampus. Broad spectrum non-steroidal anti-inflammatory drugs attenuated these effects but selective Cox-2 inhibition was ineffective while PPARγ activation was surprisingly effective at protecting both neurogenesis and memory from the effects of LPS-produced transient illness. These data may highlight novel mechanisms behind chronic inflammatory and neuroinflammatory episodes that are known to compromise hippocampus-dependent forms of learning and memory.

  17. PPARγ activation prevents impairments in spatial memory and neurogenesis following transient illness

    PubMed Central

    Ormerod, Brandi K.; Hanft, Simon J.; Asokan, Aditya; Haditsch, Ursula; Lee, Star W.; Palmer, Theo D.

    2012-01-01

    The detrimental effects of illness on cognition are familiar to virtually everyone. Some effects resolve quickly while others may linger after the illness resolves. We found that a transient immune response stimulated by lipopolysaccharide (LPS) compromised hippocampal neurogenesis and impaired hippocampus-dependent spatial memory. The immune event caused a 50% reduction in the number of neurons generated during the illness and the onset of the memory impairment was delayed and coincided with the time when neurons generated during the illness would have become functional within the hippocampus. Broad spectrum non-steroidal anti-inflammatory drugs attenuated these effects but selective Cox-2 inhibition was ineffective while PPARγ activation was surprisingly effective at protecting both neurogenesis and memory from the effects of LPS-produced transient illness. These data may highlight novel mechanisms behind chronic inflammatory and neuroinflammatory episodes that are known to compromise hippocampus-dependent forms of learning and memory. PMID:23108061

  18. Hippocampal PER1: a circadian sentinel controlling RSKy activity during memory formation.

    PubMed

    Yoo, Seung-Hee; Eckel-Mahan, Kristin

    2016-09-01

    Studies have demonstrated a pronounced dependence of memory formation on circadian time; however, the numerous mechanisms underlying this reliance are only beginning to be understood. While the 24-h cellular clock controls various aspects of hippocampal memory formation, its consolidation in particular (i.e., its conversion from short-term to long-term memory), appears to be heavily dependent on circadian activity in hippocampal neurons. Hippocampal memory consolidation requires phosphorylation of the cAMP Response Element-Binding protein, CREB, which upon phosphorylation promotes the transcription of genes necessary for long-term memory formation. Rhythmic cAMP/ERK-MAPK activity upstream of CREB is a necessary component. This Editorial highlights a study by Rawashdeh and coworkers, in which the authors establish the circadian clock gene Period1 (Per1) as a regulator of CREB phosphorylation in the mouse hippocampus, and thus reveal a functional link between circadian rhythms and learning efficiency. Read the highlighted article 'Period1 gates the circadian modulation of memory-relevant signaling in mouse hippocampus by regulating the nuclear shuttling of the CREB kinase pP90RSK' on page 731. PMID:27554418

  19. Non-volatile memory devices with redox-active diruthenium molecular compound.

    PubMed

    Pookpanratana, S; Zhu, H; Bittle, E G; Natoli, S N; Ren, T; Richter, C A; Li, Q; Hacker, C A

    2016-03-01

    Reduction-oxidation (redox) active molecules hold potential for memory devices due to their many unique properties. We report the use of a novel diruthenium-based redox molecule incorporated into a non-volatile Flash-based memory device architecture. The memory capacitor device structure consists of a Pd/Al2O3/molecule/SiO2/Si structure. The bulky ruthenium redox molecule is attached to the surface by using a 'click' reaction and the monolayer structure is characterized by x-ray photoelectron spectroscopy to verify the Ru attachment and molecular density. The 'click' reaction is particularly advantageous for memory applications because of (1) ease of chemical design and synthesis, and (2) provides an additional spatial barrier between the oxide/silicon to the diruthenium molecule. Ultraviolet photoelectron spectroscopy data identified the energy of the electronic levels of the surface before and after surface modification. The molecular memory devices display an unsaturated charge storage window attributed to the intrinsic properties of the redox-active molecule. Our findings demonstrate the strengths and challenges with integrating molecular layers within solid-state devices, which will influence the future design of molecular memory devices.

  20. Non-volatile memory devices with redox-active diruthenium molecular compound

    NASA Astrophysics Data System (ADS)

    Pookpanratana, S.; Zhu, H.; Bittle, E. G.; Natoli, S. N.; Ren, T.; Richter, C. A.; Li, Q.; Hacker, C. A.

    2016-03-01

    Reduction-oxidation (redox) active molecules hold potential for memory devices due to their many unique properties. We report the use of a novel diruthenium-based redox molecule incorporated into a non-volatile Flash-based memory device architecture. The memory capacitor device structure consists of a Pd/Al2O3/molecule/SiO2/Si structure. The bulky ruthenium redox molecule is attached to the surface by using a ‘click’ reaction and the monolayer structure is characterized by x-ray photoelectron spectroscopy to verify the Ru attachment and molecular density. The ‘click’ reaction is particularly advantageous for memory applications because of (1) ease of chemical design and synthesis, and (2) provides an additional spatial barrier between the oxide/silicon to the diruthenium molecule. Ultraviolet photoelectron spectroscopy data identified the energy of the electronic levels of the surface before and after surface modification. The molecular memory devices display an unsaturated charge storage window attributed to the intrinsic properties of the redox-active molecule. Our findings demonstrate the strengths and challenges with integrating molecular layers within solid-state devices, which will influence the future design of molecular memory devices.

  1. Patterns of Brain-Electrical Activity during Declarative Memory Performance in 10-Month-Old Infants

    ERIC Educational Resources Information Center

    Morasch, Katherine C.; Bell, Martha Ann

    2009-01-01

    This study of infant declarative memory concurrently examined brain-electrical activity and deferred imitation performance in 10-month-old infants. Continuous electroencephalogram (EEG) measures were collected throughout the activity-matched baseline, encoding (modeling) and retrieval (delayed test) phases of a within-subjects deferred imitation…

  2. Memory retrieval of inhibitory avoidance requires histamine H1 receptor activation in the hippocampus.

    PubMed

    Fabbri, Roberta; Furini, Cristiane Regina Guerino; Passani, Maria Beatrice; Provensi, Gustavo; Baldi, Elisabetta; Bucherelli, Corrado; Izquierdo, Ivan; de Carvalho Myskiw, Jociane; Blandina, Patrizio

    2016-05-10

    Retrieval represents a dynamic process that may require neuromodulatory signaling. Here, we report that the integrity of the brain histaminergic system is necessary for retrieval of inhibitory avoidance (IA) memory, because rats depleted of histamine through lateral ventricle injections of α-fluoromethylhistidine (a-FMHis), a suicide inhibitor of histidine decarboxylase, displayed impaired IA memory when tested 2 d after training. a-FMHis was administered 24 h after training, when IA memory trace was already formed. Infusion of histamine in hippocampal CA1 of brain histamine-depleted rats (hence, amnesic) 10 min before the retention test restored IA memory but was ineffective when given in the basolateral amygdala (BLA) or the ventral medial prefrontal cortex (vmPFC). Intra-CA1 injections of selective H1 and H2 receptor agonists showed that histamine exerted its effect by activating the H1 receptor. Noteworthy, the H1 receptor antagonist pyrilamine disrupted IA memory retrieval in rats, thus strongly supporting an active involvement of endogenous histamine; 90 min after the retention test, c-Fos-positive neurons were significantly fewer in the CA1s of a-FMHis-treated rats that displayed amnesia compared with in the control group. We also found reduced levels of phosphorylated cAMP-responsive element binding protein (pCREB) in the CA1s of a-FMHis-treated animals compared with in controls. Increases in pCREB levels are associated with retrieval of associated memories. Targeting the histaminergic system may modify the retrieval of emotional memory; hence, histaminergic ligands might reduce dysfunctional aversive memories and improve the efficacy of exposure psychotherapies.

