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Sample records for effector xopn suppresses

  1. Tomato TFT1 Is Required for PAMP-Triggered Immunity and Mutations that Prevent T3S Effector XopN from Binding to TFT1 Attenuate Xanthomonas Virulence

    E-print Network

    Aakre, Christopher David

    XopN is a type III effector protein from Xanthomonas campestris pathovar vesicatoria that suppresses PAMP-triggered immunity (PTI) in tomato. Previous work reported that XopN interacts with the tomato 14-3-3 isoform TFT1; ...

  2. Tomato TFT1 Is Required for PAMP-Triggered Immunity and Mutations that Prevent T3S Effector XopN from

    E-print Network

    protein from Xanthomonas campestris pathovar vesicatoria that suppresses PAMP-triggered immunity (PTIN from Binding to TFT1 Attenuate Xanthomonas Virulence Kyle W. Taylor1. , Jung-Gun Kim1. , Xue B. Su1¤a-Triggered Immunity and Mutations that Prevent T3S Effector XopN from Binding to TFT1 Attenuate Xanthomonas Virulence

  3. The Pseudomonas syringae type III effector HopD1 suppresses effector-triggered immunity, localizes to the endoplasmic

    E-print Network

    pathogen Pseudomonas syringae pv. tomato DC3000 is the causative agent of bacterial speck disease on tomato to inject type III effector (T3E) proteins into host cells using a type III protein secretion system. OnceThe Pseudomonas syringae type III effector HopD1 suppresses effector-triggered immunity, localizes

  4. Bacterial Effector HopF2 Suppresses Arabidopsis Immunity by Targeting BAK1 

    E-print Network

    Zhou, Jinggeng

    2013-07-19

    Pseudomonas syringae delivers a plethora of effector proteins into host cells to sabotage host immune responses and physiology to favor infection. We have previously shown that P. syringae pv. tomato DC3000 effector HopF2 suppresses Arabidopsis...

  5. Identication of Pseudomonas syringae type III effectors that can suppress programmed cell death in plants and yeast

    E-print Network

    that suppressing plant immunity is one of the primary roles for DC3000 effectors and a central requirement for P effector genes in genome of P. syringae tomato DC3000 has revealed 33 con®rmed effectors and several-apoptotic protein, Bax to induce PCD in plants and yeast, indicating that these effectors function as anti

  6. Bacterial effector HopF2 interacts with AvrPto and suppresses Arabidopsis innate immunity at the plasma membrane

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant pathogenic bacteria inject a cocktail of effector proteins into host plant cells to modulate the host immune response, thereby promoting pathogenicity. How or whether these effectors work cooperatively is largely unknown. The Pseudomonas syringae DC3000 effector HopF2 suppresses the host plan...

  7. Bacterial effector HopF2 suppresses arabidopsis innate immunity at the plasma membrane.

    PubMed

    Wu, Shujing; Lu, Dongping; Kabbage, Mehdi; Wei, Hai-Lei; Swingle, Bryan; Records, Angela R; Dickman, Martin; He, Ping; Shan, Libo

    2011-05-01

    Many bacterial pathogens inject a cocktail of effector proteins into host cells through type III secretion systems. These effectors act in concert to modulate host physiology and immune signaling, thereby promoting pathogenicity. In a search for additional Pseudomonas syringae effectors in suppressing plant innate immunity triggered by pathogen or microbe-associated molecular patterns (PAMPs or MAMPs), we identified P. syringae tomato DC3000 effector HopF2 as a potent suppressor of early immune-response gene transcription and mitogen-activated protein kinase (MAPK) signaling activated by multiple MAMPs, including bacterial flagellin, elongation factor Tu, peptidoglycan, lipopolysaccharide and HrpZ1 harpin, and fungal chitin. The conserved surface-exposed residues of HopF2 are essential for its MAMP suppression activity. HopF2 is targeted to the plant plasma membrane through a putative myristoylation site, and the membrane association appears to be required for its MAMP-suppression function. Expression of HopF2 in plants potently diminished the flagellin-induced phosphorylation of BIK1, a plasma membrane-associated cytoplasmic kinase that is rapidly phosphorylated within one minute upon flagellin perception. Thus, HopF2 likely intercepts MAMP signaling at the plasma membrane immediately of signal perception. Consistent with the potent suppression function of multiple MAMP signaling, expression of HopF2 in transgenic plants compromised plant nonhost immunity to bacteria P. syringae pv. Phaseolicola and plant immunity to the necrotrophic fungal pathogen Botrytis cinerea. PMID:21198360

  8. The majority of the type III effector inventory of Pseudomonas syringae pv. tomato DC3000 can suppress plant immunity.

    PubMed

    Guo, Ming; Tian, Fang; Wamboldt, Yashitola; Alfano, James R

    2009-09-01

    The Pseudomonas syringae type III protein secretion system (T3SS) and the type III effectors it injects into plant cells are required for plant pathogenicity and the ability to elicit a hypersensitive response (HR). The HR is a programmed cell death that is associated with effector-triggered immunity (ETI). A primary function of P. syringae type III effectors appears to be the suppression of ETI and pathogen-associated molecular pattern-triggered immunity (PTI), which is induced by conserved molecules on microorganisms. We reported that seven type III effectors from P. syringae pv. tomato DC3000 were capable of suppressing an HR induced by P. fluorescens(pHIR11) and have now tested 35 DC3000 type III effectors in this assay, finding that the majority of them can suppress the HR induced by HopA1. One newly identified type III effector with particularly strong HR suppression activity was HopS2. We used the pHIR11 derivative pLN1965, which lacks hopA1, in related assays and found that a subset of the type III effectors that suppressed HopA1-induced ETI also suppressed an ETI response induced by AvrRpm1 in Arabidopsis thaliana. A. thaliana plants expressing either HopAO1 or HopF2, two type III effectors that suppressed the HopA1-induced HR, were reduced in the flagellin-induced PTI response as well as PTI induced by other PAMPs and allowed enhanced in planta growth of P. syringae. Collectively, our results suggest that the majority of DC3000 type III effectors can suppress plant immunity. Additionally, the construct pLN1965 will likely be a useful tool in determining whether other type III effectors or effectors from other types of pathogens can suppress either ETI, PTI, or both. PMID:19656042

  9. Suppression of Plant Resistance Gene-Based Immunity by a Fungal Effector

    PubMed Central

    Houterman, Petra M.; Cornelissen, Ben J. C.; Rep, Martijn

    2008-01-01

    The innate immune system of plants consists of two layers. The first layer, called basal resistance, governs recognition of conserved microbial molecules and fends off most attempted invasions. The second layer is based on Resistance (R) genes that mediate recognition of effectors, proteins secreted by pathogens to suppress or evade basal resistance. Here, we show that a plant-pathogenic fungus secretes an effector that can both trigger and suppress R gene-based immunity. This effector, Avr1, is secreted by the xylem-invading fungus Fusarium oxysporum f.sp. lycopersici (Fol) and triggers disease resistance when the host plant, tomato, carries a matching R gene (I or I-1). At the same time, Avr1 suppresses the protective effect of two other R genes, I-2 and I-3. Based on these observations, we tentatively reconstruct the evolutionary arms race that has taken place between tomato R genes and effectors of Fol. This molecular analysis has revealed a hitherto unpredicted strategy for durable disease control based on resistance gene combinations. PMID:18464895

  10. The Pseudomonas syringae effector HopF2 suppresses Arabidopsis immunity by targeting BAK1.

    PubMed

    Zhou, Jinggeng; Wu, Shujing; Chen, Xin; Liu, Chenglong; Sheen, Jen; Shan, Libo; He, Ping

    2014-01-01

    Pseudomonas syringae delivers a plethora of effector proteins into host cells to sabotage immune responses and modulate physiology to favor infection. The P. syringae pv. tomato DC3000 effector HopF2 suppresses Arabidopsis innate immunity triggered by multiple microbe-associated molecular patterns (MAMP) at the plasma membrane. We show here that HopF2 possesses distinct mechanisms for suppression of two branches of MAMP-activated MAP kinase (MAPK) cascades. In addition to blocking MKK5 (MAPK kinase 5) activation in the MEKK1 (MAPK kinase kinase 1)/MEKKs-MKK4/5-MPK3/6 cascade, HopF2 targets additional component(s) upstream of MEKK1 in the MEKK1-MKK1/2-MPK4 cascade and the plasma membrane-localized receptor-like cytoplasmic kinase BIK1 and its homologs. We further show that HopF2 directly targets BAK1, a plasma membrane-localized receptor-like kinase that is involved in multiple MAMP signaling. The interaction between BAK1 and HopF2 and between two other P. syringae effectors, AvrPto and AvrPtoB, was confirmed in vivo and in vitro. Consistent with BAK1 as a physiological target of AvrPto, AvrPtoB and HopF2, the strong growth defects or lethality associated with ectopic expression of these effectors in wild-type Arabidopsis transgenic plants were largely alleviated in bak1 mutant plants. Thus, our results provide genetic evidence to show that BAK1 is a physiological target of AvrPto, AvrPtoB and HopF2. Identification of BAK1 as an additional target of HopF2 virulence not only explains HopF2 suppression of multiple MAMP signaling at the plasma membrane, but also supports the notion that pathogen virulence effectors act through multiple targets in host cells. PMID:24237140

  11. A bacterial cysteine protease effector protein interferes with photosynthesis to suppress plant innate immune responses.

    PubMed

    Rodríguez-Herva, José J; González-Melendi, Pablo; Cuartas-Lanza, Raquel; Antúnez-Lamas, María; Río-Alvarez, Isabel; Li, Ziduo; López-Torrejón, Gema; Díaz, Isabel; Del Pozo, Juan C; Chakravarthy, Suma; Collmer, Alan; Rodríguez-Palenzuela, Pablo; López-Solanilla, Emilia

    2012-05-01

    The bacterial pathogen Pseudomonas syringae pv tomato DC3000 suppresses plant innate immunity with effector proteins injected by a type III secretion system (T3SS). The cysteine protease effector HopN1, which reduces the ability of DC3000 to elicit programmed cell death in non-host tobacco, was found to also suppress the production of defence-associated reactive oxygen species (ROS) and callose when delivered by Pseudomonas fluorescens heterologously expressing a P. syringae T3SS. Purified His(6) -tagged HopN1 was used to identify tomato PsbQ, a member of the oxygen evolving complex of photosystem II (PSII), as an interacting protein. HopN1 localized to chloroplasts and both degraded PsbQ and inhibited PSII activity in chloroplast preparations, whereas a HopN1(D299A) non-catalytic mutant lost these abilities. Gene silencing of NtPsbQ in tobacco compromised ROS production and programmed cell death by DC3000. Our data reveal PsbQ as a contributor to plant immunity responses and a target for pathogen suppression. PMID:22233353

  12. Multiple Candidate Effectors from the Oomycete Pathogen Hyaloperonospora arabidopsidis Suppress Host Plant Immunity

    PubMed Central

    Fabro, Georgina; Steinbrenner, Jens; Coates, Mary; Ishaque, Naveed; Baxter, Laura; Studholme, David J.; Körner, Evelyn; Allen, Rebecca L.; Piquerez, Sophie J. M.; Rougon-Cardoso, Alejandra; Greenshields, David; Lei, Rita; Badel, Jorge L.; Caillaud, Marie-Cecile; Sohn, Kee-Hoon; Van den Ackerveken, Guido; Parker, Jane E.; Beynon, Jim; Jones, Jonathan D. G.

    2011-01-01

    Oomycete pathogens cause diverse plant diseases. To successfully colonize their hosts, they deliver a suite of effector proteins that can attenuate plant defenses. In the oomycete downy mildews, effectors carry a signal peptide and an RxLR motif. Hyaloperonospora arabidopsidis (Hpa) causes downy mildew on the model plant Arabidopsis thaliana (Arabidopsis). We investigated if candidate effectors predicted in the genome sequence of Hpa isolate Emoy2 (HaRxLs) were able to manipulate host defenses in different Arabidopsis accessions. We developed a rapid and sensitive screening method to test HaRxLs by delivering them via the bacterial type-three secretion system (TTSS) of Pseudomonas syringae pv tomato DC3000-LUX (Pst-LUX) and assessing changes in Pst-LUX growth in planta on 12 Arabidopsis accessions. The majority (?70%) of the 64 candidates tested positively contributed to Pst-LUX growth on more than one accession indicating that Hpa virulence likely involves multiple effectors with weak accession-specific effects. Further screening with a Pst mutant (?CEL) showed that HaRxLs that allow enhanced Pst-LUX growth usually suppress callose deposition, a hallmark of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). We found that HaRxLs are rarely strong avirulence determinants. Although some decreased Pst-LUX growth in particular accessions, none activated macroscopic cell death. Fewer HaRxLs conferred enhanced Pst growth on turnip, a non-host for Hpa, while several reduced it, consistent with the idea that turnip's non-host resistance against Hpa could involve a combination of recognized HaRxLs and ineffective HaRxLs. We verified our results by constitutively expressing in Arabidopsis a sub-set of HaRxLs. Several transgenic lines showed increased susceptibility to Hpa and attenuation of Arabidopsis PTI responses, confirming the HaRxLs' role in Hpa virulence. This study shows TTSS screening system provides a useful tool to test whether candidate effectors from eukaryotic pathogens can suppress/trigger plant defense mechanisms and to rank their effectiveness prior to subsequent mechanistic investigation. PMID:22072967

  13. Multiple candidate effectors from the oomycete pathogen Hyaloperonospora arabidopsidis suppress host plant immunity.

    PubMed

    Fabro, Georgina; Steinbrenner, Jens; Coates, Mary; Ishaque, Naveed; Baxter, Laura; Studholme, David J; Körner, Evelyn; Allen, Rebecca L; Piquerez, Sophie J M; Rougon-Cardoso, Alejandra; Greenshields, David; Lei, Rita; Badel, Jorge L; Caillaud, Marie-Cecile; Sohn, Kee-Hoon; Van den Ackerveken, Guido; Parker, Jane E; Beynon, Jim; Jones, Jonathan D G

    2011-11-01

    Oomycete pathogens cause diverse plant diseases. To successfully colonize their hosts, they deliver a suite of effector proteins that can attenuate plant defenses. In the oomycete downy mildews, effectors carry a signal peptide and an RxLR motif. Hyaloperonospora arabidopsidis (Hpa) causes downy mildew on the model plant Arabidopsis thaliana (Arabidopsis). We investigated if candidate effectors predicted in the genome sequence of Hpa isolate Emoy2 (HaRxLs) were able to manipulate host defenses in different Arabidopsis accessions. We developed a rapid and sensitive screening method to test HaRxLs by delivering them via the bacterial type-three secretion system (TTSS) of Pseudomonas syringae pv tomato DC3000-LUX (Pst-LUX) and assessing changes in Pst-LUX growth in planta on 12 Arabidopsis accessions. The majority (~70%) of the 64 candidates tested positively contributed to Pst-LUX growth on more than one accession indicating that Hpa virulence likely involves multiple effectors with weak accession-specific effects. Further screening with a Pst mutant (?CEL) showed that HaRxLs that allow enhanced Pst-LUX growth usually suppress callose deposition, a hallmark of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). We found that HaRxLs are rarely strong avirulence determinants. Although some decreased Pst-LUX growth in particular accessions, none activated macroscopic cell death. Fewer HaRxLs conferred enhanced Pst growth on turnip, a non-host for Hpa, while several reduced it, consistent with the idea that turnip's non-host resistance against Hpa could involve a combination of recognized HaRxLs and ineffective HaRxLs. We verified our results by constitutively expressing in Arabidopsis a sub-set of HaRxLs. Several transgenic lines showed increased susceptibility to Hpa and attenuation of Arabidopsis PTI responses, confirming the HaRxLs' role in Hpa virulence. This study shows TTSS screening system provides a useful tool to test whether candidate effectors from eukaryotic pathogens can suppress/trigger plant defense mechanisms and to rank their effectiveness prior to subsequent mechanistic investigation. PMID:22072967

  14. The Fusarium oxysporum effector Six6 contributes to virulence and suppresses I-2-mediated cell death.

    PubMed

    Gawehns, F; Houterman, P M; Ichou, F Ait; Michielse, C B; Hijdra, M; Cornelissen, B J C; Rep, M; Takken, F L W

    2014-04-01

    Plant pathogens secrete effectors to manipulate their host and facilitate colonization. Fusarium oxysporum f. sp. lycopersici is the causal agent of Fusarium wilt disease in tomato. Upon infection, F. oxysporum f. sp. lycopersici secretes numerous small proteins into the xylem sap (Six proteins). Most Six proteins are unique to F. oxysporum, but Six6 is an exception; a homolog is also present in two Colletotrichum spp. SIX6 expression was found to require living host cells and a knockout of SIX6 in F. oxysporum f. sp. lycopersici compromised virulence, classifying it as a genuine effector. Heterologous expression of SIX6 did not affect growth of Agrobacterium tumefaciens in Nicotiana benthamiana leaves or susceptibility of Arabidopsis thaliana toward Verticillium dahliae, Pseudomonas syringae, or F. oxysporum, suggesting a specific function for F. oxysporum f. sp. lycopersici Six6 in the F. oxysporum f. sp. lycopersici- tomato pathosystem. Remarkably, Six6 was found to specifically suppress I-2-mediated cell death (I2CD) upon transient expression in N. benthamiana, whereas it did not compromise the activity of other cell-death-inducing genes. Still, this I2CD suppressing activity of Six6 does not allow the fungus to overcome I-2 resistance in tomato, suggesting that I-2-mediated resistance is independent from cell death. PMID:24313955

  15. Pseudomonas syringae effector AvrPtoB suppresses basal defence in Arabidopsis.

    PubMed

    de Torres, Marta; Mansfield, John W; Grabov, Nina; Brown, Ian R; Ammouneh, Hassan; Tsiamis, George; Forsyth, Alec; Robatzek, Silke; Grant, Murray; Boch, Jens

    2006-08-01

    The virulence and avirulence activities of members of the Pseudomonas syringae HopAB family of effectors and AvrPto were examined in bean, tomato and Arabidopsis. Proteins were delivered by the RW60 strain of P. syringae pv. phaseolicola. RW60 causes a hypersensitive reaction (HR) in bean and tomato but is restricted without the HR in Arabidopsis. Dual avirulence and virulence functions in tomato and bean, respectively, were identified in virPphA homologues but only avrPtoB strongly enhanced virulence to Arabidopsis, overcoming basal defences operating against RW60. Virulence activity in both bean and Arabidopsis required regions of the C-terminus of the AvrPtoB protein, whereas elicitation of the rapid HR in tomato, with the matching Pto resistance gene, did not. The effect of AvrPtoB on Arabidopsis was accession-specific; most obvious in Wassilewskija (Ws-3), intermediate in Columbia and not detectable in Niedersenz (Nd-1) after inoculation with RW60 + avrPtoB. Analysis of crosses between Ws-3 and Nd-1 indicated co-segregation for the AvrPtoB virulence function with the absence of the Nd-1 FLS2 gene which mediates recognition of bacterial flagellin. In planta expression of AvrPtoB did not prevent the HR activated by P. syringae pv. tomato DC3000 + avrB, avrRpm1, avrRps4 or avrRpt2, but suppressed cell wall alterations, including callose deposition, characteristic of basal defence and was associated with reprogramming of the plant's transcriptional response. The success or failure of AvrPtoB in suppressing basal defences in Nd-1 depended on the timing of exposure of plant cells to the effector and the flagellin flg22 peptide. PMID:16792692

  16. Prophage-Encoded Peroxidase in 'Candidatus Liberibacter asiaticus' Is a Secreted Effector That Suppresses Plant Defenses.