  3. Active, passive and snapshot exploration in a virtual environment: influence on scene memory, reorientation and path memory.

    PubMed

    Gaunet, F; Vidal, M; Kemeny, A; Berthoz, A

    2001-06-01

    We investigated the importance of active, passive and snapshot exploration on spatial memory in a virtual city. The exploration consisted in traveling along a series of streets. 'Active exploration' was performed by giving directions to the subject who controlled his displacement with a joystick. During 'passive' exploration, the travel was imposed by the computer. Finally, during 'snapshot exploration', simple views of the scene were presented sequentially every 4 m. Travel velocity was the same in all cases. The three visual exploration modes were compared with three spatial memory measures: (1) scene recognition, (2) at the end of the path, reorientation toward the departure point and (3) drawings of the path shape. Scene recognition and estimation of the direction of the starting point of the path were not affected by the mode of exploration. In contrast, reproduction of the shape of the path was affected: the errors of reproduction were greater for the snapshot exploration than for the other two conditions; there was no difference between the other two conditions. These results suggest that (1) 2D image features from a visual scene are memorized. Moreover, (2) pointing towards the origin of the path relies on motion duration integration or a frame of reference integrated during displacement. Finally, (3) drawing the path shape involves a deliberate reconstruction process.

  4. Memory Maintenance in Synapses with Calcium-Based Plasticity in the Presence of Background Activity

    PubMed Central

    Higgins, David; Graupner, Michael; Brunel, Nicolas

    2014-01-01

    Most models of learning and memory assume that memories are maintained in neuronal circuits by persistent synaptic modifications induced by specific patterns of pre- and postsynaptic activity. For this scenario to be viable, synaptic modifications must survive the ubiquitous ongoing activity present in neural circuits in vivo. In this paper, we investigate the time scales of memory maintenance in a calcium-based synaptic plasticity model that has been shown recently to be able to fit different experimental data-sets from hippocampal and neocortical preparations. We find that in the presence of background activity on the order of 1 Hz parameters that fit pyramidal layer 5 neocortical data lead to a very fast decay of synaptic efficacy, with time scales of minutes. We then identify two ways in which this memory time scale can be extended: (i) the extracellular calcium concentration in the experiments used to fit the model are larger than estimated concentrations in vivo. Lowering extracellular calcium concentration to in vivo levels leads to an increase in memory time scales of several orders of magnitude; (ii) adding a bistability mechanism so that each synapse has two stable states at sufficiently low background activity leads to a further boost in memory time scale, since memory decay is no longer described by an exponential decay from an initial state, but by an escape from a potential well. We argue that both features are expected to be present in synapses in vivo. These results are obtained first in a single synapse connecting two independent Poisson neurons, and then in simulations of a large network of excitatory and inhibitory integrate-and-fire neurons. Our results emphasise the need for studying plasticity at physiological extracellular calcium concentration, and highlight the role of synaptic bi- or multistability in the stability of learned synaptic structures. PMID:25275319

  5. Impact of lead sub-chronic toxicity on recognition memory and motor activity of Wistar rat.

    PubMed

    Azzaoui, F Z; Ahami, A O T; Khadmaoui, A

    2009-01-15

    The aim of this research was to investigate the impact of lead nitrate administered in drinking water during 90 days (sub-chronic toxicity), on body weight gain, motor activity, brain lead accumulation and especially on recognition memory of Wistar rats. Two groups of young female Wistar rats were used. Treated rats received 20 mg L(-1) of lead nitrate diluted in drinking water, while control rats received drinking water only, for 3 months. An evolution of body weight, motor activity, object recognition memory and measure of brain lead levels has been evaluated. The body weight was taken weekly, whereas the memory abilities and the motor activity are measured once every fortnight alternatively, by submitting rats to the Open Field (OF) test and to the Novel Object Recognizing (NOR) memory test. The results have shown a non significant effect in gain of body weight. However, a high significance was shown for horizontal activity (p<0.01), long memory term (p<0.01), at the end of testing period and for brain lead levels (p<0.05) between studied groups.

  6. Long-term memory consolidation depends on proteasome activity in the crab Chasmagnathus.

    PubMed

    Merlo, E; Romano, A

    2007-06-15

    Long-term memory formation depends on protein and mRNA synthesis that subserves synaptic reorganization. The removal of pre-existing inhibitory proteins by the ubiquitin-proteasome system (UPS) is proposed as a crucial step to support these modifications. The activation of the constitutive transcription factor nuclear factor kappaB (NF-kappaB) depends on the degradation of the inhibitor of NF-kappaB (IkappaB) by the UPS. Here we study the effect of a UPS inhibitor, MG132, on long-term memory consolidation and NF-kappaB activation in the learning paradigm of the crab Chasmagnathus, a model in which this transcription factor plays a key role. Here we found that administration of MG132 interferes with long-term memory but not with short-term memory, and no facilitatory effects were found. Then we studied the effect of the UPS inhibitor on NF-kappaB pathway, finding that MG132 blocks the activation of NF-kappaB induced by training. These results suggest that the UPS is necessary for long-term memory consolidation, allowing for the activation of NF-kappaB as one of the target molecular pathways.

  7. Dissociated sequential activity and stimulus encoding in the dorsomedial striatum during spatial working memory

    PubMed Central

    Akhlaghpour, Hessameddin; Wiskerke, Joost; Choi, Jung Yoon; Taliaferro, Joshua P; Au, Jennifer; Witten, Ilana B

    2016-01-01

    Several lines of evidence suggest that the striatum has an important role in spatial working memory. The neural dynamics in the striatum have been described in tasks with short delay periods (1–4 s), but remain largely uncharacterized for tasks with longer delay periods. We collected and analyzed single unit recordings from the dorsomedial striatum of rats performing a spatial working memory task with delays up to 10 s. We found that neurons were activated sequentially, with the sequences spanning the entire delay period. Surprisingly, this sequential activity was dissociated from stimulus encoding activity, which was present in the same neurons, but preferentially appeared towards the onset of the delay period. These observations contrast with descriptions of sequential dynamics during similar tasks in other brains areas, and clarify the contribution of the striatum to spatial working memory. DOI: http://dx.doi.org/10.7554/eLife.19507.001 PMID:27636864

  8. Fencing direct memory access data transfers in a parallel active messaging interface of a parallel computer

    DOEpatents

    Blocksome, Michael A; Mamidala, Amith R

    2014-02-11

    Fencing direct memory access (`DMA`) data transfers in a parallel active messaging interface (`PAMI`) of a parallel computer, the PAMI including data communications endpoints, each endpoint including specifications of a client, a context, and a task, the endpoints coupled for data communications through the PAMI and through DMA controllers operatively coupled to segments of shared random access memory through which the DMA controllers deliver data communications deterministically, including initiating execution through the PAMI of an ordered sequence of active DMA instructions for DMA data transfers between two endpoints, effecting deterministic DMA data transfers through a DMA controller and a segment of shared memory; and executing through the PAMI, with no FENCE accounting for DMA data transfers, an active FENCE instruction, the FENCE instruction completing execution only after completion of all DMA instructions initiated prior to execution of the FENCE instruction for DMA data transfers between the two endpoints.