    PubMed

    Jain, Mukesh; Fleites, Laura A; Gabriel, Dean W

    2015-12-01

    'Candidatus Liberibacter asiaticus' is transmitted by psyllids and causes huanglongbing (HLB), a lethal disease of citrus. Most pathogenic 'Ca. L. asiaticus' strains carry two nearly identical prophages similar to SC1 and SC2 in strain UF506. SC2 was observed to replicate as a moderately high-copy excision plasmid encoding a reactive oxygen species-scavenging peroxidase (SC2_gp095), a predicted lysogenic conversion factor. SC2_gp095 was expressed at significantly higher levels in periwinkle than in citrus and was suppressed in psyllids. SC2_gp095 was cloned in a shuttle vector and transformed into Escherichia coli and Liberibacter crescens, a culturable proxy for 'Ca. L. asiaticus'. Transformed L. crescens cells showed 20 to 25% enhanced resistance to H2O2 on agar plates, 47% greater enzymatic activity, and enhanced growth in liquid cultures. A nonclassical secretion potential was predicted for SC2_gp095 and secretion from L. crescens was confirmed by enzymatic and Western blot analyses. Transient expression of SC2_gp095 in planta resulted in strong transcriptional downregulation of RbohB, the key gatekeeper of the H2O2-mediated defense signaling in plants, helping explain the surprisingly long incubation period (years) before HLB symptoms appear in 'Ca. L. asiaticus'-infected citrus. 'Ca. L. asiaticus' peroxidase is likely a secreted, horizontally acquired effector that suppresses host symptom development, a tactic used by most biotrophic plant pathogens. PMID:26313412

  17. The Pseudomonas syringae type III effector HopG1 targets mitochondria, alters plant development and suppresses plant innate immunity.

    PubMed

    Block, Anna; Guo, Ming; Li, Guangyong; Elowsky, Christian; Clemente, Thomas E; Alfano, James R

    2010-03-01

    The bacterial plant pathogen Pseudomonas syringae uses a type III protein secretion system to inject type III effectors into plant cells. Primary targets of these effectors appear to be effector-triggered immunity (ETI) and pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). The type III effector HopG1 is a suppressor of ETI that is broadly conserved in bacterial plant pathogens. Here we show that HopG1 from P. syringae pv. tomato DC3000 also suppresses PTI. Interestingly, HopG1 localizes to plant mitochondria, suggesting that its suppression of innate immunity may be linked to a perturbation of mitochondrial function. While HopG1 possesses no obvious mitochondrial signal peptide, its N-terminal two-thirds was sufficient for mitochondrial localization. A HopG1-GFP fusion lacking HopG1's N-terminal 13 amino acids was not localized to the mitochondria reflecting the importance of the N-terminus for targeting. Constitutive expression of HopG1 in Arabidopsis thaliana, Nicotiana tabacum (tobacco) and Lycopersicon esculentum (tomato) dramatically alters plant development resulting in dwarfism, increased branching and infertility. Constitutive expression of HopG1 in planta leads to reduced respiration rates and an increased basal level of reactive oxygen species. These findings suggest that HopG1's target is mitochondrial and that effector/target interaction promotes disease by disrupting mitochondrial functions. PMID:19863557

  18. Functionally redundant RXLR effectors from Phytophthora infestans act at different steps to suppress early flg22-triggered immunity.

    PubMed

    Zheng, Xiangzi; McLellan, Hazel; Fraiture, Malou; Liu, Xiaoyu; Boevink, Petra C; Gilroy, Eleanor M; Chen, Ying; Kandel, Kabindra; Sessa, Guido; Birch, Paul R J; Brunner, Frédéric

    2014-04-01

    Genome sequences of several economically important phytopathogenic oomycetes have revealed the presence of large families of so-called RXLR effectors. Functional screens have identified RXLR effector repertoires that either compromise or induce plant defense responses. However, limited information is available about the molecular mechanisms underlying the modes of action of these effectors in planta. The perception of highly conserved pathogen- or microbe-associated molecular patterns (PAMPs/MAMPs), such as flg22, triggers converging signaling pathways recruiting MAP kinase cascades and inducing transcriptional re-programming, yielding a generic anti-microbial response. We used a highly synchronizable, pathogen-free protoplast-based assay to identify a set of RXLR effectors from Phytophthora infestans (PiRXLRs), the causal agent of potato and tomato light blight that manipulate early stages of flg22-triggered signaling. Of thirty-three tested PiRXLR effector candidates, eight, called Suppressor of early Flg22-induced Immune response (SFI), significantly suppressed flg22-dependent activation of a reporter gene under control of a typical MAMP-inducible promoter (pFRK1-Luc) in tomato protoplasts. We extended our analysis to Arabidopsis thaliana, a non-host plant species of P. infestans. From the aforementioned eight SFI effectors, three appeared to share similar functions in both Arabidopsis and tomato by suppressing transcriptional activation of flg22-induced marker genes downstream of post-translational MAP kinase activation. A further three effectors interfere with MAMP signaling at, or upstream of, the MAP kinase cascade in tomato, but not in Arabidopsis. Transient expression of the SFI effectors in Nicotiana benthamiana enhances susceptibility to P. infestans and, for the most potent effector, SFI1, nuclear localization is required for both suppression of MAMP signaling and virulence function. The present study provides a framework to decipher the molecular mechanisms underlying the manipulation of host MAMP-triggered immunity (MTI) by P. infestans and to understand the basis of host versus non-host resistance in plants towards P. infestans. PMID:24763622

  19. Candidate effector proteins of the necrotrophic apple canker pathogen Valsa mali can suppress BAX-induced PCD

    PubMed Central

    Li, Zhengpeng; Yin, Zhiyuan; Fan, Yanyun; Xu, Ming; Kang, Zhensheng; Huang, Lili

    2015-01-01

    Canker caused by the Ascomycete Valsa mali is the most destructive disease of apple in Eastern Asia, resulting in yield losses of up to 100%. This necrotrophic fungus induces severe necrosis on apple, eventually leading to the death of the whole tree. Identification of necrosis inducing factors may help to unravel the molecular bases for colonization of apple trees by V. mali. As a first step toward this goal, we identified and characterized the V. mali repertoire of candidate effector proteins (CEPs). In total, 193 secreted proteins with no known function were predicted from genomic data, of which 101 were V. mali-specific. Compared to non-CEPs predicted for the V. mali secretome, CEPs have shorter sequence length and a higher content of cysteine residues. Based on transient over-expression in Nicotiana benthamiana performed for 70 randomly selected CEPs, seven V. mali Effector Proteins (VmEPs) were shown to significantly suppress BAX-induced PCD. Furthermore, targeted deletion of VmEP1 resulted in a significant reduction of virulence. These results suggest that V. mali expresses secreted proteins that can suppress PCD usually associated with effector-triggered immunity (ETI). ETI in turn may play an important role in the V. mali–apple interaction. The ability of V. mali to suppress plant ETI sheds a new light onto the interaction of a necrotrophic fungus with its host plant. PMID:26284095

  20. Homologous RXLR effectors from Hyaloperonospora arabidopsidis and Phytophthora sojae suppress immunity in distantly related plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Diverse pathogens secrete effector proteins into plant cells to manipulate host cellular processes. Oomycete pathogens contain very large complements of predicted effector genes defined by an RXLR host cell entry motif. The genome of Hyaloperonospora arabidopsidis (Hpa, downy mildew of Arabidopsis) ...

  1. A Virulence Essential CRN Effector of Phytophthora capsici Suppresses Host Defense and Induces Cell Death in Plant Nucleus

    PubMed Central

    Mafurah, Joseph Juma; Ma, Huifei; Zhang, Meixiang; Xu, Jing; He, Feng; Ye, Tingyue; Shen, Danyu; Chen, Yanyu; Rajput, Nasir Ahmed; Dou, Daolong

    2015-01-01

    Phytophthora capsici is a soil-borne plant pathogen with a wide range of hosts. The pathogen secretes a large array of effectors during infection of host plants, including Crinkler (CRN) effectors. However, it remains largely unknown on the roles of these effectors in virulence especially in P. capsici. In this study, we identified a cell death-inducing CRN effector PcCRN4 using agroinfiltration approach. Transient expression of PcCRN4 gene induced cell death in N. benthamiana, N. tabacum and Solanum lycopersicum. Overexpression of the gene in N. benthamiana enhanced susceptibility to P. capsici. Subcellular localization results showed that PcCRN4 localized to the plant nucleus, and the localization was required for both of its cell death-inducing activity and virulent function. Silencing PcCRN4 gene in P. capsici significantly reduced pathogen virulence. The expression of the pathogenesis-related gene PR1b in N. benthamiana was significantly induced when plants were inoculated with PcCRN4-silenced P. capsici transformant compared to the wilt-type. Callose deposits were also abundant at sites inoculated with PcCRN4-silenced transformant, indicating that silencing of PcCRN4 in P. capsici reduced the ability of the pathogen to suppress plant defenses. Transcriptions of cell death-related genes were affected when PcCRN4-silenced line were inoculated on Arabidopsis thaliana, suggesting that PcCRN4 may induce cell death by manipulating cell death-related genes. Overall, our results demonstrate that PcCRN4 is a virulence essential effector and it needs target to the plant nucleus to suppress plant immune responses. PMID:26011314

  2. The Pseudomonas syringae type III effector tyrosine phosphatase HopAO1 suppresses innate immunity in Arabidopsis thaliana.

    PubMed

    Underwood, William; Zhang, Shuqun; He, Sheng Y

    2007-11-01

    The bacterial pathogen Pseudomonas syringae pv. tomato (Pst) strain DC3000 infects tomato and Arabidopsis plants, and is a model for studying the molecular basis of bacterial disease. Pst DC3000 secretes a battery of largely uncharacterized effector proteins into host cells via a type-III secretion system (TTSS). Little is currently known about the molecular mechanisms by which individual TTSS effectors promote virulence. The effector HopAO1 has similarity to protein tyrosine phosphatases, including a conserved catalytic site, and suppresses the hypersensitive response (HR) in some non-host plants. Whether HopAO1 has a similar effect in the host Arabidopsis is not clear. Here, we show that transgenic expression of HopAO1 in Arabidopsis suppresses callose deposition elicited by the Pst DC3000 hrpA mutant, and allows the normally non-pathogenic hrpA mutant to multiply within the leaf tissue. HopAO1 also suppresses resistance to Pst DC3000 induced by flg22, a pathogen-associated molecular pattern (PAMP). However, HopAO1 does not suppress the HR triggered by several classical avirulence genes. These results suggest that HopAO1 targets primarily PAMP-induced innate immunity in Arabidopsis. The virulence function of HopAO1 is dependent on an intact phosphatase catalytic site, as transgenic plants expressing a catalytically inactive derivative do not show these effects. Intriguingly, expression of the catalytically inactive HopAO1 has a dominant-negative effect on the function of the wild-type HopAO1. Analysis of mitogen-activated protein kinase (MAPK) activity suggests that HopAO1 targets a step downstream or independent of MAPK activation. Genome-wide expression analysis revealed that expression of several well-known defense genes was suppressed in hrpA mutant-infected HopAO1 transgenic plants. PMID:17877704

  3. Two Host Cytoplasmic Effectors Are Required for Pathogenesis of Phytophthora sojae by Suppression of Host Defenses1[W][OA

    PubMed Central

    Liu, Tingli; Ye, Wenwu; Ru, Yanyan; Yang, Xinyu; Gu, Biao; Tao, Kai; Lu, Shan; Dong, Suomeng; Zheng, Xiaobo; Shan, Weixing; Wang, Yuanchao; Dou, Daolong

    2011-01-01

    Phytophthora sojae encodes hundreds of putative host cytoplasmic effectors with conserved FLAK motifs following signal peptides, termed crinkling- and necrosis-inducing proteins (CRN) or Crinkler. Their functions and mechanisms in pathogenesis are mostly unknown. Here, we identify a group of five P. sojae-specific CRN-like genes with high levels of sequence similarity, of which three are putative pseudogenes. Functional analysis shows that the two functional genes encode proteins with predicted nuclear localization signals that induce contrasting responses when expressed in Nicotiana benthamiana and soybean (Glycine max). PsCRN63 induces cell death, while PsCRN115 suppresses cell death elicited by the P. sojae necrosis-inducing protein (PsojNIP) or PsCRN63. Expression of CRN fragments with deleted signal peptides and FLAK motifs demonstrates that the carboxyl-terminal portions of PsCRN63 or PsCRN115 are sufficient for their activities. However, the predicted nuclear localization signal is required for PsCRN63 to induce cell death but not for PsCRN115 to suppress cell death. Furthermore, silencing of the PsCRN63 and PsCRN115 genes in P. sojae stable transformants leads to a reduction of virulence on soybean. Intriguingly, the silenced transformants lose the ability to suppress host cell death and callose deposition on inoculated plants. These results suggest a role for CRN effectors in the suppression of host defense responses. PMID:21071601

  4. Functional analysis of plant defense suppression and activation by the Xanthomonas core type III effector XopX

    PubMed Central

    Stork, William; Kim, Jung-Gun; Mudgett, Mary Beth

    2014-01-01

    Many phytopathogenic type III secretion effectors (T3Es) have been shown to target and suppress plant immune signaling, but perturbation of the plant immune system by T3Es can also elicit a plant response. XopX is a “core” Xanthomonas T3E that contributes to growth and symptom development during Xanthomonas euvesicatoria (Xe) infection of tomato, but its functional role is undefined. We tested the effect of XopX on several aspects of plant immune signaling. XopX promoted ethylene production and plant cell death (PCD) during Xe infection of susceptible tomato and in transient expression assays in Nicotiana benthamiana, which is consistent with its requirement for the development of Xe-induced disease symptoms. Additionally, although XopX suppressed flagellin-induced reactive oxygen species, it promoted the accumulation of pattern-triggered immunity (PTI) gene transcripts. Surprisingly, XopX co-expression with other PCD elicitors resulted in delayed PCD, suggesting antagonism between XopX-dependent PCD and other PCD pathways. However, we found no evidence that XopX contributed to the suppression of effector-triggered immunity during Xe-tomato interactions, suggesting that XopX’s primary virulence role is to modulate PTI. These results highlight the dual role of a core Xanthomonas T3E in simultaneously suppressing and activating plant defense responses. PMID:25338145

  5. Repression of SATB1 in regulatory T cells is required for suppressive function and inhibition of effector differentiation

    PubMed Central

    Beyer, Marc; Thabet, Yasser; Müller, Roman-Ulrich; Sadlon, Timothy; Classen, Sabine; Lahl, Katharina; Basu, Samik; Zhou, Xuyu; Bailey-Bucktrout, Samantha L.; Krebs, Wolfgang; Schönfeld, Eva A.; Böttcher, Jan; Golovina, Tatiana; Mayer, Christian T.; Hofmann, Andrea; Sommer, Daniel; Debey-Pascher, Svenja; Endl, Elmar; Limmer, Andreas; Hippen, Keli L.; Blazar, Bruce R.; Balderas, Robert; Quast, Thomas; Waha, Andreas; Mayer, Günter; Famulok, Michael; Knolle, Percy A.; Wickenhauser, Claudia; Kolanus, Waldemar; Schermer, Bernhard; Bluestone, Jeffrey A.; Barry, Simon C.; Sparwasser, Tim; Riley, James L.; Schultze, Joachim L.

    2013-01-01

    Regulatory T (Treg) cells are essential for self-tolerance and immune homeostasis. Lack of effector T cell (Teff) function and gain of suppressive activity by Treg are dependent on the transcriptional program induced by Foxp3. Here we report repression of SATB1, a genome organizer regulating chromatin structure and gene expression, as crucial for Treg phenotype and function. Foxp3, acting as a transcriptional repressor, directly suppressed the SATB1 locus and indirectly through induction of microRNAs that bound the SATB1 3?UTR. Release of SATB1 from Foxp3 control in Treg caused loss of suppressive function, establishment of transcriptional Teff programs and induction of Teff cytokines. These data support that inhibition of SATB1-mediated modulation of global chromatin remodelling is pivotal for maintaining Treg functionality. PMID:21841785

  6. Toxicity and SidJ-Mediated Suppression of Toxicity Require Distinct Regions in the SidE Family of Legionella pneumophila Effectors.

    PubMed

    Havey, James C; Roy, Craig R

    2015-09-01

    Intracellular bacteria use a variety of strategies to evade degradation and create a replicative niche. Legionella pneumophila is an intravacuolar pathogen that establishes a replicative niche through the secretion of more than 300 effector proteins. The function of most effectors remains to be determined. Toxicity in yeast has been used to identify functional domains and elucidate the biochemical function of effectors. A library of L. pneumophila effectors was screened using an expression plasmid that produces low levels of each protein. This screen identified the effector SdeA as a protein that confers a strong toxic phenotype that inhibits yeast replication. The toxicity of SdeA was suppressed in cells producing the effector SidJ. The effector SdeA is a member of the SidE family of L. pneumophila effector proteins. All SidE orthologs encoded by the Philadelphia isolate of Legionella pneumophila were toxic to yeast, and SidJ suppressed the toxicity of each. We identified a conserved central region in the SidE proteins that was sufficient to mediate yeast toxicity. Surprisingly, SidJ did not suppress toxicity when this central region was produced in yeast. We determined that the amino-terminal region of SidE was essential for SidJ-mediated suppression of toxicity. Thus, there is a genetic interaction that links the activity of SidJ and the amino-terminal region of SidE, which is required to modulate the toxic activity displayed by the central region of the SidE protein. This suggests a complex mechanism by which the L. pneumophila effector SidJ modulates the function of the SidE proteins after translocation into host cells. PMID:26099583

  7. Analysis of Globodera rostochiensis effectors reveals conserved functions of SPRYSEC proteins in suppressing and eliciting plant immune responses

    PubMed Central

    Ali, Shawkat; Magne, Maxime; Chen, Shiyan; Obradovic, Natasa; Jamshaid, Lubna; Wang, Xiaohong; Bélair, Guy; Moffett, Peter

    2015-01-01

    Potato cyst nematodes (PCNs), including Globodera rostochiensis (Woll.), are important pests of potato. Plant parasitic nematodes produce multiple effector proteins, secreted from their stylets, to successfully infect their hosts. These include proteins delivered to the apoplast and to the host cytoplasm. A number of effectors from G. rostochiensis predicted to be delivered to the host cytoplasm have been identified, including several belonging to the secreted SPRY domain (SPRYSEC) family. SPRYSEC proteins are unique to members of the genus Globodera and have been implicated in both the induction and the repression of host defense responses. We have tested the properties of six different G. rostochiensis SPRYSEC proteins by expressing them in Nicotiana benthamiana and N. tabacum. We have found that all SPRYSEC proteins tested are able to suppress defense responses induced by NB-LRR proteins as well as cell death induced by elicitors, suggesting that defense repression is a common characteristic of members of this effector protein family. At the same time, GrSPRYSEC-15 elicited a defense responses in N. tabacum, which was found to be resistant to a virus expressing GrSPRYSEC-15. These results suggest that SPRYSEC proteins may possess characteristics that allow them to be recognized by the plant immune system. PMID:26322064

  8. The Pseudomonas syringae pv. tomato type III effector HopM1 suppresses Arabidopsis defenses independent of suppressing salicylic acid signaling and of targeting AtMIN7.

    PubMed

    Gangadharan, Anju; Sreerekha, Mysore-Venkatarau; Whitehill, Justin; Ham, Jong Hyun; Mackey, David

    2013-01-01

    Pseudomonas syringae pv tomato strain DC3000 (Pto) delivers several effector proteins promoting virulence, including HopM1, into plant cells via type III secretion. HopM1 contributes to full virulence of Pto by inducing degradation of Arabidopsis proteins, including AtMIN7, an ADP ribosylation factor-guanine nucleotide exchange factor. Pseudomonas syringae pv phaseolicola strain NPS3121 (Pph) lacks a functional HopM1 and elicits robust defenses in Arabidopsis thaliana, including accumulation of pathogenesis related 1 (PR-1) protein and deposition of callose-containing cell wall fortifications. We have examined the effects of heterologously expressed HopM1Pto on Pph-induced defenses. HopM1 suppresses Pph-induced PR-1 expression, a widely used marker for salicylic acid (SA) signaling and systemic acquired resistance. Surprisingly, HopM1 reduces PR-1 expression without affecting SA accumulation and also suppresses the low levels of PR-1 expression apparent in SA-signaling deficient plants. Further, HopM1 enhances the growth of Pto in SA-signaling deficient plants. AtMIN7 contributes to Pph-induced PR-1 expression. However, HopM1 fails to degrade AtMIN7 during Pph infection and suppresses Pph-induced PR-1 expression and callose deposition in wild-type and atmin7 plants. We also show that the HopM1-mediated suppression of PR-1 expression is not observed in plants lacking the TGA transcription factor, TGA3. Our data indicate that HopM1 promotes bacterial virulence independent of suppressing SA-signaling and links TGA3, AtMIN7, and other HopM1 targets to pathways distinct from the canonical SA-signaling pathway contributing to PR-1 expression and callose deposition. Thus, efforts to understand this key effector must consider multiple targets and unexpected outputs of its action. PMID:24324742

  9. The 1,4-benzodiazepine Ro5-4864 (4-chlorodiazepam) suppresses multiple pro-inflammatory mast cell effector functions

    PubMed Central

    2013-01-01

    Activation of mast cells (MCs) can be achieved by the high-affinity receptor for IgE (Fc?RI) as well as by additional receptors such as the lipopolysaccharide (LPS) receptor and the receptor tyrosine kinase Kit (stem cell factor [SCF] receptor). Thus, pharmacological interventions which stabilize MCs in response to different receptors would be preferable in diseases with pathological systemic MC activation such as systemic mastocytosis. 1,4-Benzodiazepines (BDZs) have been reported to suppress MC effector functions. In the present study, our aim was to analyze molecularly the effects of BDZs on MC activation by comparison of the effects of the two BDZs Ro5-4864 and clonazepam, which markedly differ in their affinities for the archetypical BDZ recognition sites, i.e., the GABAA receptor and TSPO (previously termed peripheral-type BDZ receptor). Ro5-4864 is a selective agonist at TSPO, whereas clonazepam is a selective agonist at the GABAA receptor. Ro5-4864 suppressed pro-inflammatory MC effector functions in response to antigen (Ag) (degranulation/cytokine production) and LPS and SCF (cytokine production), whereas clonazepam was inactive. Signaling pathway analyses revealed inhibitory effects of Ro5-4864 on Ag-triggered production of reactive oxygen species, calcium mobilization and activation of different downstream kinases. The initial activation of Src family kinases was attenuated by Ro5-4864 offering a molecular explanation for the observed impacts on various downstream signaling elements. In conclusion, BDZs structurally related to Ro5-4864 might serve as multifunctional MC stabilizers without the sedative effect of GABAA receptor-interacting BDZs. PMID:23425659

  10. Suppression of plant defenses by a Myzus persicae (green peach aphid) salivary effector protein

    PubMed Central

    Elzinga, Dezi A.; De Vos, Martin

    2014-01-01

    The complex interactions between aphids and their host plant are species-specific and involve multiple layers of recognition and defense. Aphid salivary proteins, which are released into the plant during phloem feeding, are a likely mediator of these interactions. In an approach to identify aphid effectors that facilitate feeding from host plants, eleven Myzus persicae (green peach aphid) salivary proteins and the GroEL protein of Buchnera aphidicola, a bacterial endosymbiont of this aphid species, were expressed transiently in Nicotiana tabacum (tobacco). Whereas two salivary proteins increased aphid reproduction, expression of three other aphid proteins and GroEL significantly decreased aphid reproduction on N. tabacum. These effects were recapitulated in stable transgenic Arabidopsis thaliana (Arabidopsis) plants. Further experiments with A. thaliana expressing Mp55, a salivary protein that increased aphid reproduction, showed lower accumulation of 4-methoxyindol-3-ylmethylglucosinolate, callose, and hydrogen peroxide in response to aphid feeding. Mp55-expressing plants also were more attractive for aphids in choice assays. Silencing Mp55 gene expression in M. persicae using RNA interference approaches reduced aphid reproduction on N. tabacum, A. thaliana, and Nicotiana benthamiana. Together, these results demonstrate a role for Mp55, a protein with as yet unknown molecular function, in the interaction of M. persicae with its host plants. PMID:24654979

  11. Analysis of Putative Apoplastic Effectors from the Nematode, Globodera rostochiensis, and Identification of an Expansin-Like Protein That Can Induce and Suppress Host Defenses

    PubMed Central

    Ali, Shawkat; Magne, Maxime; Chen, Shiyan; Côté, Olivier; Stare, Barbara Geri?; Obradovic, Natasa; Jamshaid, Lubna; Wang, Xiaohong; Bélair, Guy; Moffett, Peter

    2015-01-01

    The potato cyst nematode, Globodera rostochiensis, is an important pest of potato. Like other pathogens, plant parasitic nematodes are presumed to employ effector proteins, secreted into the apoplast as well as the host cytoplasm, to alter plant cellular functions and successfully infect their hosts. We have generated a library of ORFs encoding putative G. rostochiensis putative apoplastic effectors in vectors for expression in planta. These clones were assessed for morphological and developmental effects on plants as well as their ability to induce or suppress plant defenses. Several CLAVATA3/ESR-like proteins induced developmental phenotypes, whereas predicted cell wall-modifying proteins induced necrosis and chlorosis, consistent with roles in cell fate alteration and tissue invasion, respectively. When directed to the apoplast with a signal peptide, two effectors, an ubiquitin extension protein (GrUBCEP12) and an expansin-like protein (GrEXPB2), suppressed defense responses including NB-LRR signaling induced in the cytoplasm. GrEXPB2 also elicited defense response in species- and sequence-specific manner. Our results are consistent with the scenario whereby potato cyst nematodes secrete effectors that modulate host cell fate and metabolism as well as modifying host cell walls. Furthermore, we show a novel role for an apoplastic expansin-like protein in suppressing intra-cellular defense responses. PMID:25606855

  12. SOBER1 phospholipase activity suppresses phosphatidic acid accumulation and plant immunity in response to bacterial effector AvrBsT.

    PubMed

    Kirik, Angela; Mudgett, Mary Beth

    2009-12-01

    Arabidopsis thaliana ecotype Pi-0 is resistant to Pseudomonas syringae pathovar tomato (Pst) strain DC3000 expressing the T3S effector protein AvrBsT. Resistance is due to a loss of function mutation (sober1-1) in a conserved alpha/beta hydrolase, SOBER1 (Suppressor of AvrBsT Elicited Resistance1). Members of this superfamily possess phospholipase and carboxylesterase activity with diverse substrate specificity. The nature of SOBER1 enzymatic activity and substrate specificity was not known. SOBER1-dependent suppression of the hypersensitive response (HR) in Pi-0 suggested that it might hydrolyze a plant lipid or precursor required for HR induction. Here, we show that Pi-0 leaves infected with Pst DC3000 expressing AvrBsT accumulated higher levels of phosphatidic acid (PA) compared to leaves infected with Pst DC3000. Phospholipase D (PLD) activity was required for high PA levels and AvrBsT-dependent HR in Pi-0. Overexpression of SOBER1 in Pi-0 reduced PA levels and inhibited HR. These data implicated PA, phosphatidylcholine (PC) and lysophosphatidylcholine (LysoPC) as potential SOBER1 substrates. Recombinant His(6)-SOBER1 hydrolyzed PC but not PA or LysoPC in vitro indicating that the enzyme has phospholipase A(2) (PLA(2)) activity. Chemical inhibition of PLA(2) activity in leaves expressing SOBER1 resulted in HR in response to Pst DC3000 AvrBsT. These data are consistent with the model that SOBER1 PLA(2) activity suppresses PLD-dependent production of PA in response to AvrBsT elicitation. This work highlights an important role for SOBER1 in the regulation of PA levels generated in plants in response to biotic stress. PMID:19918071

  13. Lactose inhibits regulatory T-cell-mediated suppression of effector T-cell interferon-? and IL-17 production.

    PubMed

    Paasela, Monika; Kolho, Kaija-Leena; Vaarala, Outi; Honkanen, Jarno

    2014-12-14

    Our interest in lactose as an immunomodulatory molecule results from studies showing that lactose binds to galectin-9, which has been shown to have various regulatory functions in the immune system including regulation of T-cell responses. Impaired regulation of T helper (Th)1 and Th17 type immune responses and dysfunction of regulatory T cells (Treg) have been implicated in many human immune-mediated diseases. In the present study, we investigated the effects of lactose on immune regulation using co-cultures of human peripheral blood mononuclear cell (PBMC)-derived Treg and effector T cells (Teff) obtained from twenty healthy adults. Treg, i.e. CD4+CD25+CD127-, were isolated from PBMC by immunomagnetic separation. The fraction of CD4+CD127- cells that was depleted of CD25+ cells was used as Teff. Treg and Teff at a ratio 1:5 were activated and the effects of lactose on the secretion of interferon-? (IFN-?) and IL-17 were analysed using ELISA for protein and quantitative RT-PCR for mRNA. Treg down-regulated the secretion of both IFN-? (8.8-3.9 ng/ml, n 20, P= 0.003) and IL-17 (0.83-0.64 ng/ml, n 15, P= 0.04) in co-cultures, while in the presence of lactose the levels of secreted IFN-? and IL-17 remained high and no down-regulation was observed (16.4 v. 3.99 ng/ml, n 20, P< 0.0001, and 0.74 v. 0.64 ng/ml, n 15, P= 0.005, respectively). We showed that lactose inhibits human Treg-mediated suppression of Th1 and Th17 immune responses in vitro. PMID:25331548

  14. Broadly Conserved Fungal Effector BEC1019 Suppresses Host Cell Death and Enhances Pathogen Virulence in Powdery Mildew of Barley (Hordeum vulgare L.).

    PubMed

    Whigham, Ehren; Qi, Shan; Mistry, Divya; Surana, Priyanka; Xu, Ruo; Fuerst, Gregory; Pliego, Clara; Bindschedler, Laurence V; Spanu, Pietro D; Dickerson, Julie A; Innes, Roger W; Nettleton, Dan; Bogdanove, Adam J; Wise, Roger P

    2015-09-01

    The interaction of barley, Hordeum vulgare L., with the powdery mildew fungus Blumeria graminis f. sp. hordei is a well-developed model to investigate resistance and susceptibility to obligate biotrophic pathogens. The 130-Mb Blumeria genome encodes approximately 540 predicted effectors that are hypothesized to suppress or induce host processes to promote colonization. Blumeria effector candidate (BEC)1019, a single-copy gene encoding a putative, secreted metalloprotease, is expressed in haustorial feeding structures, and host-induced gene silencing of BEC1019 restricts haustorial development in compatible interactions. Here, we show that Barley stripe mosaic virus-induced gene silencing of BEC1019 significantly reduces fungal colonization of barley epidermal cells, demonstrating that BEC1019 plays a central role in virulence. In addition, delivery of BEC1019 to the host cytoplasm via Xanthomonas type III secretion suppresses cultivar nonspecific hypersensitive reaction (HR) induced by Xanthomonas oryzae pv. oryzicola, as well as cultivar-specific HR induced by AvrPphB from Pseudomonas syringae pv. phaseolicola. BEC1019 homologs are present in 96 of 241 sequenced fungal genomes, including plant pathogens, human pathogens, and free-living nonpathogens. Comparative analysis revealed variation at several amino acid positions that correlate with fungal lifestyle and several highly conserved, noncorrelated motifs. Site-directed mutagenesis of one of these, ETVIC, compromises the HR-suppressing activity of BEC1019. We postulate that BEC1019 represents an ancient, broadly important fungal protein family, members of which have evolved to function as effectors in plant and animal hosts. PMID:25938194

  15. Basal resistance against bacteria in Nicotiana benthamiana leaves is accompanied by reduced vascular staining and suppressed by multiple Pseudomonas syringae type III secretion system effector proteins.