  9. Fencing direct memory access data transfers in a parallel active messaging interface of a parallel computer

    DOEpatents

    Blocksome, Michael A.; Mamidala, Amith R.

    2013-09-03

    Fencing direct memory access (`DMA`) data transfers in a parallel active messaging interface (`PAMI`) of a parallel computer, the PAMI including data communications endpoints, each endpoint including specifications of a client, a context, and a task, the endpoints coupled for data communications through the PAMI and through DMA controllers operatively coupled to segments of shared random access memory through which the DMA controllers deliver data communications deterministically, including initiating execution through the PAMI of an ordered sequence of active DMA instructions for DMA data transfers between two endpoints, effecting deterministic DMA data transfers through a DMA controller and a segment of shared memory; and executing through the PAMI, with no FENCE accounting for DMA data transfers, an active FENCE instruction, the FENCE instruction completing execution only after completion of all DMA instructions initiated prior to execution of the FENCE instruction for DMA data transfers between the two endpoints.

  10. Rheb Protein Binds CAD (Carbamoyl-phosphate Synthetase 2, Aspartate Transcarbamoylase, and Dihydroorotase) Protein in a GTP- and Effector Domain-dependent Manner and Influences Its Cellular Localization and Carbamoyl-phosphate Synthetase (CPSase) Activity*

    PubMed Central

    Sato, Tatsuhiro; Akasu, Hitomi; Shimono, Wataru; Matsu, Chisa; Fujiwara, Yuki; Shibagaki, Yoshio; Heard, Jeffrey J.; Tamanoi, Fuyuhiko; Hattori, Seisuke

    2015-01-01

    Rheb small GTPases, which consist of Rheb1 and Rheb2 (also known as RhebL1) in mammalian cells, are unique members of the Ras superfamily and play central roles in regulating protein synthesis and cell growth by activating mTOR. To gain further insight into the function of Rheb, we carried out a search for Rheb-binding proteins and found that Rheb binds to CAD protein (carbamoyl-phosphate synthetase 2, aspartate transcarbamoylase, and dihydroorotase), a multifunctional enzyme required for the de novo synthesis of pyrimidine nucleotides. CAD binding is more pronounced with Rheb2 than with Rheb1. Rheb binds CAD in a GTP- and effector domain-dependent manner. The region of CAD where Rheb binds is located at the C-terminal region of the carbamoyl-phosphate synthetase domain and not in the dihydroorotase and aspartate transcarbamoylase domains. Rheb stimulated carbamoyl-phosphate synthetase activity of CAD in vitro. In addition, an elevated level of intracellular UTP pyrimidine nucleotide was observed in Tsc2-deficient cells, which was attenuated by knocking down of Rheb. Immunostaining analysis showed that expression of Rheb leads to increased accumulation of CAD on lysosomes. Both a farnesyltransferase inhibitor that blocks membrane association of Rheb and knockdown of Rheb mislocalized CAD. These results establish CAD as a downstream effector of Rheb and suggest a possible role of Rheb in regulating de novo pyrimidine nucleotide synthesis. PMID:25422319

  11. Auditory short-term memory activation during score reading.

    PubMed

    Simoens, Veerle L; Tervaniemi, Mari

    2013-01-01

    Performing music on the basis of reading a score requires reading ahead of what is being played in order to anticipate the necessary actions to produce the notes. Score reading thus not only involves the decoding of a visual score and the comparison to the auditory feedback, but also short-term storage of the musical information due to the delay of the auditory feedback during reading ahead. This study investigates the mechanisms of encoding of musical information in short-term memory during such a complicated procedure. There were three parts in this study. First, professional musicians participated in an electroencephalographic (EEG) experiment to study the slow wave potentials during a time interval of short-term memory storage in a situation that requires cross-modal translation and short-term storage of visual material to be compared with delayed auditory material, as it is the case in music score reading. This delayed visual-to-auditory matching task was compared with delayed visual-visual and auditory-auditory matching tasks in terms of EEG topography and voltage amplitudes. Second, an additional behavioural experiment was performed to determine which type of distractor would be the most interfering with the score reading-like task. Third, the self-reported strategies of the participants were also analyzed. All three parts of this study point towards the same conclusion according to which during music score reading, the musician most likely first translates the visual score into an auditory cue, probably starting around 700 or 1300 ms, ready for storage and delayed comparison with the auditory feedback.

  12. Human intracranial high-frequency activity maps episodic memory formation in space and time.

    PubMed

    Burke, John F; Long, Nicole M; Zaghloul, Kareem A; Sharan, Ashwini D; Sperling, Michael R; Kahana, Michael J

    2014-01-15

    Noninvasive neuroimaging studies have revealed a network of brain regions that activate during human memory encoding; however, the relative timing of such activations remains unknown. Here we used intracranially recorded high-frequency activity (HFA) to first identify regions that activate during successful encoding. Then, we leveraged the high-temporal precision of HFA to investigate the timing of such activations. We found that memory encoding invokes two spatiotemporally distinct activations: early increases in HFA that involve the ventral visual pathway as well as the medial temporal lobe and late increases in HFA that involve the left inferior frontal gyrus, left posterior parietal cortex, and left ventrolateral temporal cortex. We speculate that these activations reflect higher-order visual processing and top-down modulation of attention/semantic information, respectively.

  13. Photo-induced optical activity in phase-change memory materials.

    PubMed

    Borisenko, Konstantin B; Shanmugam, Janaki; Williams, Benjamin A O; Ewart, Paul; Gholipour, Behrad; Hewak, Daniel W; Hussain, Rohanah; Jávorfi, Tamás; Siligardi, Giuliano; Kirkland, Angus I

    2015-03-05

    We demonstrate that optical activity in amorphous isotropic thin films of pure Ge2Sb2Te5 and N-doped Ge2Sb2Te5N phase-change memory materials can be induced using rapid photo crystallisation with circularly polarised laser light. The new anisotropic phase transition has been confirmed by circular dichroism measurements. This opens up the possibility of controlled induction of optical activity at the nanosecond time scale for exploitation in a new generation of high-density optical memory, fast chiroptical switches and chiral metamaterials.

  14. Cerebral Activity Associated with Transient Sleep-Facilitated Reduction in Motor Memory Vulnerability to Interference

    PubMed Central

    Albouy, Geneviève; King, Bradley R.; Schmidt, Christina; Desseilles, Martin; Dang-Vu, Thien Thanh; Balteau, Evelyne; Phillips, Christophe; Degueldre, Christian; Orban, Pierre; Benali, Habib; Peigneux, Philippe; Luxen, André; Karni, Avi; Doyon, Julien; Maquet, Pierre; Korman, Maria

    2016-01-01

    Motor memory consolidation is characterized, in part, by a sleep-facilitated decrease in susceptibility to subsequent interfering experiences. Surprisingly, the cerebral substrates supporting this phenomenon have never been examined. We used fMRI to investigate the neural correlates of the influence of sleep on interference to motor memory consolidation. Healthy young adults were trained on a sequential motor task, and subsequently practiced a second competing sequence after an interval including diurnal sleep or wakefulness. Participants were then retested on the initial sequence 8 h and 24 h (including nocturnal sleep) after training. Results demonstrated that a post-training nap significantly protected memory against interference at 8 h and modulated the link between cerebral activity and behavior, such that a smaller post-interference decrease in cortico-striatal activity was associated with better performance. Interestingly, the protective effect of a nap was only transitory, as both groups performed similarly at 24 h. Activity in cortico-striatal areas that was disrupted during the day, presumably due to interference and accentuated in the absence of a nap, was restored overnight. Altogether, our findings offer the first evidence that cortico-striatal areas play a critical role in the transient sleep-facilitated reduction in motor memory vulnerability and in the overnight restoration of previously degraded memories. PMID:27725727

  15. Medial temporal lobe activity associated with the successful retrieval of destination memory.

    PubMed

    Mugikura, Shunji; Abe, Nobuhito; Ito, Ayahito; Kawasaki, Iori; Ueno, Aya; Takahashi, Shoki; Fujii, Toshikatsu

    2016-01-01

    Destination memory is the process of remembering to whom we tell particular things. Although recent behavioral studies have clarified the cognitive nature of destination memory, the neural mechanisms underlying destination memory retrieval remain unclear. We used functional magnetic resonance imaging (fMRI) to determine whether the medial temporal lobe (MTL), a structure that has been implicated in recollection-based memory, is activated during the successful retrieval of destination information. During a study phase before fMRI scanning, the subjects told a series of facts to either a woman or a man. During fMRI scanning, the subjects were asked to judge whether each fact presented was old or new, and if they judged it as old, to indicate, including a confidence rating (high or low), whether the subjects had told that fact to either a man or a woman. We found that successful destination retrieval, when compared to failed destination retrieval, was associated with increased activity in the parahippocampal gyrus. We also found that the confidence level (high vs. low) for destination memory retrieval was associated with increased activity in another (posterior) region of the parahippocampal gyrus. The present study suggests that the successful retrieval of destination information depends highly on MTL-mediated recollection processes. PMID:26378005

  16. Medial temporal lobe activity associated with the successful retrieval of destination memory.

    PubMed

    Mugikura, Shunji; Abe, Nobuhito; Ito, Ayahito; Kawasaki, Iori; Ueno, Aya; Takahashi, Shoki; Fujii, Toshikatsu

    2016-01-01

    Destination memory is the process of remembering to whom we tell particular things. Although recent behavioral studies have clarified the cognitive nature of destination memory, the neural mechanisms underlying destination memory retrieval remain unclear. We used functional magnetic resonance imaging (fMRI) to determine whether the medial temporal lobe (MTL), a structure that has been implicated in recollection-based memory, is activated during the successful retrieval of destination information. During a study phase before fMRI scanning, the subjects told a series of facts to either a woman or a man. During fMRI scanning, the subjects were asked to judge whether each fact presented was old or new, and if they judged it as old, to indicate, including a confidence rating (high or low), whether the subjects had told that fact to either a man or a woman. We found that successful destination retrieval, when compared to failed destination retrieval, was associated with increased activity in the parahippocampal gyrus. We also found that the confidence level (high vs. low) for destination memory retrieval was associated with increased activity in another (posterior) region of the parahippocampal gyrus. The present study suggests that the successful retrieval of destination information depends highly on MTL-mediated recollection processes.

  17. Behind the lines–actions of bacterial type III effector proteins in plant cells

    PubMed Central

    Büttner, Daniela

    2016-01-01

    Pathogenicity of most Gram-negative plant-pathogenic bacteria depends on the type III secretion (T3S) system, which translocates bacterial effector proteins into plant cells. Type III effectors modulate plant cellular pathways to the benefit of the pathogen and promote bacterial multiplication. One major virulence function of type III effectors is the suppression of plant innate immunity, which is triggered upon recognition of pathogen-derived molecular patterns by plant receptor proteins. Type III effectors also interfere with additional plant cellular processes including proteasome-dependent protein degradation, phytohormone signaling, the formation of the cytoskeleton, vesicle transport and gene expression. This review summarizes our current knowledge on the molecular functions of type III effector proteins with known plant target molecules. Furthermore, plant defense strategies for the detection of effector protein activities or effector-triggered alterations in plant targets are discussed. PMID:27526699

  18. Modeling the effector - regulatory T cell cross-regulation reveals the intrinsic character of relapses in Multiple Sclerosis

    PubMed Central

    2011-01-01

    Background The relapsing-remitting dynamics is a hallmark of autoimmune diseases such as Multiple Sclerosis (MS). Although current understanding of both cellular and molecular mechanisms involved in the pathogenesis of autoimmune diseases is significant, how their activity generates this prototypical dynamics is not understood yet. In order to gain insight about the mechanisms that drive these relapsing-remitting dynamics, we developed a computational model using such biological knowledge. We hypothesized that the relapsing dynamics in autoimmunity can arise through the failure in the mechanisms controlling cross-regulation between regulatory and effector T cells with the interplay of stochastic events (e.g. failure in central tolerance, activation by pathogens) that are able to trigger the immune system. Results The model represents five concepts: central tolerance (T-cell generation by the thymus), T-cell activation, T-cell memory, cross-regulation (negative feedback) between regulatory and effector T-cells and tissue damage. We enriched the model with reversible and irreversible tissue damage, which aims to provide a comprehensible link between autoimmune activity and clinical relapses and active lesions in the magnetic resonances studies in patients with Multiple Sclerosis. Our analysis shows that the weakness in this negative feedback between effector and regulatory T-cells, allows the immune system to generate the characteristic relapsing-remitting dynamics of autoimmune diseases, without the need of additional environmental triggers. The simulations show that the timing at which relapses appear is highly unpredictable. We also introduced targeted perturbations into the model that mimicked immunotherapies that modulate effector and regulatory populations. The effects of such therapies happened to be highly dependent on the timing and/or dose, and on the underlying dynamic of the immune system. Conclusion The relapsing dynamic in MS derives from the emergent

  19. Pre-stimulus BOLD-network activation modulates EEG spectral activity during working memory retention.

    PubMed

    Kottlow, Mara; Schlaepfer, Anthony; Baenninger, Anja; Michels, Lars; Brandeis, Daniel; Koenig, Thomas