    PubMed

    Oh, Hye-Sook; Collmer, Alan

    2005-10-01

    Basal resistance in plants is induced by flagellin and several other common bacterial molecules and is implicated in the immunity of plants to most bacteria and other microbes. However, basal resistance can be suppressed by effector proteins that are injected by the type III secretion system (TTSS) of pathogens such as Pseudomonas syringae. This study demonstrates that basal resistance in the leaves of Nicotiana benthamiana is accompanied by reduced vascular flow into minor veins. Reduced vascular flow was assayed by feeding leaves, via freshly excised petioles, with 1% (weight in volume, w/v) neutral red (NR) and then observing differential staining of minor veins or altered levels of extractable dye in excised leaf samples. The reduced vascular staining was localized to tissues expressing basal resistance and was observable when resistance was induced by either the non-pathogen Pseudomonas fluorescens, a TTSS-deficient mutant of P. syringae pv. tabaci, or flg22 (a flagellin-derived peptide elicitor of basal resistance). Nicotiana benthamiana leaf areas expressing basal resistance no longer elicited the hypersensitive response when challenge inoculated with P. syringae pv. tomato DC3000. The reduced vascular staining effect was suppressed by wild-type P. syringae pv. tabaci and P. fluorescens heterologously expressing a P. syringae TTSS and AvrPto1(PtoJL1065). TTSS-proficient P. fluorescens was used to test the ability of several P. syringae pv. tomato DC3000 effectors for their ability to suppress the basal resistance-associated reduced vascular staining effect. AvrE(PtoDC3000), HopM1(PtoDC3000) (formerly known as HopPtoM), HopF2(PtoDC3000) (HopPtoF) and HopG1(PtoDC3000) (HopPtoG) suppressed basal resistance by this test, whereas HopC1(PtoDC3000) (HopPtoC) did not. In summary, basal resistance locally alters vascular function and the vascular dye uptake assay should be a useful tool for characterizing effectors that suppress basal resistance. PMID:16212612

  16. CTLA4-Ig suppresses development of experimental autoimmune uveitis in the induction and effector phases: Comparison with blockade of interleukin-6.

    PubMed

    Iwahashi, Chiharu; Fujimoto, Minoru; Nomura, Shintaro; Serada, Satoshi; Nakai, Kei; Ohguro, Nobuyuki; Nishida, Kohji; Naka, Tetsuji

    2015-11-01

    Recently, a number of biologics have been used in the treatment of autoimmune diseases. However, in the treatment of severe autoimmune uveitis, only TNF-alpha inhibitors are preferably used and the effect of other biologics such as interleukin-6 (IL-6) signaling blockade or cytotoxic T-lymphocyte antigen-4-immunoglobulin fusion protein (CTLA4-Ig) has not been well studied. Previously, we reported that IL-6 blockade effectively suppresses the development of experimental autoimmune uveitis (EAU), a mouse model for uveitis, by inhibiting Th17 cell development. In this study, we investigated the effect of CTLA4-Ig on EAU development and compared it with the effect of anti-IL-6 receptor monoclonal antibody (MR16-1). C57BL/6J mice were immunized with interphotoreceptor retinoid-binding protein (IRBP) and treated once with CTLA4-Ig or MR16-1. Both CTLA4-Ig and MR16-1 administered in the induction phase (the same day as immunization) significantly reduced the clinical and histopathological scores of EAU. Fluorescence-activated cell sorting studies using draining lymph node (LN) cells from EAU mice 10 days after immunization showed that CTLA4-Ig can suppress early T-helper cell activation. CTLA4-Ig administered in the effector phase of the disease (one week after immunization), when IRBP-reactive T cells have been primed, also significantly reduced the clinical and histopathological scores of EAU. In contrast, MR16-1 administered in the effector phase did not ameliorate EAU. To investigate the differences between these biologics in the effector phase, in vitro restimulation analysis of LN cells obtained from EAU mice one week after immunization was performed and revealed that CTLA4-Ig, but not MR16-1, added to culture media could inhibit the proliferation of IRBP-specific CD4(+) T cells which possessed capacities of producing IFN-gamma and/or IL-17. Collectively, CTLA4-Ig ameliorated EAU through preventing initial T-cell activation in the induction phase and suppressing proliferation of IRBP-specific T cells in the effector phase. Blockade of IL-6 signaling did not have such inhibitory effects after T-cell priming. CTLA4-Ig may have therapeutic effects on human chronic uveitis. PMID:26297802

  17. Cutting edge: Genetic variation in TLR1 is associated with Pam3CSK4-induced effector T cell resistance to regulatory T cell suppression.

    PubMed

    Mikacenic, Carmen; Schneider, Anya; Radella, Frank; Buckner, Jane H; Wurfel, Mark M

    2014-12-15

    TLR play essential roles in the initiation and modulation of immune responses. TLR1/TLR2 heterodimers recognize triacylated bacterial lipopeptides, including the synthetic TLR1/2 lipopeptide Pam3CSK4. Genetic variation in TLR1 is associated with outcomes in diseases in which regulatory T cells (Treg) play a role, including asthma and allergy. To determine whether genetic polymorphisms in TLR1 are associated with alterations in Treg suppression of effector T cells (Teff), we performed in vitro suppression assays in healthy individuals with various haplotypes in TLR1. We show that functional genetic polymorphisms in TLR1 modify surface expression of TLR1 on T lymphocytes and confer enhanced Teff resistance to Treg suppression in the presence of Pam3CSK4. These effects are mediated, in part, by IL-6 and inhibited by blocking IL-6 signaling through STAT3. These findings suggest that TLR1 polymorphisms could influence immune-related disease through Teff resistance to Treg suppression. PMID:25378593

  18. Pseudomonas syringae Effector AvrPphB Suppresses AvrB-Induced Activation of RPM1 but Not AvrRpm1-Induced Activation.

    PubMed

    Russell, Andrew R; Ashfield, Tom; Innes, Roger W

    2015-06-01

    The Pseudomonas syringae effector AvrB triggers a hypersensitive resistance response in Arabidopsis and soybean plants expressing the disease resistance (R) proteins RPM1 and Rpg1b, respectively. In Arabidopsis, AvrB induces RPM1-interacting protein kinase (RIPK) to phosphorylate a disease regulator known as RIN4, which subsequently activates RPM1-mediated defenses. Here, we show that AvrPphB can suppress activation of RPM1 by AvrB and this suppression is correlated with the cleavage of RIPK by AvrPphB. Significantly, AvrPphB does not suppress activation of RPM1 by AvrRpm1, suggesting that RIPK is not required for AvrRpm1-induced modification of RIN4. This observation indicates that AvrB and AvrRpm1 recognition is mediated by different mechanisms in Arabidopsis, despite their recognition being determined by a single R protein. Moreover, AvrB recognition but not AvrRpm1 recognition is suppressed by AvrPphB in soybean, suggesting that AvrB recognition requires a similar molecular mechanism in soybean and Arabidopsis. In support of this, we found that phosphodeficient mutations in the soybean GmRIN4a and GmRIN4b proteins are sufficient to block Rpg1b-mediated hypersensitive response in transient assays in Nicotiana glutinosa. Taken together, our results indicate that AvrB and AvrPphB target a conserved defense signaling pathway in Arabidopsis and soybean that includes RIPK and RIN4. PMID:25625821

  19. Xanthomonas Type III Effector XopD Desumoylates Tomato Transcription Factor SlERF4 to Suppress Ethylene Responses and Promote Pathogen Growth

    PubMed Central

    Kim, Jung-Gun; Stork, William; Mudgett, Mary Beth

    2013-01-01

    SUMMARY XopD, a type III secretion effector from Xanthomonas euvesicatoria (Xcv), the causal agent of bacterial spot of tomato is required for pathogen growth and delay of host symptom development. XopD carries a C-terminal SUMO protease domain, a host range determining non-specific DNA-binding domain and two EAR motifs typically found in repressors of stress-induced transcription. The precise target(s) and mechanism(s) of XopD are obscure. We report that XopD directly targets the tomato ethylene responsive transcription factor SlERF4 to suppress ethylene production, which is required for anti-Xcv immunity and symptom development. SlERF4 expression was required for Xcv ?xopD-induced ethylene production and ethylene -stimulated immunity. XopD colocalized with SlERF4 in subnuclear foci and catalyzed SUMO1 hydrolysis from lysine 53 of SlERF4 causing SlERF4 destabilization. Mutation of lysine 53 prevented SlERF4 sumoylation, decreased SlERF4 levels, and reduced SlERF4 transcription. These data suggest that XopD desumoylates SlERF4 to repress ethylene induced-transcription required for anti-Xcv immunity. PMID:23414755

  20. The Parasitic Worm Product ES-62 Targets Myeloid Differentiation Factor 88–Dependent Effector Mechanisms to Suppress Antinuclear Antibody Production and Proteinuria in MRL/lpr Mice

    PubMed Central

    Rodgers, David T; McGrath, Mairi A; Pineda, Miguel A; Al-Riyami, Lamyaa; Rzepecka, Justyna; Lumb, Felicity; Harnett, William; Harnett, Margaret M

    2015-01-01

    Objective The hygiene hypothesis suggests that parasitic helminths (worms) protect against the development of autoimmune disease via a serendipitous side effect of worm-derived immunomodulators that concomitantly promote parasite survival and limit host pathology. The aim of this study was to investigate whether ES-62, a phosphorylcholine-containing glycoprotein secreted by the filarial nematode Acanthocheilonema viteae, protects against kidney damage in an MRL/lpr mouse model of systemic lupus erythematosus (SLE). Methods MRL/lpr mice progressively produce high levels of autoantibodies, and the resultant deposition of immune complexes drives kidney pathology. The effects of ES-62 on disease progression were assessed by measurement of proteinuria, assessment of kidney histology, determination of antinuclear antibody (ANA) production and cytokine levels, and flow cytometric analysis of relevant cellular populations. Results ES-62 restored the disrupted balance between effector and regulatory B cells in MRL/lpr mice by inhibiting plasmablast differentiation, with a consequent reduction in ANA production and deposition of immune complexes and C3a in the kidneys. Moreover, by reducing interleukin-22 production, ES-62 may desensitize downstream effector mechanisms in the pathogenesis of kidney disease. Highlighting the therapeutic importance of resetting B cell responses, adoptive transfer of purified splenic B cells from ES-62–treated MRL/lpr mice mimicked the protection afforded by the helminth product. Mechanistically, this reflects down-regulation of myeloid differentiation factor 88 expression by B cells and also kidney cells, resulting in inhibition of pathogenic cross-talk among Toll-like receptor–, C3a-, and immune complex–mediated effector mechanisms. Conclusion This study provides the first demonstration of protection against kidney pathology by a parasitic worm–derived immunomodulator in a model of SLE and suggests therapeutic potential for drugs based on the mechanism of action of ES-62. PMID:25546822

  1. Initial fungal effector production is mediated by early endosome motility.

    PubMed

    Higuchi, Yujiro

    2015-01-01

    Fungal plant pathogenicity is facilitated by effector proteins that are specifically expressed during infection and are responsible for suppressing plant defense mechanisms. Recent studies have elucidated the detailed molecular mechanisms of effector action throughout fungal infection. However, little is known about the trafficking and secretion of effectors in fungal hyphae during the initial stage of infection. Using state-of-the-art microscopy we have demonstrated that early endosome (EE) motility is required for effector production during fungal infection. Moreover, the MAPK Crk1 has been shown to travel on EEs and to function as a negative regulator of effector expression, suggesting that motile EEs are involved in signal transduction. Here I further discuss possible mechanisms whereby EE motility regulates effector expression in the initial stages of infection. PMID:26480479

  2. 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.

  3. Robotic end effector

    DOEpatents

    Minichan, Richard L. (23 Pineview Dr., Warrenville, SC 29851)

    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.

  4. 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.

  5. The haustorial transcriptomes of Uromyces appendiculatus and Phakopsora pachyrhizi and their candidate effector families

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The haustoria of the biotrophic rust fungi are responsible for the uptake of nutrients from host cells, and they produce secreted proteins known as effectors that suppress host defenses. Effectors hold essential keys for elucidating the plant-fungal interactions, and they are promising targets for p...

  6. The Pseudomonas syringae type III effector HopG1 targets mitochondria, alters plant development and

    E-print Network

    pathogen Pseudomonas syrin- gae uses a type III protein secretion system to inject type III effectors- ringae pv. tomato DC3000 also suppresses PTI. Interestingly, HopG1 localizes to plant mitochon- dria effector (T3E) proteins into plant cells to promote pathogenicity (Alfano and Collmer, 2004). The T3SS

  7. T Cell Signaling Targets for Enhancing Regulatory or Effector Function

    PubMed Central

    Pan, Fan; Fan, Huimin; Liu, Zhongmin; Jiang, Shuiping

    2015-01-01

    To respond to infection, resting or naïve T cells must undergo activation, clonal expansion, and differentiation into specialized functional subsets of effector T cells. However, to prevent excessive or self-destructive immune responses, regulatory T cells (Tregs) are instrumental in suppressing the activation and function of effector cells, including effector T cells. The transcription factor Forkhead box P3 (Foxp3) regulates the expression of genes involved in the development and function of Tregs. Foxp3 interacts with other transcription factors and with epigenetic elements such as histone deacetylases (HDACs) and histone acetyltransferases. Treg suppressive function can be increased by exposure to HDAC inhibitors. The individual contributions of different HDAC family members to Treg function and their respective mechanisms of action, however, remain unclear. A study showed that HDAC6, HDAC9, and Sirtuin-1 had distinct effects on Foxp3 expression and function, suggesting that selectively targeting HDACs individually or in combination may enhance Treg stability and suppressive function. Another study showed that the receptor programmed death 1 (PD-1), a well-known inhibitor of T cell activation, halted cell cycle progression in effector T cells by inhibiting the transcription of the gene encoding the substrate-recognition component (Skp2) of the ubiquitin ligase SCFSkp2. Together, these findings reveal new signaling targets for enhancing Treg or effector T cell function that may be helpful in designing future therapies, either to increase Treg suppressive function in transplantation and autoimmune diseases or to block PD-1 function, thus increasing the magnitude of antiviral or antitumor immune responses of effector T cells. PMID:22855503

  8. Sequential Delivery of Host-Induced Virulence Effectors by Appressoria and Intracellular Hyphae of the Phytopathogen Colletotrichum higginsianum

    PubMed Central

    Kleemann, Jochen; Neumann, Ulla; van Themaat, Emiel Ver Loren; van der Does, H. Charlotte; Hacquard, Stéphane; Stüber, Kurt; Will, Isa; Schmalenbach, Wolfgang; Schmelzer, Elmon; O'Connell, Richard J.

    2012-01-01

    Phytopathogens secrete effector proteins to manipulate their hosts for effective colonization. Hemibiotrophic fungi must maintain host viability during initial biotrophic growth and elicit host death for subsequent necrotrophic growth. To identify effectors mediating these opposing processes, we deeply sequenced the transcriptome of Colletotrichum higginsianum infecting Arabidopsis. Most effector genes are host-induced and expressed in consecutive waves associated with pathogenic transitions, indicating distinct effector suites are deployed at each stage. Using fluorescent protein tagging and transmission electron microscopy-immunogold labelling, we found effectors localised to stage-specific compartments at the host-pathogen interface. In particular, we show effectors are focally secreted from appressorial penetration pores before host invasion, revealing new levels of functional complexity for this fungal organ. Furthermore, we demonstrate that antagonistic effectors either induce or suppress plant cell death. Based on these results we conclude that hemibiotrophy in Colletotrichum is orchestrated through the coordinated expression of antagonistic effectors supporting either cell viability or cell death. PMID:22496661

  9. Two-axis angular effector

    DOEpatents

    Vaughn, Mark R. (Albuquerque, NM); Robinett, III, Rush D. (Tijeras, NM); Phelan, John R. (Albuquerque, NM); Van Zuiden, Don M. (Albuquerque, NM)

    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.

  10. Long-distance endosome trafficking drives fungal effector production during plant infection.

    PubMed

    Bielska, Ewa; Higuchi, Yujiro; Schuster, Martin; Steinberg, Natascha; Kilaru, Sreedhar; Talbot, Nicholas J; Steinberg, Gero

    2014-01-01

    To cause plant disease, pathogenic fungi can secrete effector proteins into plant cells to suppress plant immunity and facilitate fungal infection. Most fungal pathogens infect plants using very long strand-like cells, called hyphae, that secrete effectors from their tips into host tissue. How fungi undergo long-distance cell signalling to regulate effector production during infection is not known. Here we show that long-distance retrograde motility of early endosomes (EEs) is necessary to trigger transcription of effector-encoding genes during plant infection by the pathogenic fungus Ustilago maydis. We demonstrate that motor-dependent retrograde EE motility is necessary for regulation of effector production and secretion during host cell invasion. We further show that retrograde signalling involves the mitogen-activated kinase Crk1 that travels on EEs and participates in control of effector production. Fungal pathogens therefore undergo long-range signalling to orchestrate host invasion. PMID:25283249

  11. Pseudomonas syringae type III secretion system effectors: repertoires in search of functions.

    PubMed

    Cunnac, Sébastien; Lindeberg, Magdalen; Collmer, Alan

    2009-02-01

    The ability of Pseudomonas syringae to grow and cause diseases in plants is dependent on the injection of multiple effector proteins into plant cells via the type III secretion system (T3SS). Genome-enabled bioinformatic/experimental methods have comprehensively identified the repertoires of effectors and related T3SS substrates for P. syringae pv. tomato DC3000 and three other sequenced strains. The effector repertoires are diverse and internally redundant. Insights into effector functions are being gained through the construction of mutants lacking one or more effector genes, which may be reduced in growth in planta, and through gain-of-function assays for the ability of single effectors to suppress plant innate immune defenses, manipulate hormone signaling, elicit cell death, and/or display biochemical activities on plant protein targets. PMID:19168384

  12. Long-distance endosome trafficking drives fungal effector production during plant infection

    PubMed Central

    Bielska, Ewa; Higuchi, Yujiro; Schuster, Martin; Steinberg, Natascha; Kilaru, Sreedhar; Talbot, Nicholas J.; Steinberg, Gero

    2014-01-01

    To cause plant disease, pathogenic fungi can secrete effector proteins into plant cells to suppress plant immunity and facilitate fungal infection. Most fungal pathogens infect plants using very long strand-like cells, called hyphae, that secrete effectors from their tips into host tissue. How fungi undergo long-distance cell signalling to regulate effector production during infection is not known. Here we show that long-distance retrograde motility of early endosomes (EEs) is necessary to trigger transcription of effector-encoding genes during plant infection by the pathogenic fungus Ustilago maydis. We demonstrate that motor-dependent retrograde EE motility is necessary for regulation of effector production and secretion during host cell invasion. We further show that retrograde signalling involves the mitogen-activated kinase Crk1 that travels on EEs and participates in control of effector production. Fungal pathogens therefore undergo long-range signalling to orchestrate host invasion. PMID:25283249

  13. In planta effector competition assays detect Hyaloperonospora arabidopsidis effectors that contribute to virulence and localize to different plant subcellular compartments.