    2015-01-01

    Working memory (WM) processes depend on our momentary mental state and therefore exhibit considerable fluctuations. Here, we investigate the interplay of task-preparatory and task-related brain activity as represented by pre-stimulus BOLD-fluctuations and spectral EEG from the retention periods of a visual WM task. Visual WM is used to maintain sensory information in the brain enabling the performance of cognitive operations and is associated with mental health. We tested 22 subjects simultaneously with EEG and fMRI while performing a visuo-verbal Sternberg task with two different loads, allowing for the temporal separation of preparation, encoding, retention and retrieval periods. Four temporally coherent networks (TCNs)-the default mode network (DMN), the dorsal attention, the right and the left WM network-were extracted from the continuous BOLD data by means of a group ICA. Subsequently, the modulatory effect of these networks' pre-stimulus activation upon retention-related EEG activity in the theta, alpha, and beta frequencies was analyzed. The obtained results are informative in the context of state-dependent information processing. We were able to replicate two well-known load-dependent effects: the frontal-midline theta increase during the task and the decrease of pre-stimulus DMN activity. As our main finding, these two measures seem to depend on each other as the significant negative correlations at frontal-midline channels suggested. Thus, suppressed pre-stimulus DMN levels facilitated later task related frontal midline theta increases. In general, based on previous findings that neuronal coupling in different frequency bands may underlie distinct functions in WM retention, our results suggest that processes reflected by spectral oscillations during retention seem not only to be "online" synchronized with activity in different attention-related networks but are also modulated by activity in these networks during preparation intervals. PMID:25999828

  20. CA1 hippocampal network activity changes during sleep-dependent memory consolidation

    PubMed Central

    Ognjanovski, Nicolette; Maruyama, Daniel; Lashner, Nora; Zochowski, Michal; Aton, Sara J.

    2014-01-01

    A period of sleep over the first few hours following single-trial contextual fear conditioning (CFC) is essential for hippocampally-mediated memory consolidation. Recent studies have uncovered intracellular mechanisms required for memory formation which are affected by post-conditioning sleep and sleep deprivation. However, almost nothing is known about the circuit-level activity changes during sleep that underlie activation of these intracellular pathways. Here we continuously recorded from the CA1 region of freely-behaving mice to characterize neuronal and network activity changes occurring during active memory consolidation. C57BL/6J mice were implanted with custom stereotrode recording arrays to monitor activity of individual CA1 neurons, local field potentials (LFPs), and electromyographic activity. Sleep architecture and state-specific CA1 activity patterns were assessed during a 24 h baseline recording period, and for 24 h following either single-trial CFC or Sham conditioning. We find that consolidation of CFC is not associated with significant sleep architecture changes, but is accompanied by long-lasting increases in CA1 neuronal firing, as well as increases in delta, theta, and gamma-frequency CA1 LFP activity. These changes occurred in both sleep and wakefulness, and may drive synaptic plasticity within the hippocampus during memory formation. We also find that functional connectivity within the CA1 network, assessed through functional clustering algorithm (FCA) analysis of spike timing relationships among recorded neurons, becomes more stable during consolidation of CFC. This increase in network stability was not present following Sham conditioning, was most evident during post-CFC slow wave sleep (SWS), and was negligible during post-CFC wakefulness. Thus in the interval between encoding and recall, SWS may stabilize the hippocampal contextual fear memory (CFM) trace by promoting CA1 network stability. PMID:24860440

  1. The wick in the candle of learning: epistemic curiosity activates reward circuitry and enhances memory.

    PubMed

    Kang, Min Jeong; Hsu, Ming; Krajbich, Ian M; Loewenstein, George; McClure, Samuel M; Wang, Joseph Tao-yi; Camerer, Colin F

    2009-08-01

    Curiosity has been described as a desire for learning and knowledge, but its underlying mechanisms are not well understood. We scanned subjects with functional magnetic resonance imaging while they read trivia questions. The level of curiosity when reading questions was correlated with activity in caudate regions previously suggested to be involved in anticipated reward. This finding led to a behavioral study, which showed that subjects spent more scarce resources (either limited tokens or waiting time) to find out answers when they were more curious. The functional imaging also showed that curiosity increased activity in memory areas when subjects guessed incorrectly, which suggests that curiosity may enhance memory for surprising new information. This prediction about memory enhancement was confirmed in a behavioral study: Higher curiosity in an initial session was correlated with better recall of surprising answers 1 to 2 weeks later.

  2. Age differences in medial prefrontal activity for subsequent memory of truth value

    PubMed Central

    Cassidy, Brittany S.; Hedden, Trey; Yoon, Carolyn; Gutchess, Angela H.

    2014-01-01

    Much research has demonstrated that aging is marked by decreased source memory relative to young adults, yet a smaller body of work has demonstrated that increasing the socioemotional content of source information may be one way to reduce age-related performance differences. Although dorsomedial prefrontal cortex (dmPFC) activity may support source memory among young and older adults, the extent to which one activates dorsal vs. ventral mPFC may reflect one's personal connection with incoming information. Because truth value may be one salient marker that impacts one's connection with information and allocation of attention toward incoming material, we investigated whether the perceived truth value of information differently impacts differences in mPFC activity associated with encoding source information, particularly with age. Twelve young (18–23 years) and 12 older adults (63–80 years) encoded true and false statements. Behavioral results showed similar memory performance between the age groups. With respect to neural activity associated with subsequent memory, young adults, relative to older adults, exhibited greater activity in dmPFC while older adults displayed enhanced ventromedial prefrontal cortex (vmPFC) and insula engagement relative to young. These results may potentially indicate that young adults focus on a general knowledge acquisition goal, while older adults focus on emotionally relevant aspects of the material. The findings demonstrate that age-related differences in recruitment of mPFC associated with encoding source information may in some circumstances underlie age-equivalent behavioral performance. PMID:24570672

  3. Optogenetic activation of presynaptic inputs in lateral amygdala forms associative fear memory.

    PubMed

    Kwon, Jeong-Tae; Nakajima, Ryuichi; Kim, Hyung-Su; Jeong, Yire; Augustine, George J; Han, Jin-Hee

    2014-11-01

    In Pavlovian fear conditioning, the lateral amygdala (LA) has been highlighted as a key brain site for association between sensory cues and aversive stimuli. However, learning-related changes are also found in upstream sensory regions such as thalamus and cortex. To isolate the essential neural circuit components for fear memory association, we tested whether direct activation of presynaptic sensory inputs in LA, without the participation of upstream activity, is sufficient to form fear memory in mice. Photostimulation of axonal projections from the two main auditory brain regions, the medial geniculate nucleus of the thalamus and the secondary auditory cortex, was paired with aversive footshock. Twenty-four hours later the same photostimulation induced robust conditioned freezing and this fear memory formation was disrupted when glutamatergic synaptic transmission was locally blocked in the LA. Therefore, our results prove for the first time that synapses between sensory input areas and the LA, previously implicated as a crucial brain site for fear memory formation, actually are sufficient to serve as a conditioned stimulus. Our results strongly support the idea that the LA may be sufficient to encode and store associations between neutral cue and aversive stimuli during natural fear conditioning as a critical part of a broad fear memory engram.

  4. Uncovering Camouflage: Amygdala Activation Predicts Long-Term Memory of Induced Perceptual Insight

    PubMed Central

    Ludmer, Rachel; Dudai, Yadin; Rubin, Nava

    2012-01-01

    What brain mechanisms underlie learning of new knowledge from single events? We studied encoding in long-term memory of a unique type of one-shot experience, induced perceptual insight. While undergoing an fMRI brain scan, participants viewed degraded images of real-world pictures where the underlying objects were hard to recognize (‘camouflage’), followed by brief exposures to the original images (‘solution’), which led to induced insight (“Aha!”). A week later, participants’ memory was tested; a solution image was classified as ‘remembered’ if detailed perceptual knowledge was elicited from the camouflage image alone. During encoding, subsequently remembered images enjoyed higher activity in mid-level visual cortex and medial frontal cortex, but most pronouncedly in the amygdala, whose activity could be used to predict which solutions will remain in long-term memory. Our findings extend the known roles of amygdala in memory to include promoting of long-term memory of the sudden reorganization of internal representations. PMID:21382558

  5. Cross-modal activation of auditory regions during visuo-spatial working memory in early deafness.

    PubMed

    Ding, Hao; Qin, Wen; Liang, Meng; Ming, Dong; Wan, Baikun; Li, Qiang; Yu, Chunshui

    2015-09-01

    Early deafness can reshape deprived auditory regions to enable the processing of signals from the remaining intact sensory modalities. Cross-modal activation has been observed in auditory regions during non-auditory tasks in early deaf subjects. In hearing subjects, visual working memory can evoke activation of the visual cortex, which further contributes to behavioural performance. In early deaf subjects, however, whether and how auditory regions participate in visual working memory remains unclear. We hypothesized that auditory regions may be involved in visual working memory processing and activation of auditory regions may contribute to the superior behavioural performance of early deaf subjects. In this study, 41 early deaf subjects (22 females and 19 males, age range: 20-26 years, age of onset of deafness < 2 years) and 40 age- and gender-matched hearing controls underwent functional magnetic resonance imaging during a visuo-spatial delayed recognition task that consisted of encoding, maintenance and recognition stages. The early deaf subjects exhibited faster reaction times on the spatial working memory task than did the hearing controls. Compared with hearing controls, deaf subjects exhibited increased activation in the superior temporal gyrus bilaterally during the recognition stage. This increased activation amplitude predicted faster and more accurate working memory performance in deaf subjects. Deaf subjects also had increased activation in the superior temporal gyrus bilaterally during the maintenance stage and in the right superior temporal gyrus during the encoding stage. These increased activation amplitude also predicted faster reaction times on the spatial working memory task in deaf subjects. These findings suggest that cross-modal plasticity occurs in auditory association areas in early deaf subjects. These areas are involved in visuo-spatial working memory. Furthermore, amplitudes of cross-modal activation during the maintenance stage were

  6. Interaction with the Src Homology (SH3-SH2) Region of the Src-family Kinase Hck Structures the HIV-1 Nef Dimer for Kinase Activation and Effector Recruitment*

    PubMed Central

    Alvarado, John Jeff; Tarafdar, Sreya; Yeh, Joanne I.; Smithgall, Thomas E.

    2014-01-01

    HIV-1 Nef supports high titer viral replication in vivo and is essential for AIDS progression. Nef function depends on interactions with multiple host cell effectors, including Hck and other Src-family kinases. Here we describe the x-ray crystal structure of Nef in complex with the Hck SH3-SH2 regulatory region to a resolution of 1.86 Å. The complex crystallized as a dimer of complexes, with the conserved Nef PXXPXR motif engaging the Hck SH3 domain. A new intercomplex contact was found between SH3 Glu-93, and Nef Arg-105. Mutagenesis of Hck SH3 Glu-93 interfered with Nef·Hck complex formation and kinase activation in cells. The Hck SH2 domains impinge on the N-terminal region of Nef to stabilize a dimer conformation that exposes Asp-123, a residue critical for Nef function. Our results suggest that in addition to serving as a kinase effector for Nef, Hck binding may reorganize the Nef dimer for functional interaction with other signaling partners. PMID:25122770

  7. Oomycetes, effectors, and all that jazz.

    PubMed

    Bozkurt, Tolga O; Schornack, Sebastian; Banfield, Mark J; Kamoun, Sophien

    2012-08-01

    Plant pathogenic oomycetes secrete a diverse repertoire of effector proteins that modulate host innate immunity and enable parasitic infection. Understanding how effectors evolve, translocate and traffic inside host cells, and perturb host processes are major themes in the study of oomycete-plant interactions. The last year has seen important progress in the study of oomycete effectors with, notably, the elucidation of the 3D structures of five RXLR effectors, and novel insights into how cytoplasmic effectors subvert host cells. In this review, we discuss these and other recent advances and highlight the most important open questions in oomycete effector biology.

  8. Persistent neural activity in auditory cortex is related to auditory working memory in humans and nonhuman primates.

    PubMed

    Huang, Ying; Matysiak, Artur; Heil, Peter; König, Reinhard; Brosch, Michael

    2016-01-01

    Working memory is the cognitive capacity of short-term storage of information for goal-directed behaviors. Where and how this capacity is implemented in the brain are unresolved questions. We show that auditory cortex stores information by persistent changes of neural activity. We separated activity related to working memory from activity related to other mental processes by having humans and monkeys perform different tasks with varying working memory demands on the same sound sequences. Working memory was reflected in the spiking activity of individual neurons in auditory cortex and in the activity of neuronal populations, that is, in local field potentials and magnetic fields. Our results provide direct support for the idea that temporary storage of information recruits the same brain areas that also process the information. Because similar activity was observed in the two species, the cellular bases of some auditory working memory processes in humans can be studied in monkeys. PMID:27438411

  9. A pilot study examining functional brain activity 6 months after memory retraining in MS: the MEMREHAB trial.

    PubMed

    Dobryakova, Ekaterina; Wylie, Glenn R; DeLuca, John; Chiaravalloti, Nancy D

    2014-09-01

    Cognitive impairment in individuals with multiple sclerosis (MS) is now well recognized. One of the most common cognitive deficits is found in memory functioning, largely due to impaired acquisition. We examined functional brain activity 6 months after memory retraining in individuals with MS. The current report presents long term follow-up results from a randomized clinical trial on a memory rehabilitation protocol known as the modified Story Memory Technique. Behavioral memory performance and brain activity of all participants were evaluated at baseline, immediately after treatment, and 6 months after treatment. Results revealed that previously observed increases in patterns of cerebral activation during learning immediately after memory training were maintained 6 months post training.

  10. Expression of CD39 on Activated T Cells Impairs their Survival in Older Individuals.

    PubMed

    Fang, Fengqin; Yu, Mingcan; Cavanagh, Mary M; Hutter Saunders, Jessica; Qi, Qian; Ye, Zhongde; Le Saux, Sabine; Sultan, William; Turgano, Emerson; Dekker, Cornelia L; Tian, Lu; Weyand, Cornelia M; Goronzy, Jörg J

    2016-02-01

    In an immune response, CD4(+) T cells expand into effector T cells and then contract to survive as long-lived memory cells. To identify age-associated defects in memory cell formation, we profiled activated CD4(+) T cells and found an increased induction of the ATPase CD39 with age. CD39(+) CD4(+) T cells resembled effector T cells with signs of metabolic stress and high susceptibility to undergo apoptosis. Pharmacological inhibition of ATPase activity dampened effector cell differentiation and improved survival, suggesting that CD39 activity influences T cell fate. Individuals carrying a low-expressing CD39 variant responded better to vaccination with an increase in vaccine-specific memory T cells. Increased inducibility of CD39 after activation may contribute to the impaired vaccine response with age. PMID:26832412

  11. Differential associations between impulsivity and risk-taking and brain activations underlying working memory in adolescents

    PubMed Central

    Panwar, Karni; Rutherford, Helena J.V.; Mencl, W. Einar; Lacadie, Cheryl M.; Potenza, Marc N.; Mayes, Linda C.