    PubMed

    Badel, Jorge Luis; Piquerez, Sophie J M; Greenshields, David; Rallapalli, Ghanasyam; Fabro, Georgina; Ishaque, Naveed; Jones, Jonathan D G

    2013-07-01

    The genome of the pathogenic oomycete Hyaloperonospora arabidopsidis is predicted to encode at least 134 high-confidence effectors (HaRxL) carrying the RxLR motif implicated in their translocation into plant cells. However, only four avirulence genes (ATR1, ATR13, ATR5, and ATR39) have been isolated. This indicates that identification of HaRxL effectors based on avirulence is low throughput. We aimed at rapidly identifying H. arabidopsidis effectors that contribute to virulence by developing methods to detect and quantify multiple candidates in bacterial mixed infections using either Illumina sequencing or capillary electrophoresis. In these assays, referred to here as in planta effector competition assays, we estimate the contribution to virulence of individual effectors by calculating the abundance of each HaRxL in the bacterial population recovered from leaves 3 days after inoculation relative to abundance in the initial mixed inoculum. We identified HaRxL that enhance Pseudomonas syringae pv. tomato DC3000 growth in some but not all Arabidopsis accessions. Further analysis showed that HaRxLL464, HaRxL75, HaRxL22, HaRxLL441, and HaRxL89 suppress pathogen-associated molecular pattern-triggered immunity (PTI) and localize to different subcellular compartments in Nicotiana benthamiana, providing evidence for a multilayered suppression of PTI by pathogenic oomycetes and molecular probes for the dissection of PTI. PMID:23734779

  14. End Effector Coupler/Decoupler

    NASA Technical Reports Server (NTRS)

    Smith, Russell W.; Rhodes, Marvin D.

    1995-01-01

    Coupling/decoupling device offers benefits of both stiffness and repositioning. Permits end effector to be released (decoupled) from robot arm while attached to workpiece, and realigned and recoupled to arm when end effector task completed. Relieves necessity for accurate positioning of robot and workpiece, and compensates for deformations initiated by changes in temperature as well as normal machining variations.

  15. Orbital maneuvering end effectors

    NASA Technical Reports Server (NTRS)

    Myers, W. Neill (inventor); Forbes, John C. (inventor); Barnes, Wayne L. (inventor)

    1986-01-01

    This invention relates to an end effector device for grasping and maneuvering objects such as berthing handles of a space telescope. The device includes a V-shaped capture window defined as inclined surfaces in parallel face plates which converge toward a retainer recess in which the handle is retained. A pivotal finger (30) meshes with a pair of pivoted fingers which rotate in counterrotation. The fingers rotate to pull a handle within the capture window into recess where latches lock handle in the recess. To align the capture window, plates may be cocked plus or minus five degrees on base. Drive means is included in the form of a motor coupled with a harmonic drive speed reducer, which provides for slow movement of the fingers at a high torque so that large articles may be handled. Novelty of the invention is believed to reside in the combined intermeshing finger structure, drive means and the harmonic drive speed reducer, which features provide the required maneuverability and strength.

  16. A functional genomics approach identifies candidate effectors from the aphid species Myzus persicae (green peach aphid).

    PubMed

    Bos, Jorunn I B; Prince, David; Pitino, Marco; Maffei, Massimo E; Win, Joe; Hogenhout, Saskia A

    2010-11-01

    Aphids are amongst the most devastating sap-feeding insects of plants. Like most plant parasites, aphids require intimate associations with their host plants to gain access to nutrients. Aphid feeding induces responses such as clogging of phloem sieve elements and callose formation, which are suppressed by unknown molecules, probably proteins, in aphid saliva. Therefore, it is likely that aphids, like plant pathogens, deliver proteins (effectors) inside their hosts to modulate host cell processes, suppress plant defenses, and promote infestation. We exploited publicly available aphid salivary gland expressed sequence tags (ESTs) to apply a functional genomics approach for identification of candidate effectors from Myzus persicae (green peach aphid), based on common features of plant pathogen effectors. A total of 48 effector candidates were identified, cloned, and subjected to transient overexpression in Nicotiana benthamiana to assay for elicitation of a phenotype, suppression of the Pathogen-Associated Molecular Pattern (PAMP)-mediated oxidative burst, and effects on aphid reproductive performance. We identified one candidate effector, Mp10, which specifically induced chlorosis and local cell death in N. benthamiana and conferred avirulence to recombinant Potato virus X (PVX) expressing Mp10, PVX-Mp10, in N. tabacum, indicating that this protein may trigger plant defenses. The ubiquitin-ligase associated protein SGT1 was required for the Mp10-mediated chlorosis response in N. benthamiana. Mp10 also suppressed the oxidative burst induced by flg22, but not by chitin. Aphid fecundity assays revealed that in planta overexpression of Mp10 and Mp42 reduced aphid fecundity, whereas another effector candidate, MpC002, enhanced aphid fecundity. Thus, these results suggest that, although Mp10 suppresses flg22-triggered immunity, it triggers a defense response, resulting in an overall decrease in aphid performance in the fecundity assays. Overall, we identified aphid salivary proteins that share features with plant pathogen effectors and therefore may function as aphid effectors by perturbing host cellular processes. PMID:21124944

  17. A Functional Genomics Approach Identifies Candidate Effectors from the Aphid Species Myzus persicae (Green Peach Aphid)

    PubMed Central

    Bos, Jorunn I. B.; Prince, David; Pitino, Marco; Maffei, Massimo E.; Win, Joe; Hogenhout, Saskia A.

    2010-01-01

    Aphids are amongst the most devastating sap-feeding insects of plants. Like most plant parasites, aphids require intimate associations with their host plants to gain access to nutrients. Aphid feeding induces responses such as clogging of phloem sieve elements and callose formation, which are suppressed by unknown molecules, probably proteins, in aphid saliva. Therefore, it is likely that aphids, like plant pathogens, deliver proteins (effectors) inside their hosts to modulate host cell processes, suppress plant defenses, and promote infestation. We exploited publicly available aphid salivary gland expressed sequence tags (ESTs) to apply a functional genomics approach for identification of candidate effectors from Myzus persicae (green peach aphid), based on common features of plant pathogen effectors. A total of 48 effector candidates were identified, cloned, and subjected to transient overexpression in Nicotiana benthamiana to assay for elicitation of a phenotype, suppression of the Pathogen-Associated Molecular Pattern (PAMP)–mediated oxidative burst, and effects on aphid reproductive performance. We identified one candidate effector, Mp10, which specifically induced chlorosis and local cell death in N. benthamiana and conferred avirulence to recombinant Potato virus X (PVX) expressing Mp10, PVX-Mp10, in N. tabacum, indicating that this protein may trigger plant defenses. The ubiquitin-ligase associated protein SGT1 was required for the Mp10-mediated chlorosis response in N. benthamiana. Mp10 also suppressed the oxidative burst induced by flg22, but not by chitin. Aphid fecundity assays revealed that in planta overexpression of Mp10 and Mp42 reduced aphid fecundity, whereas another effector candidate, MpC002, enhanced aphid fecundity. Thus, these results suggest that, although Mp10 suppresses flg22-triggered immunity, it triggers a defense response, resulting in an overall decrease in aphid performance in the fecundity assays. Overall, we identified aphid salivary proteins that share features with plant pathogen effectors and therefore may function as aphid effectors by perturbing host cellular processes. PMID:21124944

  18. Recent Progress in RXLR Effector Research.

    PubMed

    Anderson, Ryan G; Deb, Devdutta; Fedkenheuer, Kevin; McDowell, John M

    2015-10-01

    Some of the most devastating oomycete pathogens deploy effector proteins, with the signature amino acid motif RXLR, that enter plant cells to promote virulence. Research on the function and evolution of RXLR effectors has been very active over the decade that has transpired since their discovery. Comparative genomics indicate that RXLR genes play a major role in virulence for Phytophthora and downy mildew species. Importantly, gene-for-gene resistance against these oomycete lineages is based on recognition of RXLR proteins. Comparative genomics have revealed several mechanisms through which this resistance can be broken, most notably involving epigenetic control of RXLR gene expression. Structural studies have revealed a core fold that is present in the majority of RXLR proteins, providing a foundation for detailed mechanistic understanding of virulence and avirulence functions. Finally, functional studies have demonstrated that suppression of host immunity is a major function for RXLR proteins. Host protein targets are being identified in a variety of plant cell compartments. Some targets comprise hubs that are also manipulated by bacteria and fungi, thereby revealing key points of vulnerability in the plant immune network. PMID:26125490

  19. How Diverse—CD4 Effector T Cells and their Functions

    PubMed Central

    Wan, Yisong Y.; Flavell, Richard A.

    2009-01-01

    CD4 effector T cells, also called helper T (Th) cells, are the functional cells for executing immune functions. Balanced immune responses can only be achieved by proper regulation of the differentiation and function of Th cells. Dysregulated Th cell function often leads to inefficient clearance of pathogens and causes inflammatory diseases and autoimmunity. Since the establishment of the Th1–Th2 dogma in the 1980s, different lineages of effector T cells have been identified that not only promote but also suppress immune responses. Through years of collective efforts, much information was gained on the function and regulation of different subsets of Th cells. In this review, we attempt to sample the essence of what has been learnt in this field over the past two decades. We will discuss the classification and immunological functions of effector T cells, the determinants for effector T cell differentiation, as well as the relationship between different lineages of effector T cells. PMID:19482777

  20. Single-cell quantification of IL-2 response by effector and regulatory T cells reveals critical plasticity in

    E-print Network

    Emonet, Thierry

    ) and effector (Teff) Tcells, whereby access to IL-2 can either increase the survival of Teff cells function of Treg cells, namely the specific suppression of survival signals for weakly activated Teff cells

  1. Distinct Pseudomonas type-III effectors use a cleavable transit peptide to target chloroplasts.

    PubMed

    Li, Guangyong; Froehlich, John E; Elowsky, Christian; Msanne, Joseph; Ostosh, Andrew C; Zhang, Chi; Awada, Tala; Alfano, James R

    2014-01-01

    The pathogen Pseudomonas syringae requires a type-III protein secretion system and the effector proteins it injects into plant cells for pathogenesis. The primary role for P. syringae type-III effectors is the suppression of plant immunity. The P. syringae pv. tomato DC3000 HopK1 type-III effector was known to suppress the hypersensitive response (HR), a programmed cell death response associated with effector-triggered immunity. Here we show that DC3000 hopK1 mutants are reduced in their ability to grow in Arabidopsis, and produce reduced disease symptoms. Arabidopsis transgenically expressing HopK1 are reduced in PAMP-triggered immune responses compared with wild-type plants. An N-terminal region of HopK1 shares similarity with the corresponding region in the well-studied type-III effector AvrRps4; however, their C-terminal regions are dissimilar, indicating that they have different effector activities. HopK1 is processed in planta at the same processing site found in AvrRps4. The processed forms of HopK1 and AvrRps4 are chloroplast localized, indicating that the shared N-terminal regions of these type-III effectors represent a chloroplast transit peptide. The HopK1 contribution to virulence and the ability of HopK1 and AvrRps4 to suppress immunity required their respective transit peptides, but the AvrRps4-induced HR did not. Our results suggest that a primary virulence target of these type-III effectors resides in chloroplasts, and that the recognition of AvrRps4 by the plant immune system occurs elsewhere. Moreover, our results reveal that distinct type-III effectors use a cleavable transit peptide to localize to chloroplasts, and that targets within this organelle are important for immunity. PMID:24299018

  2. Differential modulation of CNS-specific effector and regulatory T cells during tolerance induction by recombinant invariant chains in vivo.

    PubMed

    Lange, Christian; Doster, Hong; Steinbach, Karin; Kalbacher, Hubert; Scholl, Matthias; Melms, Arthur; Bischof, Felix

    2009-08-01

    Inflammation within the Central Nervous System (CNS) is largely controlled by the balance between CNS-specific effector and regulatory T lymphocytes. To suppress CNS-inflammation in an antigen-specific manner, CNS-specific effector and regulatory T cells thus have to be differentially regulated. We employed recombinant peptide/MHC class II tetramers to assess CNS-specific effector and regulatory T cells during the specific suppression of myelin proteolipid protein aa139-151 (PLP139-151)-induced experimental autoimmune encephalomyelitis (EAE) by intravenous injection of recombinant invariant chains (Ii) in which the CLIP region has been replaced by the PLP139-151 epitope (Ii-PLP139-151). Injection of Ii-PLP139-151 induced apoptosis in CNS-specific effector T cells. In contrast, the proportion of specific regulatory T cells was increased and these cells expressed larger amounts of molecules that mediate regulatory T cell function including transforming growth factor beta and the inducible costimulator (ICOS). Consequently, regulatory T cells from Ii-treated mice were more potent than regulatory T cells from control-treated animals in suppressing effector T cell proliferation. These data demonstrate that effector T cells and regulatory T cells directed against the same CNS-antigen can be differentially regulated in vivo to suppress CNS-autoimmunity. Recombinant Ii induce apoptosis in CNS-specific effector T cells and provoke qualitative changes in specific regulatory T cells that enhance their immunosuppressive properties. PMID:19362139

  3. Effector proteins of rust fungi

    PubMed Central

    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. PMID:25191335

  4. Improving a Gripper End Effector

    SciTech Connect

    Mullen, O Dennis; Smith, Christopher M.; Gervais, Kevin L.

    2001-01-31

    This paper discusses the improvement made to an existing four-bar linkage gripping end effector to adapt it for use in a current project. The actuating linkage was modified to yield higher jaw force overall and particularly in the critical range of jaw displacement

  5. Using hierarchical clustering of secreted protein families to classify and rank candidate effectors of rust fungi.

    PubMed

    Saunders, Diane G O; Win, Joe; Cano, Liliana M; Szabo, Les J; Kamoun, Sophien; Raffaele, Sylvain

    2012-01-01

    Rust fungi are obligate biotrophic pathogens that cause considerable damage on crop plants. Puccinia graminis f. sp. tritici, the causal agent of wheat stem rust, and Melampsora larici-populina, the poplar leaf rust pathogen, have strong deleterious impacts on wheat and poplar wood production, respectively. Filamentous pathogens such as rust fungi secrete molecules called disease effectors that act as modulators of host cell physiology and can suppress or trigger host immunity. Current knowledge on effectors from other filamentous plant pathogens can be exploited for the characterisation of effectors in the genome of recently sequenced rust fungi. We designed a comprehensive in silico analysis pipeline to identify the putative effector repertoire from the genome of two plant pathogenic rust fungi. The pipeline is based on the observation that known effector proteins from filamentous pathogens have at least one of the following properties: (i) contain a secretion signal, (ii) are encoded by in planta induced genes, (iii) have similarity to haustorial proteins, (iv) are small and cysteine rich, (v) contain a known effector motif or a nuclear localization signal, (vi) are encoded by genes with long intergenic regions, (vii) contain internal repeats, and (viii) do not contain PFAM domains, except those associated with pathogenicity. We used Markov clustering and hierarchical clustering to classify protein families of rust pathogens and rank them according to their likelihood of being effectors. Using this approach, we identified eight families of candidate effectors that we consider of high value for functional characterization. This study revealed a diverse set of candidate effectors, including families of haustorial expressed secreted proteins and small cysteine-rich proteins. This comprehensive classification of candidate effectors from these devastating rust pathogens is an initial step towards probing plant germplasm for novel resistance components. PMID:22238666

  6. Dexterous end effector flight demonstration

    NASA Technical Reports Server (NTRS)

    Carter, Edward L.; Monford, Leo G.

    1994-01-01

    The Dexterous End Effector Flight Experiment is a flight demonstration of newly developed equipment and methods which make for more dexterous manipulation of robotic arms. The following concepts are to be demonstrated: The Force Torque Sensor is a six axis load cell located at the end of the RMS which displays load data to the operator on the orbiter CCTV monitor. TRAC is a target system which provides six axis positional information to the operator. It has the characteristic of having high sensitivity to attitude misalignment while being flat. AUTO-TRAC is a variation of TRAC in which a computer analyzes a target, displays translational and attitude misalignment information, and provides cues to the operator for corrective inputs. The Magnetic End Effector is a fault tolerant end effector which grapples payloads using magnetic attraction. The Carrier Latch Assembly is a fault tolerant payload carrier, which uses mechanical latches and/or magnetic attraction to hold small payloads during launch/landing and to release payloads as desired. The flight experiment goals and objectives are explained. The experiment equipment is described, and the tasks to be performed during the demonstration are discussed.

  7. Pseudomonas syringae pv. Tomato DC3000 Type III secretion effector polymutants reveal an interplay between hopAD1 and AvrPtoB

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The model pathogen Pseudomonas syringae pv. tomato DC3000 suppresses the two-tiered innate immune system of plants by injecting a complex repertoire of effector proteins into host cells via the type III secretion system. The model effector AvrPtoB has multiple domains and plant protein interactors i...

  8. The N-terminal region of Pseudomonas type III effector AvrPtoB elicits Pto-dependent immunity and has two

    E-print Network

    Sheen, Jen

    with either of the sequence-dissimilar type III effector proteins AvrPto or AvrPtoB (HopAB2) from Pseudo of a battery of `effector' proteins, many of which act to suppress basal defenses (Alfano and Collmer, 2004. tomato DC3000, for ex

  9. Space Station end effector strategy study

    NASA Technical Reports Server (NTRS)

    Katzberg, Stephen J.; Jensen, Robert L.; Willshire, Kelli F.; Satterthwaite, Robert E.

    1987-01-01

    The results of a study are presented for terminology definition, identification of functional requirements, technolgy assessment, and proposed end effector development strategies for the Space Station Program. The study is composed of a survey of available or under-developed end effector technology, identification of requirements from baselined Space Station documents, a comparative assessment of the match between technology and requirements, and recommended strategies for end effector development for the Space Station Program.

  10. Novel Type III Effectors in Pseudomonas aeruginosa

    PubMed Central

    Burstein, David; Satanower, Shirley; Simovitch, Michal; Belnik, Yana; Zehavi, Meital; Yerushalmi, Gal; Ben-Aroya, Shay

    2015-01-01

    ABSTRACT Pseudomonas aeruginosa is a Gram-negative, opportunistic pathogen that causes chronic and acute infections in immunocompromised patients. Most P. aeruginosa strains encode an active type III secretion system (T3SS), utilized by the bacteria to deliver effector proteins from the bacterial cell directly into the cytoplasm of the host cell. Four T3SS effectors have been discovered and extensively studied in P. aeruginosa: ExoT, ExoS, ExoU, and ExoY. This is especially intriguing in light of P. aeruginosa’s ability to infect a wide range of hosts. We therefore hypothesized that additional T3SS effectors that have not yet been discovered are encoded in the genome of P. aeruginosa. Here, we applied a machine learning classification algorithm to identify novel P. aeruginosa effectors. In this approach, various types of data are integrated to differentiate effectors from the rest of the open reading frames of the bacterial genome. Due to the lack of a sufficient learning set of positive effectors, our machine learning algorithm integrated genomic information from another Pseudomonas species and utilized dozens of features accounting for various aspects of the effector coding genes and their products. Twelve top-ranking predictions were experimentally tested for T3SS-specific translocation, leading to the discovery of two novel T3SS effectors. We demonstrate that these effectors are not part of the injection structural complex and report initial efforts toward their characterization. PMID:25784698

  11. Structure and evolution of barley powdery mildew effector candidates

    PubMed Central

    2012-01-01

    Background Protein effectors of pathogenicity are instrumental in modulating host immunity and disease resistance. The powdery mildew pathogen of grasses Blumeria graminis causes one of the most important diseases of cereal crops. B. graminis is an obligate biotrophic pathogen and as such has an absolute requirement to suppress or avoid host immunity if it is to survive and cause disease. Results Here we characterise a superfamily predicted to be the full complement of Candidates for Secreted Effector Proteins (CSEPs) in the fungal barley powdery mildew parasite B. graminis f.sp. hordei. The 491 genes encoding these proteins constitute over 7% of this pathogen’s annotated genes and most were grouped into 72 families of up to 59 members. They were predominantly expressed in the intracellular feeding structures called haustoria, and proteins specifically associated with the haustoria were identified by large-scale mass spectrometry-based proteomics. There are two major types of effector families: one comprises shorter proteins (100–150 amino acids), with a high relative expression level in the haustoria and evidence of extensive diversifying selection between paralogs; the second type consists of longer proteins (300–400 amino acids), with lower levels of differential expression and evidence of purifying selection between paralogs. An analysis of the predicted protein structures underscores their overall similarity to known fungal effectors, but also highlights unexpected structural affinities to ribonucleases throughout the entire effector super-family. Candidate effector genes belonging to the same family are loosely clustered in the genome and are associated with repetitive DNA derived from retro-transposons. Conclusions We employed the full complement of genomic, transcriptomic and proteomic analyses as well as structural prediction methods to identify and characterize the members of the CSEPs superfamily in B. graminis f.sp. hordei. Based on relative intron position and the distribution of CSEPs with a ribonuclease-like domain in the phylogenetic tree we hypothesize that the associated genes originated from an ancestral gene, encoding a secreted ribonuclease, duplicated successively by repetitive DNA-driven processes and diversified during the evolution of the grass and cereal powdery mildew lineage. PMID:23231440

  12. Deployment of the Burkholderia glumae type III secretion system as an efficient tool for translocating pathogen effectors to monocot cells.

    PubMed

    Sharma, Shailendra; Sharma, Shiveta; Hirabuchi, Akiko; Yoshida, Kentaro; Fujisaki, Koki; Ito, Akiko; Uemura, Aiko; Terauchi, Ryohei; Kamoun, Sophien; Sohn, Kee Hoon; Jones, Jonathan D G; Saitoh, Hiromasa

    2013-05-01

    Genome sequences of plant fungal pathogens have enabled the identification of effectors that cooperatively modulate the cellular environment for successful fungal growth and suppress host defense. Identification and characterization of novel effector proteins are crucial for understanding pathogen virulence and host-plant defense mechanisms. Previous reports indicate that the Pseudomonas syringae pv. tomato DC3000 type III secretion system (T3SS) can be used to study how non-bacterial effectors manipulate dicot plant cell function using the effector detector vector (pEDV) system. Here we report a pEDV-based effector delivery system in which the T3SS of Burkholderia glumae, an emerging rice pathogen, is used to translocate the AVR-Pik and AVR-Pii effectors of the fungal pathogen Magnaporthe oryzae to rice cytoplasm. The translocated AVR-Pik and AVR-Pii showed avirulence activity when tested in rice cultivars containing the cognate R genes. AVR-Pik reduced and delayed the hypersensitive response triggered by B. glumae in the non-host plant Nicotiana benthamiana, indicative of an immunosuppressive virulence activity. AVR proteins fused with fluorescent protein and nuclear localization signal were delivered by B. glumae T3SS and observed in the nuclei of infected cells in rice, wheat, barley and N. benthamiana. Our bacterial T3SS-enabled eukaryotic effector delivery and subcellular localization assays provide a useful method for identifying and studying effector functions in monocot plants. PMID:23451734

  13. The interplay of effector and regulatory T cells in cancer.

    PubMed

    Roychoudhuri, Rahul; Eil, Robert L; Restifo, Nicholas P

    2015-04-01

    Regulatory T (Treg) cells suppress effector T (Teff) cells and prevent immune-mediated rejection of cancer. Much less appreciated are mechanisms by which Teff cells antagonize Treg cells. Herein, we consider how complex reciprocal interactions between Teff and Treg cells shape their population dynamics within tumors. Under states of tolerance, including during tumor escape, suppressed Teff cells support Treg cell populations through antigen-dependent provision of interleukin (IL)-2. During immune activation, Teff cells can lose this supportive capacity and directly antagonize Treg cell populations to neutralize their immunosuppressive function. While this latter state is rarely achieved spontaneously within tumors, we propose that therapeutic induction of immune activation has the potential to stably disrupt immunosuppressive population states resulting in durable cancer regression. PMID:25728990

  14. Tansley review Nematode effector proteins: an emerging

    E-print Network

    Hussey, Richard S.

    head that is used to pierce host plant cell walls to access host cell contents for ingestion effector regulation and delivery into host cells 3 III. Nematode effectors as probes of plant cell biology. Elling5 , Martin Wubben6 and Eric L. Davis7 1 Division of Plant Sciences and Bond Life Sciences Center

  15. Distinct Sets of Rab6 Effectors Contribute to ZW10- and COG-Dependent Golgi Homeostasis

    PubMed Central

    Majeed, Waqar; Liu, Shijie; Storrie, Brian

    2014-01-01

    The organization of the Golgi apparatus is determined in part by the interaction of Rab proteins and their diverse array of effectors. Here, we used multiple approaches to identify and characterize a small subset of effectors that mimicked the effects of Rab6 on Golgi ribbon organization. In a visual-based, candidate-protein screen, we found that the individual depletion of any of three Rab6 effectors, myosin IIA (MyoIIA), Kif20A, and Bicaudal D (BicD), was sufficient to suppress Golgi ribbon fragmentation/dispersal coupled to retrograde tether proteins in a manner paralleling Rab6. MyoIIA and Kif20A depletion were pathway selective and suppressed ZW10-dependent Golgi ribbon fragmentation/dispersal only while BicD depletion, like Rab6, suppressed both ZW10- and COG-dependent Golgi ribbon fragmentation. The MyoIIA effects could be produced in short term assays by the reversible myosin inhibitor, blebbistatin. At the electron microscope level, the effects of BicD-depletion mimicked many of those of Rab6-depletion: longer and more continuous Golgi cisternae and a pronounced accumulation of coated vesicles. Functionally, BicD-depleted cells were inhibited in transport of newly synthesized VSV-G protein to the cell surface. In sum, our results indicate small, partially overlapping subsets of Rab6 effectors are differentially important to two tether-dependent pathways essential to Golgi organization and function. PMID:24575842

  16. Orbital maneuvering vehicle end effectors

    NASA Technical Reports Server (NTRS)

    Myers, W. Neill (Inventor); Forbes, John C. (Inventor); Barnes, Wayne L. (Inventor)

    1988-01-01

    An end effector device (A) for grasping and holding an article such as a handle (18) of a space telescope is disclosed. The device includes a V-shaped capture window (74) defined as inclined surfaces (76, 78) in parallel face plates (22, 24) which converge toward a retainer recess (54) in which the handle is retained. A pivotal finger (30) meshes with a pair of pivoted fingers (26, 28) which rotate in counterrotation. The fingers rotate to pull a handle within the capture window into recess (54) where latches (50) lock handle (18) in the recess. To align the capture window, plates (22, 24) may be cocked plus or minus five degrees on base (64).

  17. ROBOTIC TANK INSPECTION END EFFECTOR

    SciTech Connect

    Rachel Landry

    1999-10-01

    The objective of this contract between Oceaneering Space Systems (OSS) and the Department of Energy (DOE) was to provide a tool for the DOE to inspect the inside tank walls of underground radioactive waste storage tanks in their tank farms. Some of these tanks are suspected to have leaks, but the harsh nature of the environment within the tanks precludes human inspection of tank walls. As a result of these conditions only a few inspection methods can fulfill this task. Of the methods available, OSS chose to pursue Alternating Current Field Measurement (ACFM), because it does not require clean surfaces for inspection, nor any contact with the Surface being inspected, and introduces no extra by-products in the inspection process (no coupling fluids or residues are left behind). The tool produced by OSS is the Robotic Tank Inspection End Effector (RTIEE), which is initially deployed on the tip of the Light Duty Utility Arm (LDUA). The RTEE combines ACFM with a color video camera for both electromagnetic and visual inspection The complete package consists of an end effector, its corresponding electronics and software, and a user's manual to guide the operator through an inspection. The system has both coarse and fine inspection modes and allows the user to catalog defects and suspected areas of leakage in a database for further examination, which may lead to emptying the tank for repair, decommissioning, etc.. The following is an updated report to OSS document OSS-21100-7002, which was submitted in 1995. During the course of the contract, two related subtasks arose, the Wall and Coating Thickness Sensor and the Vacuum Scarifying and Sampling Tool Assembly. The first of these subtasks was intended to evaluate the corrosion and wall thinning of 55-gallon steel drums. The second was retrieved and characterized the waste material trapped inside the annulus region of the underground tanks on the DOE's tank farms. While these subtasks were derived from the original intent of the contract, the focus remains on the RTIEE.

  18. Advances in experimental methods for the elucidation of Pseudomonas syringae effector function with a focus on AvrPtoB

    PubMed Central

    Munkvold, Kathy R.; Martin, Gregory B.

    2010-01-01

    SUMMARY Pseudomonas syringae infects a wide range of plant species through the use of a type III secretion system. The effector proteins injected into the plant cell through this molecular syringe serve as promoters of disease by subverting the plant immune response to the benefit of the bacteria in the intercellular space. The targets and activities of a subset of effectors have been elucidated recently. In this article, we focus on the experimental approaches that have proved most successful in probing the molecular basis of effectors, ranging from loss-of-function to gain-of-function analyses utilizing several techniques for effector delivery into plants. In particular, we highlight how these diverse approaches have been applied to the study of one effector—AvrPtoB—a multifunctional protein with the ability to suppress both effector-triggered immunity and pathogen (or microbe)-associated molecular pattern-triggered immunity. Taken together, advances in this field illustrate the need for multiple experimental approaches when elucidating the function of a single effector. PMID:19849784

  19. A TAL Effector Toolbox for Genome Engineering

    E-print Network

    Sanjana, Neville E.

    Transcription activator-like effectors (TALEs) are a class of naturally occurring DNA-binding proteins found in the plant pathogen Xanthomonas sp. The DNA-binding domain of each TALE consists of tandem 34–amino acid repeat ...

  20. The Salmonella effector protein SpvC, a phosphothreonine lyase is functional in plant cells

    PubMed Central

    Neumann, Christina; Fraiture, Malou; Hernàndez-Reyes, Casandra; Akum, Fidele N.; Virlogeux-Payant, Isabelle; Chen, Ying; Pateyron, Stephanie; Colcombet, Jean; Kogel, Karl-Heinz; Hirt, Heribert; Brunner, Frédéric; Schikora, Adam

    2014-01-01

    Salmonella is one of the most prominent causes of food poisoning and growing evidence indicates that contaminated fruits and vegetables are an increasing concern for human health. Successful infection demands the suppression of the host immune system, which is often achieved via injection of bacterial effector proteins into host cells. In this report we present the function of Salmonella effector protein in plant cell, supporting the new concept of trans-kingdom competence of this bacterium. We screened a range of Salmonella Typhimurium effector proteins for interference with plant immunity. Among these, the phosphothreonine lyase SpvC attenuated the induction of immunity-related genes when present in plant cells. Using in vitro and in vivo systems we show that this effector protein interacts with and dephosphorylates activated Arabidopsis Mitogen-activated Protein Kinase 6 (MPK6), thereby inhibiting defense signaling. Moreover, the requirement of Salmonella SpvC was shown by the decreased proliferation of the ?spvC mutant in Arabidopsis plants. These results suggest that some Salmonella effector proteins could have a conserved function during proliferation in different hosts. The fact that Salmonella and other Enterobacteriaceae use plants as hosts strongly suggests that plants represent a much larger reservoir for animal pathogens than so far estimated. PMID:25368608

  1. 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.

  2. 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.

  3. Differences and Similarities of Soybean Defense-Related Genes Suppressed by Pathogenic and Symbiotic Bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacterial effector proteins secreted through type III secretion systems (T3SS) play a crucial role in establishing plant and human diseases. Type III effectors have been shown to trigger defense responses when recognized by resistant plants, and to suppress defense responses in susceptible host plan...

  4. Genetic disassembly and combinatorial reassembly identify a minimal functional repertoire of type III effectors in Pseudomonas syringae.