    2014-01-01

    Increased impulsivity and risk-taking are common during adolescence and relate importantly to addictive behaviors. However, the extent to which impulsivity and risk-taking relate to brain activations that mediate cognitive processing is not well understood. Here we examined the relationships between impulsivity and risk-taking and the neural correlates of working memory. Neural activity was measured in 18 adolescents (13–18 years) while they engaged in a working memory task that included verbal and visuospatial components that each involved encoding, rehearsal and recognition stages. Risk-taking and impulsivity were assessed using the Balloon Analogue Risk Task (BART) and the adolescent version of the Barratt Impulsiveness Scale -11 (BIS-11A), respectively. We found overlapping as well as distinct regions subserving the different stages of verbal and visuospatial working memory. In terms of risk-taking, we found a positive correlation between BART scores and activity in subcortical regions (e.g., thalamus, dorsal striatum) recruited during verbal rehearsal, and an inverse correlation between BART scores and cortical regions (e.g., parietal and temporal regions) recruited during visuospatial rehearsal. The BIS-11A evidenced that motor impulsivity was associated with activity in regions recruited during all stages of working memory, while attention and non-planning impulsivity was only associated with activity in regions recruited during recognition. In considering working memory, impulsivity and risk-taking together, both impulsivity and risk-taking were associated with activity in regions recruited during rehearsal; however, during verbal rehearsal, differential correlations were found. Specifically, positive correlations were found between: (1) risk-taking and activity in subcortical regions, including the thalamus and dorsal striatum; and, (2) motor impulsivity and activity in the left inferior frontal gyrus, insula, dorsolateral and ventrolateral prefrontal

  12. Differential associations between impulsivity and risk-taking and brain activations underlying working memory in adolescents.

    PubMed

    Panwar, Karni; Rutherford, Helena J V; Mencl, W Einar; Lacadie, Cheryl M; Potenza, Marc N; Mayes, Linda C

    2014-11-01

    Increased impulsivity and risk-taking are common during adolescence and relate importantly to addictive behaviors. However, the extent to which impulsivity and risk-taking relate to brain activations that mediate cognitive processing is not well understood. Here we examined the relationships between impulsivity and risk-taking and the neural correlates of working memory. Neural activity was measured in 18 adolescents (13-18 years) while they engaged in a working memory task that included verbal and visuospatial components that each involved encoding, rehearsal and recognition stages. Risk-taking and impulsivity were assessed using the Balloon Analogue Risk Task (BART) and the adolescent version of the Barratt Impulsiveness Scale-11 (BIS-11A), respectively. We found overlapping as well as distinct regions subserving the different stages of verbal and visuospatial working memory. In terms of risk-taking, we found a positive correlation between BART scores and activity in subcortical regions (e.g., thalamus, dorsal striatum) recruited during verbal rehearsal, and an inverse correlation between BART scores and cortical regions (e.g., parietal and temporal regions) recruited during visuospatial rehearsal. The BIS-11A evidenced that motor impulsivity was associated with activity in regions recruited during all stages of working memory, while attention and non-planning impulsivity was only associated with activity in regions recruited during recognition. In considering working memory, impulsivity and risk-taking together, both impulsivity and risk-taking were associated with activity in regions recruited during rehearsal; however, during verbal rehearsal, differential correlations were found. Specifically, positive correlations were found between: (1) risk-taking and activity in subcortical regions, including the thalamus and dorsal striatum; and, (2) motor impulsivity and activity in the left inferior frontal gyrus, insula, and dorsolateral prefrontal cortex. Therefore

  13. Use of the heteroduplex mobility assay and cell sorting to select genome sequences of the CCR5 gene in HEK 293T cells edited by transcription activator-like effector nucleases.

    PubMed

    Nerys-Junior, Arildo; Costa, Lendel C; Braga-Dias, Luciene P; Oliveira, Márcia; Rossi, Atila D; da Cunha, Rodrigo Delvecchio; Gonçalves, Gabriel S; Tanuri, Amilcar

    2014-03-01

    Engineered nucleases such as zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) are one of the most promising tools for modifying genomes. These site-specific enzymes cause double-strand breaks that allow gene disruption or gene insertion, thereby facilitating genetic manipulation. The major problem associated with this approach is the labor-intensive procedures required to screen and confirm the cellular modification by nucleases. In this work, we produced a TALEN that targets the human CCR5 gene and developed a heteroduplex mobility assay for HEK 293T cells to select positive colonies for sequencing. This approach provides a useful tool for the quick detection and easy assessment of nuclease activity.

  14. Use of the heteroduplex mobility assay and cell sorting to select genome sequences of the CCR5 gene in HEK 293T cells edited by transcription activator-like effector nucleases

    PubMed Central

    Nerys-Junior, Arildo; Costa, Lendel C.; Braga-Dias, Luciene P.; Oliveira, Márcia; Rossi, Átila D.; da Cunha, Rodrigo Delvecchio; Gonçalves, Gabriel S.; Tanuri, Amilcar

    2014-01-01

    Engineered nucleases such as zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) are one of the most promising tools for modifying genomes. These site-specific enzymes cause double-strand breaks that allow gene disruption or gene insertion, thereby facilitating genetic manipulation. The major problem associated with this approach is the labor-intensive procedures required to screen and confirm the cellular modification by nucleases. In this work, we produced a TALEN that targets the human CCR5 gene and developed a heteroduplex mobility assay for HEK 293T cells to select positive colonies for sequencing. This approach provides a useful tool for the quick detection and easy assessment of nuclease activity. PMID:24688299

  15. Working memory is not fixed-capacity: More active storage capacity for real-world objects than for simple stimuli.

    PubMed

    Brady, Timothy F; Störmer, Viola S; Alvarez, George A

    2016-07-01

    Visual working memory is the cognitive system that holds visual information active to make it resistant to interference from new perceptual input. Information about simple stimuli-colors and orientations-is encoded into working memory rapidly: In under 100 ms, working memory ‟fills up," revealing a stark capacity limit. However, for real-world objects, the same behavioral limits do not hold: With increasing encoding time, people store more real-world objects and do so with more detail. This boost in performance for real-world objects is generally assumed to reflect the use of a separate episodic long-term memory system, rather than working memory. Here we show that this behavioral increase in capacity with real-world objects is not solely due to the use of separate episodic long-term memory systems. In particular, we show that this increase is a result of active storage in working memory, as shown by directly measuring neural activity during the delay period of a working memory task using EEG. These data challenge fixed-capacity working memory models and demonstrate that working memory and its capacity limitations are dependent upon our existing knowledge.

  16. Treating Another's Actions as One's Own: Children's Memory of and Learning from Joint Activity

    ERIC Educational Resources Information Center

    Sommerville, Jessica A.; Hammond, Amy J.