    PubMed

    Cunnac, Sébastien; Chakravarthy, Suma; Kvitko, Brian H; Russell, Alistair B; Martin, Gregory B; Collmer, Alan

    2011-02-15

    The virulence of Pseudomonas syringae and many other proteobacterial pathogens is dependent on complex repertoires of effector proteins injected into host cells by type III secretion systems. The 28 well-expressed effector genes in the repertoire of the model pathogen P. syringae pv. tomato DC3000 were deleted to produce polymutant DC3000D28E. Growth of DC3000D28E in Nicotiana benthamiana was symptomless and 4 logs lower than that of DC3000?hopQ1-1, which causes disease in this model plant. DC3000D28E seemed functionally effectorless but otherwise WT in diagnostic phenotypes relevant to plant interactions (for example, ability to inject the AvrPto-Cya reporter into N. benthamiana). Various effector genes were integrated by homologous recombination into native loci or by a programmable or random in vivo assembly shuttle (PRIVAS) system into the exchangeable effector locus in the Hrp pathogenicity island of DC3000D28E. The latter method exploited dual adapters and recombination in yeast for efficient assembly of PCR products into programmed or random combinations of multiple effector genes. Native and PRIVAS-mediated integrations were combined to identify a minimal functional repertoire of eight effector genes that restored much of the virulence of DC3000?hopQ1-1 in N. benthamiana, revealing a hierarchy in effector function: AvrPtoB acts with priority in suppressing immunity, enabling other effectors to promote further growth (HopM1 and HopE1), chlorosis (HopG1), lesion formation (HopAM1-1), and near full growth and symptom production (AvrE, HopAA1-1, and/or HopN1 functioning synergistically with the previous effectors). DC3000D28E, the PRIVAS method, and minimal functional repertoires provide new resources for probing the plant immune system. PMID:21282655

  5. Transcriptional Dynamics Driving MAMP-Triggered Immunity and Pathogen Effector-Mediated Immunosuppression in Arabidopsis Leaves Following Infection with Pseudomonas syringae pv tomato DC3000[OPEN

    PubMed Central

    Lewis, Laura A.; Polanski, Krzysztof; de Torres-Zabala, Marta; Bowden, Laura; Jenkins, Dafyd J.; Hill, Claire; Baxter, Laura; Truman, William; Prusinska, Justyna; Hickman, Richard; Wild, David L.; Ott, Sascha; Buchanan-Wollaston, Vicky; Beynon, Jim

    2015-01-01

    Transcriptional reprogramming is integral to effective plant defense. Pathogen effectors act transcriptionally and posttranscriptionally to suppress defense responses. A major challenge to understanding disease and defense responses is discriminating between transcriptional reprogramming associated with microbial-associated molecular pattern (MAMP)-triggered immunity (MTI) and that orchestrated by effectors. A high-resolution time course of genome-wide expression changes following challenge with Pseudomonas syringae pv tomato DC3000 and the nonpathogenic mutant strain DC3000hrpA- allowed us to establish causal links between the activities of pathogen effectors and suppression of MTI and infer with high confidence a range of processes specifically targeted by effectors. Analysis of this information-rich data set with a range of computational tools provided insights into the earliest transcriptional events triggered by effector delivery, regulatory mechanisms recruited, and biological processes targeted. We show that the majority of genes contributing to disease or defense are induced within 6 h postinfection, significantly before pathogen multiplication. Suppression of chloroplast-associated genes is a rapid MAMP-triggered defense response, and suppression of genes involved in chromatin assembly and induction of ubiquitin-related genes coincide with pathogen-induced abscisic acid accumulation. Specific combinations of promoter motifs are engaged in fine-tuning the MTI response and active transcriptional suppression at specific promoter configurations by P. syringae. PMID:26566919

  6. 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

  7. End effector with astronaut foot restraint

    NASA Technical Reports Server (NTRS)

    Monford, Leo G., Jr. (inventor)

    1991-01-01

    The combination of a foot restraint platform designed primarily for use by an astronaut being rigidly and permanently attached to an end effector which is suitable for attachment to the manipulator arm of a remote manipulating system is described. The foot restraint platform is attached by a brace to the end effector at a location away from the grappling interface of the end effector. The platform comprises a support plate provided with a pair of stirrups for receiving the toe portion of an astronaut's boots when standing on the platform and a pair of heel retainers in the form of raised members which are fixed to the surface of the platform and located to provide abutment surfaces for abutting engagement with the heels of the astronaut's boots when his toes are in the stirrups. The heel retainers preclude a backward sliding movement of the feet on the platform and instead require a lifting of the heels in order to extract the feet. The brace for attaching the foot restraint platform to the end effector may include a pivot or swivel joint to permit various orientations of the platform with respect to the end effector.

  8. Gene duplication and fragment recombination drive functional diversification of a superfamily of cytoplasmic effectors in Phytophthora sojae.

    PubMed

    Shen, Danyu; Liu, Tingli; Ye, Wenwu; Liu, Li; Liu, Peihan; Wu, Yuren; Wang, Yuanchao; Dou, Daolong

    2013-01-01

    Phytophthora and other oomycetes secrete a large number of putative host cytoplasmic effectors with conserved FLAK motifs following signal peptides, termed crinkling and necrosis inducing proteins (CRN), or Crinkler. Here, we first investigated the evolutionary patterns and mechanisms of CRN effectors in Phytophthora sojae and compared them to two other Phytophthora species. The genes encoding CRN effectors could be divided into 45 orthologous gene groups (OGG), and most OGGs unequally distributed in the three species, in which each underwent large number of gene gains or losses, indicating that the CRN genes expanded after species evolution in Phytophthora and evolved through pathoadaptation. The 134 expanded genes in P. sojae encoded family proteins including 82 functional genes and expressed at higher levels while the other 68 genes encoding orphan proteins were less expressed and contained 50 pseudogenes. Furthermore, we demonstrated that most expanded genes underwent gene duplication or/and fragment recombination. Three different mechanisms that drove gene duplication or recombination were identified. Finally, the expanded CRN effectors exhibited varying pathogenic functions, including induction of programmed cell death (PCD) and suppression of PCD through PAMP-triggered immunity or/and effector-triggered immunity. Overall, these results suggest that gene duplication and fragment recombination may be two mechanisms that drive the expansion and neofunctionalization of the CRN family in P. sojae, which aids in understanding the roles of CRN effectors within each oomycete pathogen. PMID:23922898

  9. Immune homeostasis enforced by co-localized effector and regulatory T cells

    PubMed Central

    Liu, Zhiduo; Gerner, Michael Y.; Van Panhuys, Nicholas; Levine, Andrew G.; Rudensky, Alexander Y.; Germain, Ronald N.

    2015-01-01

    Foxp3+ regulatory T cells (Tregs) play a critical role in preventing autoimmune disease by limiting the effector activity of conventional T cells that have escaped thymic negative selection or cell-autonomous peripheral inactivation1–3. However, despite the substantial information available about the molecular players mediating Treg functional interference with auto-aggressive effector responses4,5, the relevant cellular events in intact tissues remain largely unexplored and the issues of whether Tregs prevent activation of self-specific T cells or function primarily to limit damage from such cells have not been addressed6. Here we have employed multiplex, high-resolution, quantitative imaging to reveal that within most secondary lymphoid tissues, Tregs expressing phosphorylated STAT5 (pSTAT5) and high amounts of the suppressive molecules CD73 and CTLA-4 exist in discrete clusters with rare IL-2 producing effector T cells activated by self-antigens. This local IL-2 production induces the STAT5 phosphorylation in the Tregs and is part of a feedback circuit that augments the suppressive properties of the Tregs to limit further autoimmune responses. Inducible ablation of TCR expression by Tregs reduces their regulatory capacity and disrupts their localization in such clusters, resulting in uncontrolled effector T cell responses. Our data thus reveal that autoreactive T cells reach a state of activation and cytokine gene induction on a regular basis, with physically co-clustering, TCR-stimulated Tregs responding to this activation in a feedback manner to suppress incipient autoimmunity and maintain immune homeostasis. PMID:26605524

  10. The downy mildew effector proteins ATR1 and ATR13 promote disease susceptibility in Arabidopsis thaliana.

    PubMed

    Sohn, Kee Hoon; Lei, Rita; Nemri, Adnane; Jones, Jonathan D G

    2007-12-01

    The downy mildew (Hyaloperonospora parasitica) effector proteins ATR1 and ATR13 trigger RPP1-Nd/WsB- and RPP13-Nd-dependent resistance, respectively, in Arabidopsis thaliana. To better understand the functions of these effectors during compatible and incompatible interactions of H. parasitica isolates on Arabidopsis accessions, we developed a novel delivery system using Pseudomonas syringae type III secretion via fusions of ATRs to the N terminus of the P. syringae effector protein, AvrRPS4. ATR1 and ATR13 both triggered the hypersensitive response (HR) and resistance to bacterial pathogens in Arabidopsis carrying RPP1-Nd/WsB or RPP13-Nd, respectively, when delivered from P. syringae pv tomato (Pst) DC3000. In addition, multiple alleles of ATR1 and ATR13 confer enhanced virulence to Pst DC3000 on susceptible Arabidopsis accessions. We conclude that ATR1 and ATR13 positively contribute to pathogen virulence inside host cells. Two ATR13 alleles suppressed bacterial PAMP (for Pathogen-Associated Molecular Patterns)-triggered callose deposition in susceptible Arabidopsis when delivered by DC3000 DeltaCEL mutants. Furthermore, expression of another allele of ATR13 in plant cells suppressed PAMP-triggered reactive oxygen species production in addition to callose deposition. Intriguingly, although Wassilewskija (Ws-0) is highly susceptible to H. parasitica isolate Emco5, ATR13Emco5 when delivered by Pst DC3000 triggered localized immunity, including HR, on Ws-0. We suggest that an additional H. parasitica Emco5 effector might suppress ATR13-triggered immunity. PMID:18165328

  11. EZH2 is crucial for both differentiation of regulatory T cells and T effector cell expansion.

    PubMed

    Yang, Xiang-Ping; Jiang, Kan; Hirahara, Kiyoshi; Vahedi, Golnaz; Afzali, Behdad; Sciume, Giuseppe; Bonelli, Michael; Sun, Hong-Wei; Jankovic, Dragana; Kanno, Yuka; Sartorelli, Vittorio; O'Shea, John J; Laurence, Arian

    2015-01-01

    The roles of EZH2 in various subsets of CD4(+) T cells are controversial and its mechanisms of action are incompletely understood. FOXP3-positive Treg cells are a critical helper T cell subset, and dysregulation of Treg generation or function results in systemic autoimmunity. FOXP3 associates with EZH2 to mediate gene repression and suppressive function. Herein, we demonstrate that deletion of Ezh2 in CD4 T cells resulted in reduced numbers of Treg cells in vivo and differentiation in vitro and an increased proportion of memory CD4 T cells in part due to exaggerated production of effector cytokines. Furthermore, we found that both Ezh2-deficient Treg cells and T effector cells were functionally impaired in vivo: Tregs failed to constrain autoimmune colitis and T effector cells neither provided a protective response to T. gondii infection nor mediated autoimmune colitis. The dichotomous function of EZH2 in regulating differentiation and senescence in effector and regulatory T cells helps to explain the apparent existing contradictions in literature. PMID:26090605

  12. EZH2 is crucial for both differentiation of regulatory T cells and T effector cell expansion

    PubMed Central

    Yang, Xiang-Ping; Jiang, Kan; Hirahara, Kiyoshi; Vahedi, Golnaz; Afzali, Behdad; Sciume, Giuseppe; Bonelli, Michael; Sun, Hong-Wei; Jankovic, Dragana; Kanno, Yuka; Sartorelli, Vittorio; O’Shea, John J.; Laurence, Arian

    2015-01-01

    The roles of EZH2 in various subsets of CD4+ T cells are controversial and its mechanisms of action are incompletely understood. FOXP3-positive Treg cells are a critical helper T cell subset, and dysregulation of Treg generation or function results in systemic autoimmunity. FOXP3 associates with EZH2 to mediate gene repression and suppressive function. Herein, we demonstrate that deletion of Ezh2 in CD4 T cells resulted in reduced numbers of Treg cells in vivo and differentiation in vitro and an increased proportion of memory CD4 T cells in part due to exaggerated production of effector cytokines. Furthermore, we found that both Ezh2-deficient Treg cells and T effector cells were functionally impaired in vivo: Tregs failed to constrain autoimmune colitis and T effector cells neither provided a protective response to T. gondii infection nor mediated autoimmune colitis. The dichotomous function of EZH2 in regulating differentiation and senescence in effector and regulatory T cells helps to explain the apparent existing contradictions in literature. PMID:26090605

  13. Combover/CG10732, a Novel PCP Effector for Drosophila Wing Hair Formation

    PubMed Central

    Fagan, Jeremy K.; Dollar, Gretchen; Lu, Qiuheng; Barnett, Austen; Pechuan Jorge, Joaquin; Schlosser, Andreas; Pfleger, Cathie; Adler, Paul; Jenny, Andreas

    2014-01-01

    The polarization of cells is essential for the proper functioning of most organs. Planar Cell Polarity (PCP), the polarization within the plane of an epithelium, is perpendicular to apical-basal polarity and established by the non-canonical Wnt/Fz-PCP signaling pathway. Within each tissue, downstream PCP effectors link the signal to tissue specific readouts such as stereocilia orientation in the inner ear and hair follicle orientation in vertebrates or the polarization of ommatidia and wing hairs in Drosophila melanogaster. Specific PCP effectors in the wing such as Multiple wing hairs (Mwh) and Rho Kinase (Rok) are required to position the hair at the correct position and to prevent ectopic actin hairs. In a genome-wide screen in vitro, we identified Combover (Cmb)/CG10732 as a novel Rho kinase substrate. Overexpression of Cmb causes the formation of a multiple hair cell phenotype (MHC), similar to loss of rok and mwh. This MHC phenotype is dominantly enhanced by removal of rok or of other members of the PCP effector gene family. Furthermore, we show that Cmb physically interacts with Mwh, and cmb null mutants suppress the MHC phenotype of mwh alleles. Our data indicate that Cmb is a novel PCP effector that promotes to wing hair formation, a function that is antagonized by Mwh. PMID:25207969

  14. Minimal Mimicry: Mere Effector Matching Induces Preference

    ERIC Educational Resources Information Center

    Sparenberg, Peggy; Topolinski, Sascha; Springer, Anne; Prinz, Wolfgang

    2012-01-01

    Both mimicking and being mimicked induces preference for a target. The present experiments investigate the minimal sufficient conditions for this mimicry-preference link to occur. We argue that mere effector matching between one's own and the other person's movement is sufficient to induce preference, independent of which movement is actually…

  15. Molecular Cell Gravin Is a Transitory Effector

    E-print Network

    Scott, John D.

    Molecular Cell Article Gravin Is a Transitory Effector of Polo-like Kinase 1 during Cell Division at the plasma membrane by seques- tering protein kinases A and C with G protein- coupled receptors phosphory- lates Gravin on threonine 766 to prime the recruit- ment of the polo-like kinase Plk1 at defined

  16. Host specific toxins; effectors of necrotrophic pathogenicity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Host-specific toxins are defined as pathogen effectors that induce toxicity and promote disease only in the host species and only in cultivars of that host, and with few exceptions, expressing a specific dominant susceptibility gene. They are a feature of a small but well studied group of fungal pl...

  17. 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. PMID:26472741

  18. Kinematic evaluation of end effector design

    NASA Astrophysics Data System (ADS)

    Edwards, Gary W.

    1992-09-01

    The complex, many degree-of-freedom end effectors at the leading edge of technology would be unusable in the sea bottom research environment. Simpler designs are required to provide adequate reliability for subsea use. This work examines selection of end effector designs to achieve optimum grasping ability with minimal mechanical complexity. A new method of calculating grasp stability is developed, incorporating elements of previous works in the field. Programs are developed which evaluate the ability of different end effector configurations to grasp representative objects (a cube, sphere, and infinite cylinder). End effector designs considered had circular palms with fingers located at the periphery, oriented so that each pointed to the center of the palm. The program tested configurations of from 1 to 4 fingers and from 1 to 3 links per finger. Three sets of finger proportions were considered: equal length links, half length links, and anthropomorphic proportions. The 2 finger, 2 link per finger configuration was determined to be the optimum design, and the half length proportions were selected as the best set of proportions.

  19. 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

  20. Elevated Temperature Differentially Influences Effector-Triggered Immunity Outputs in Arabidopsis

    PubMed Central

    Menna, Alexandra; Nguyen, Dang; Guttman, David S.; Desveaux, Darrell

    2015-01-01

    Pseudomonas syringae is a Gram-negative bacterium that infects multiple plant species by manipulating cellular processes via injection of type three secreted effectors (T3SEs) into host cells. Nucleotide-binding leucine-rich repeat (NLR) resistance (R) proteins recognize specific T3SEs and trigger a robust immune response, called effector-triggered immunity (ETI), which limits pathogen proliferation and is often associated with localized programmed cell death, known as the hypersensitive response (HR). In this study, we examine the influence of elevated temperature on two ETI outputs: HR and pathogen virulence suppression. We found that in the Arabidopsis thaliana accession Col-0, elevated temperatures suppress the HR, but have minimal influence on ETI-associated P. syringae virulence suppression, thereby uncoupling these two ETI responses. We also identify accessions of Arabidopsis that exhibit impaired P. syringae virulence suppression at elevated temperature, highlighting the natural variation that exists in coping with biotic and abiotic stresses. These results not only reinforce the influence of abiotic factors on plant immunity but also emphasize the importance of carefully documented environmental conditions in studies of plant immunity. PMID:26617631

  1. Impact of end effector technology on telemanipulation performance

    NASA Technical Reports Server (NTRS)

    Bejczy, A. K.; Szakaly, Z.; Ohm, T.

    1990-01-01

    Generic requirements for end effector design are briefly summarized as derived from generic functional and operational requirements. Included is a brief summary of terms and definitions related to end effector technology. The second part contains a brief overview of end effector technology work as JPL during the past ten years, with emphasis on the evolution of new mechanical, sensing and control capabilities of end effectors. The third and major part is devoted to the description of current end effector technology. The ongoing work addresses mechanical, sensing and control details with emphasis on mechanical ruggedness, increased resolution in sensing, and close electronic and control integration with overall telemanipulator control system.

  2. A central role for Notch in effector CD8+ T cell differentiation

    PubMed Central

    Backer, Ronald A.; Helbig, Christina; Gentek, Rebecca; Kent, Andrew; Laidlaw, Brian J.; Dominguez, Claudia X.; de Souza, Yevan S.; van Trierum, Stella E.; van Beek, Ruud; Rimmelzwaan, Guus F.; ten Brinke, Anja; Willemsen, A. Marcel; van Kampen, Antoine H. C.; Kaech, Susan M.; Blander, J. Magarian; van Gisbergen, Klaas; Amsen, Derk

    2014-01-01

    Activated CD8+ T cells choose between terminal effector cell (TEC) or memory precursor cell (MPC) fates. We show that Notch controls this choice. Notch promoted differentiation of immediately protective TECs and was correspondingly required for clearance of an acute influenza virus infection. Notch activated a major portion of the TEC-specific gene expression program and suppressed the MPC-specific program. Expression of Notch receptors was induced on naïve CD8+ T cells by inflammatory mediators and interleukin 2 (IL-2) via mTOR and T-bet dependent pathways. These pathways were subsequently amplified downstream of Notch, creating a positive feedback loop. Notch thus functions as a central hub where information from different sources converges to match effector T cell differentiation to the demands of the infection. PMID:25344724

  3. Survivin as a global target of intrinsic tumor suppression networks.

    PubMed

    Guha, Minakshi; Altieri, Dario C

    2009-09-01

    Despite the constant exposure to genomic insults that may lead to malignancy, cancer is surprisingly a relatively rare occurrence, and this is largely credited to an elaborate network of endogenous tumor suppression. Many effectors of tumor suppression have been identified, and their functions when activated in damaged cells have in large part been elucidated. What is less clear is whether there are common target gene(s) of tumor suppression, whose expression must be ablated in order to block transformation and preserve cellular homeostasis. Fresh experimental evidence suggests that silencing of the mitotic regulator and cell death inhibitor, survivin, is a universal requirement for successful tumor suppression in humans. PMID:19717980

  4. Effector-triggered immunity blocks pathogen degradation of an immunity-associated vesicle traffic regulator in Arabidopsis.