    2007-01-01

    Children often overestimate their contribution to collaborative activities. Across 2 studies, the authors investigated whether this memory bias supports internalization of the actions of others in the context of joint exchanges. After taking turns with (high collaborative condition; Studies 1 and 2) or working independently of (low collaborative…

  17. The Role of Active Exploration of 3D Face Stimuli on Recognition Memory of Facial Information

    ERIC Educational Resources Information Center

    Liu, Chang Hong; Ward, James; Markall, Helena

    2007-01-01

    Research on face recognition has mainly relied on methods in which observers are relatively passive viewers of face stimuli. This study investigated whether active exploration of three-dimensional (3D) face stimuli could facilitate recognition memory. A standard recognition task and a sequential matching task were employed in a yoked design.…

  18. The Activation and Monitoring of Memories Produced by Words and Pseudohomophones

    ERIC Educational Resources Information Center

    Cortese, Michael J.; Khanna, Maya M.; White, Katherine K.; Veljkovic, Ilija; Drumm, Geoffery

    2008-01-01

    Using the DRM paradigm, our experiments examined the activation and monitoring of memories in semantic and phonological networks. Participants viewed lists of words and/or pseudohomophones (e.g., "dreem"). In Experiment 1, participants verbally recalled lists of semantic associates or attempted to write them as they appeared during study. False…

  19. Making Physical Activity Accessible to Older Adults with Memory Loss: A Feasibility Study

    ERIC Educational Resources Information Center

    Logsdon, Rebecca G.; McCurry, Susan M.; Pike, Kenneth C.; Teri, Linda

    2009-01-01

    Purpose: For individuals with mild cognitive impairment (MCI), memory loss may prevent successful engagement in exercise, a key factor in preventing additional disability. The Resources and Activities for Life Long Independence (RALLI) program uses behavioral principles to make exercise more accessible for these individuals. Exercises are broken…

  20. Activation of MAPK Is Necessary for Long-Term Memory Consolidation Following Food-Reward Conditioning

    ERIC Educational Resources Information Center

    Ribeiro, Maria J.; Schofield, Michael G.; Kemenes, Ildiko; O'Shea, Michael; Kemenes, Gyorgy; Benjamin, Paul R.

    2005-01-01

    Although an important role for the mitogen-activated protein kinase (MAPK) has been established for memory consolidation in a variety of learning paradigms, it is not known if this pathway is also involved in appetitive classical conditioning. We address this question by using a single-trial food-reward conditioning paradigm in the freshwater…

  1. A Role for Prefrontal Calcium-Sensitive Protein Phosphatase and Kinase Activities in Working Memory

    ERIC Educational Resources Information Center

    Runyan, Jason D.; Moore, Anthony N.; Dash, Pramod K.

    2005-01-01

    The prefrontal cortex is involved in the integration and interpretation of information for directing thoughts and planning action. Working memory is defined as the active maintenance of information in mind and is thought to lie at the core of many prefrontal functions. Although dopamine and other neurotransmitters have been implicated, the…

  2. Hyperglycemia induces memory impairment linked to increased acetylcholinesterase activity in zebrafish (Danio rerio).

    PubMed

    Capiotti, Katiucia Marques; De Moraes, Daiani Almeida; Menezes, Fabiano Peres; Kist, Luiza Wilges; Bogo, Maurício Reis; Da Silva, Rosane Souza

    2014-11-01

    Diabetes mellitus, which causes hyperglycemia, affects the central nervous system and can impairs cognitive functions, such as memory. The aim of this study was to investigate the effects of hyperglycemia on memory as well as on the activity of acethylcholinesterase. Hyperglycemia was induced in adult zebrafish by immersion in glucose 111mM by 14 days. The animals were divided in 4 groups: control, glucose-treated, glucose-washout 7-days and glucose-washout 14-days. We evaluated the performance in inhibitory avoidance task and locomotor activity. We also determined acethylcholinesterase activity and gene expression from whole brain. In order to counteract the effect of hyperglycemia underlined by effects on acethylcholinesterase activity, we treated the animals with galantamine (0.05ng/g), an inhibitor of this enzyme. Also we evaluated the gene expression of insulin receptor and glucose transporter from zebrafish brain. The hyperglycemia promoted memory deficit in adult zebrafish, which can be explained by increased AChE activity. The ache mRNA levels from zebrafish brain were decrease in 111mM glucose group and returned to normal levels after 7 days of glucose withdrawal. Insulin receptors (insra-1, insra-2, insrb-1 and insrb-2) and glut-3 mRNA levels were not significantly changed. Our results also demonstrated that galantamine was able to reverse the memory deficit caused by hyperglycemia, demonstrating that these effects involve modulation of AChE activity. These data suggest that the memory impairment induced by hyperglycemia is underlined by the cholinergic dysfunction caused by the mechanisms involving the control of acetylcholinesterase function and gene expression. PMID:25157430

  3. Effect of Mild Thyrotoxicosis on Performance and Brain Activations in a Working Memory Task

    PubMed Central

    Göbel, Anna; Heldmann, Marcus; Göttlich, Martin; Dirk, Anna-Luise; Brabant, Georg; Münte, Thomas F.

    2016-01-01

    Aims Disturbed levels of thyroid hormones are associated with neuropsychiatric disorders, including memory impairments. The aim of this study was to evaluate effects of mild induced thyrotoxicosis on working memory and its neural correlates. Methods Twenty-nine healthy, male subjects with normal thyroid state participated in the study. Functional MRI was acquired during a working memory task (n-back task) before and after ingesting 250 μg L-thyroxin per day for a period of eight weeks. In addition, neuropsychological tests were performed. Results In the hyperthyroid condition the subjects showed slower reaction times, but a higher accuracy in the 0-back version of the memory tasks. Fewer differences between euthyroid and hyperthyroid state were seen for the more difficult conditions of the n-back task. FMRI revealed effects of difficulty in the parahippocampal gyrus, supplementary motor area, prefrontal cortex, anterior cingulate cortex, posterior cerebellum, rolandic operculum and insula (p<0.05, FWE corrected). When comparing euthyroid and hyperthyroid condition in relation to task-induced activation, differences of activation were found in the right prefrontal cortex as well as in the right parahippocampal area. In the psychological assessment, the alerting effect in the Attention Network Task (ANT) and four out of five parameters of the auditory verbal learning test (AVLT) showed an increase from euthyroid to hyperthyroid state. Conclusions It can be concluded that even a short-term intake of thyroid hormones leads to an activation of brain areas associated with working memory and to an improvement of accuracy of working memory tasks. PMID:27536945

  4. The Pseudomonas syringae effector protein HopZ1a suppresses effector-triggered immunity.

    PubMed

    Macho, Alberto P; Guevara, Carlos M; Tornero, Pablo; Ruiz-Albert, Javier; Beuzón, Carmen R

    2010-09-01

    *The Pseudomonas syringae pv syringae type III effector HopZ1a is a member of the HopZ effector family of cysteine-proteases that triggers immunity in Arabidopsis. This immunity is dependent on HopZ1a cysteine-protease activity, and independent of known resistance genes. We have previously shown that HopZ1a-triggered immunity is partially additive to that triggered by AvrRpt2. These partially additive effects could be caused by at least two mechanisms: their signalling pathways share a common element(s), or one effector interferes with the response triggered by the other. *Here, we investigate the molecular basis for the partially additive effect displayed by AvrRpt2- and HopZ1a-triggered immunities, by analysing competitive indices, hypersensitive response and symptom induction, P