    PubMed

    Nomura, Kinya; Mecey, Christy; Lee, Young-Nam; Imboden, Lori Alice; Chang, Jeff H; He, Sheng Yang

    2011-06-28

    Innate immunity in plants can be triggered by microbe- and pathogen-associated molecular patterns. The pathogen-associated molecular pattern-triggered immunity (PTI) is often suppressed by pathogen effectors delivered into the host cell. Plants can overcome pathogen suppression of PTI and reestablish pathogen resistance through effector-triggered immunity (ETI). An unanswered question is how plants might overcome pathogen-suppression of PTI during ETI. Findings described in this paper suggest a possible mechanism. During Pseudomonas syringae pathovar tomato (Pst) DC3000 infection of Arabidopsis, a host ADP ribosylation factor guanine nucleotide exchange factor, AtMIN7, is destabilized by the pathogen effector HopM1 through the host 26S proteasome. In this study, we discovered that AtMIN7 is required for not only PTI, consistent with the notion that Pst DC3000 degrades AtMIN7 to suppress PTI, but also ETI. The AtMIN7 level in healthy plants is low, but increases posttranscriptionally in response to activation of PTI. Whereas DC3000 infection led to degradation of AtMIN7, activation of ETI by three different effectors, AvrRpt2, AvrPphB, and HopA1, in Col-0 plants blocks the ability of Pst DC3000 to destabilize AtMIN7. Further analyses of bacterial translocation of HopM1 and AtMIN7 stability in HopM1 transgenic plants show that ETI prevents HopM1-mediated degradation of AtMIN7 inside the plant cell. Both AtMIN7 and HopM1 are localized to the trans-Golgi network/early endosome, a subcellular compartment that is not previously known to be associated with bacterial pathogenesis in plants. Thus, blocking pathogen degradation of trans-Golgi network/early endosome-associated AtMIN7 is a critical part of the ETI mechanism to counter bacterial suppression of PTI. PMID:21670267

  5. Novel Control Effectors for Truss Braced Wing

    NASA Technical Reports Server (NTRS)

    White, Edward V.; Kapania, Rakesh K.; Joshi, Shiv

    2015-01-01

    At cruise flight conditions very high aspect ratio/low sweep truss braced wings (TBW) may be subject to design requirements that distinguish them from more highly swept cantilevered wings. High aspect ratio, short chord length and relative thinness of the airfoil sections all contribute to relatively low wing torsional stiffness. This may lead to aeroelastic issues such as aileron reversal and low flutter margins. In order to counteract these issues, high aspect ratio/low sweep wings may need to carry additional high speed control effectors to operate when outboard ailerons are in reversal and/or must carry additional structural weight to enhance torsional stiffness. The novel control effector evaluated in this study is a variable sweep raked wing tip with an aileron control surface. Forward sweep of the tip allows the aileron to align closely with the torsional axis of the wing and operate in a conventional fashion. Aft sweep of the tip creates a large moment arm from the aileron to the wing torsional axis greatly enhancing aileron reversal. The novelty comes from using this enhanced and controllable aileron reversal effect to provide roll control authority by acting as a servo tab and providing roll control through intentional twist of the wing. In this case the reduced torsional stiffness of the wing becomes an advantage to be exploited. The study results show that the novel control effector concept does provide roll control as described, but only for a restricted class of TBW aircraft configurations. For the configuration studied (long range, dual aisle, Mach 0.85 cruise) the novel control effector provides significant benefits including up to 12% reduction in fuel burn.

  6. Altered T Cell Memory and Effector Cell Development in Chronic Lymphatic Filarial Infection That Is Independent of Persistent Parasite Antigen

    PubMed Central

    Steel, Cathy; Nutman, Thomas B.

    2011-01-01

    Chronic lymphatic filarial (LF) infection is associated with suppression of parasite-specific T cell responses that persist even following elimination of infection. While several mechanisms have been implicated in mediating this T cell specific downregulation, a role for alterations in the homeostasis of T effector and memory cell populations has not been explored. Using multiparameter flow cytometry, we investigated the role of persistent filarial infection on the maintenance of T cell memory in patients from the filarial-endemic Cook Islands. Compared to filarial-uninfected endemic normals (EN), microfilaria (mf) positive infected patients (Inf) had a reduced CD4 central memory (TCM) compartment. In addition, Inf patients tended to have more effector memory cells (TEM) and fewer effector cells (TEFF) than did ENs giving significantly smaller TEFF ? TEM ratios. These contracted TCM and TEFF populations were still evident in patients previously mf+ who had cleared their infection (CLInf). Moreover, the density of IL-7R?, necessary for T memory cell maintenance (but decreased in T effector cells), was significantly higher on memory cells of Inf and CLInf patients, although there was no evidence for decreased IL-7 or increased soluble IL7-R?, both possible mechanisms for signaling defects in memory cells. However, effector cells that were present in Inf and CLInf patients had lower percentages of HLA-DR suggesting impaired function. These changes in T cell populations appear to reflect chronicity of infection, as filarial-infected children, despite the presence of active infection, did not show alterations in the frequencies of these T cell phenotypes. These data indicate that filarial-infected patients have contracted TCM compartments and a defect in effector cell development, defects that persist even following clearance of infection. The fact that these global changes in memory and effector cell compartments do not yet occur in infected children makes early treatment of LF even more crucial. PMID:21559422

  7. Expression Profiling during Arabidopsis/Downy Mildew Interaction Reveals a Highly-Expressed Effector That Attenuates Responses to Salicylic Acid

    PubMed Central

    Asai, Shuta; Caillaud, Marie-Cécile; Furzer, Oliver J.; Ishaque, Naveed; Wirthmueller, Lennart; Fabro, Georgina; Shirasu, Ken; Jones, Jonathan D. G.

    2014-01-01

    Plants have evolved strong innate immunity mechanisms, but successful pathogens evade or suppress plant immunity via effectors delivered into the plant cell. Hyaloperonospora arabidopsidis (Hpa) causes downy mildew on Arabidopsis thaliana, and a genome sequence is available for isolate Emoy2. Here, we exploit the availability of genome sequences for Hpa and Arabidopsis to measure gene-expression changes in both Hpa and Arabidopsis simultaneously during infection. Using a high-throughput cDNA tag sequencing method, we reveal expression patterns of Hpa predicted effectors and Arabidopsis genes in compatible and incompatible interactions, and promoter elements associated with Hpa genes expressed during infection. By resequencing Hpa isolate Waco9, we found it evades Arabidopsis resistance gene RPP1 through deletion of the cognate recognized effector ATR1. Arabidopsis salicylic acid (SA)-responsive genes including PR1 were activated not only at early time points in the incompatible interaction but also at late time points in the compatible interaction. By histochemical analysis, we found that Hpa suppresses SA-inducible PR1 expression, specifically in the haustoriated cells into which host-translocated effectors are delivered, but not in non-haustoriated adjacent cells. Finally, we found a highly-expressed Hpa effector candidate that suppresses responsiveness to SA. As this approach can be easily applied to host-pathogen interactions for which both host and pathogen genome sequences are available, this work opens the door towards transcriptome studies in infection biology that should help unravel pathogen infection strategies and the mechanisms by which host defense responses are overcome. PMID:25329884

  8. Identification of Two Legionella pneumophila Effectors that Manipulate Host Phospholipids Biosynthesis

    PubMed Central

    Viner, Ram; Chetrit, David; Ehrlich, Marcelo; Segal, Gil

    2012-01-01

    The intracellular pathogen Legionella pneumophila translocates a large number of effector proteins into host cells via the Icm/Dot type-IVB secretion system. Some of these effectors were shown to cause lethal effect on yeast growth. Here we characterized one such effector (LecE) and identified yeast suppressors that reduced its lethal effect. The LecE lethal effect was found to be suppressed by the over expression of the yeast protein Dgk1 a diacylglycerol (DAG) kinase enzyme and by a deletion of the gene encoding for Pah1 a phosphatidic acid (PA) phosphatase that counteracts the activity of Dgk1. Genetic analysis using yeast deletion mutants, strains expressing relevant yeast genes and point mutations constructed in the Dgk1 and Pah1 conserved domains indicated that LecE functions similarly to the Nem1-Spo7 phosphatase complex that activates Pah1 in yeast. In addition, by using relevant yeast genetic backgrounds we examined several L. pneumophila effectors expected to be involved in phospholipids biosynthesis and identified an effector (LpdA) that contains a phospholipase-D (PLD) domain which caused lethal effect only in a dgk1 deletion mutant of yeast. Additionally, LpdA was found to enhance the lethal effect of LecE in yeast cells, a phenomenon which was found to be dependent on its PLD activity. Furthermore, to determine whether LecE and LpdA affect the levels or distribution of DAG and PA in-vivo in mammalian cells, we utilized fluorescent DAG and PA biosensors and validated the notion that LecE and LpdA affect the in-vivo levels and distribution of DAG and PA, respectively. Finally, we examined the intracellular localization of both LecE and LpdA in human macrophages during L. pneumophila infection and found that both effectors are localized to the bacterial phagosome. Our results suggest that L. pneumophila utilize at least two effectors to manipulate important steps in phospholipids biosynthesis. PMID:23133385

  9. Effector discovery in the fungal wheat pathogen Zymoseptoria tritici.

    PubMed

    Mirzadi Gohari, Amir; Ware, Sarah B; Wittenberg, Alexander H J; Mehrabi, Rahim; Ben M'Barek, Sarrah; Verstappen, Els C P; van der Lee, Theo A J; Robert, Olivier; Schouten, Henk J; de Wit, Pierre P J G M; Kema, Gert H J

    2015-12-01

    Fungal plant pathogens, such as Zymoseptoria tritici (formerly known as Mycosphaerella graminicola), secrete repertoires of effectors to facilitate infection or trigger host defence mechanisms. The discovery and functional characterization of effectors provides valuable knowledge that can contribute to the design of new and effective disease management strategies. Here, we combined bioinformatics approaches with expression profiling during pathogenesis to identify candidate effectors of Z.?tritici. In addition, a genetic approach was conducted to map quantitative trait loci (QTLs) carrying putative effectors, enabling the validation of both complementary strategies for effector discovery. In?planta expression profiling revealed that candidate effectors were up-regulated in successive waves corresponding to consecutive stages of pathogenesis, contrary to candidates identified by QTL mapping that were, overall, expressed at low levels. Functional analyses of two top candidate effectors (SSP15 and SSP18) showed their dispensability for Z.?tritici pathogenesis. These analyses reveal that generally adopted criteria, such as protein size, cysteine residues and expression during pathogenesis, may preclude an unbiased effector discovery. Indeed, genetic mapping of genomic regions involved in specificity render alternative effector candidates that do not match the aforementioned criteria, but should nevertheless be considered as promising new leads for effectors that are crucial for the Z.?tritici-wheat pathosystem. PMID:25727413

  10. Repeat-containing protein effectors of plant-associated organisms

    PubMed Central

    Mesarich, Carl H.; Bowen, Joanna K.; Hamiaux, Cyril; Templeton, Matthew D.

    2015-01-01

    Many plant-associated organisms, including microbes, nematodes, and insects, deliver effector proteins into the apoplast, vascular tissue, or cell cytoplasm of their prospective hosts. These effectors function to promote colonization, typically by altering host physiology or by modulating host immune responses. The same effectors however, can also trigger host immunity in the presence of cognate host immune receptor proteins, and thus prevent colonization. To circumvent effector-triggered immunity, or to further enhance host colonization, plant-associated organisms often rely on adaptive effector evolution. In recent years, it has become increasingly apparent that several effectors of plant-associated organisms are repeat-containing proteins (RCPs) that carry tandem or non-tandem arrays of an amino acid sequence or structural motif. In this review, we highlight the diverse roles that these repeat domains play in RCP effector function. We also draw attention to the potential role of these repeat domains in adaptive evolution with regards to RCP effector function and the evasion of effector-triggered immunity. The aim of this review is to increase the profile of RCP effectors from plant-associated organisms. PMID:26557126

  11. Two Cytoplasmic Effectors of Phytophthora sojae Regulate Plant Cell Death via Interactions with Plant Catalases1

    PubMed Central

    Zhang, Meixiang; Li, Qi; Liu, Tingli; Liu, Li; Shen, Danyu; Zhu, Ye; Liu, Peihan; Zhou, Jian-Min; Dou, Daolong

    2015-01-01

    Plant pathogenic oomycetes, such as Phytophthora sojae, secrete an arsenal of host cytoplasmic effectors to promote infection. We have shown previously that P. sojae PsCRN63 (for crinkling- and necrosis-inducing proteins) induces programmed cell death (PCD) while PsCRN115 blocks PCD in planta; however, they are jointly required for full pathogenesis. Here, we find that PsCRN63 alone or PsCRN63 and PsCRN115 together might suppress the immune responses of Nicotiana benthamiana and demonstrate that these two cytoplasmic effectors interact with catalases from N. benthamiana and soybean (Glycine max). Transient expression of PsCRN63 increases hydrogen peroxide (H2O2) accumulation, whereas PsCRN115 suppresses this process. Transient overexpression of NbCAT1 (for N. benthamiana CATALASE1) or GmCAT1 specifically alleviates PsCRN63-induced PCD. Suppression of the PsCRN63-induced PCD by PsCRN115 is compromised when catalases are silenced in N. benthamiana. Interestingly, the NbCAT1 is recruited into the plant nucleus in the presence of PsCRN63 or PsCRN115; NbCAT1 and GmCAT1 are destabilized when PsCRN63 is coexpressed, and PsCRN115 inhibits the processes. Thus, PsCRN63/115 manipulates plant PCD through interfering with catalases and perturbing H2O2 homeostasis. Furthermore, silencing of catalase genes enhances susceptibility to Phytophthora capsici, indicating that catalases are essential for plant resistance. Taken together, we suggest that P. sojae secretes these two effectors to regulate plant PCD and H2O2 homeostasis through direct interaction with catalases and, therefore, overcome host immune responses. PMID:25424308

  12. Design and fabrication of an end effector

    NASA Technical Reports Server (NTRS)

    Crossley, F. R. E.; Umholtz, F. G.

    1975-01-01

    The construction is described of a prototype mechanical hand or 'end effector' for use on a remotely controlled robot, but with possible application as a prosthetic device. An analysis of hand motions is reported, from which it is concluded that the two most important manipulations (apart from grasps) are to be able to pick up a tool and draw it into a nested grip against the palm, and to be able to hold a pistol-grip tool such as an electric drill and pull the trigger. A model was tested and found capable of both these operations.

  13. 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.; Villen, Judit; Scott, John D.; Stadnyk, Andrew W.; Le Trong, Isolde; Stenkamp, Ronald E.; Klevit, Rachel E.; Rohde, John R.; Brzovic, Peter S.

    2014-03-03

    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.

  14. Phytophthora effector targets a novel component of small RNA pathway in plants to promote infection

    PubMed Central

    Qiao, Yongli; Shi, Jinxia; Zhai, Yi; Hou, Yingnan; Ma, Wenbo

    2015-01-01

    A broad range of parasites rely on the functions of effector proteins to subvert host immune response and facilitate disease development. The notorious Phytophthora pathogens evolved effectors with RNA silencing suppression activity to promote infection in plant hosts. Here we report that the Phytophthora Suppressor of RNA Silencing 1 (PSR1) can bind to an evolutionarily conserved nuclear protein containing the aspartate–glutamate–alanine–histidine-box RNA helicase domain in plants. This protein, designated PSR1-Interacting Protein 1 (PINP1), regulates the accumulation of both microRNAs and endogenous small interfering RNAs in Arabidopsis. A null mutation of PINP1 causes embryonic lethality, and silencing of PINP1 leads to developmental defects and hypersusceptibility to Phytophthora infection. These phenotypes are reminiscent of transgenic plants expressing PSR1, supporting PINP1 as a direct virulence target of PSR1. We further demonstrate that the localization of the Dicer-like 1 protein complex is impaired in the nucleus of PINP1-silenced or PSR1-expressing cells, indicating that PINP1 may facilitate small RNA processing by affecting the assembly of dicing complexes. A similar function of PINP1 homologous genes in development and immunity was also observed in Nicotiana benthamiana. These findings highlight PINP1 as a previously unidentified component of RNA silencing that regulates distinct classes of small RNAs in plants. Importantly, Phytophthora has evolved effectors to target PINP1 in order to promote infection. PMID:25902521

  15. An Oomycete CRN Effector Reprograms Expression of Plant HSP Genes by Targeting their Promoters.

    PubMed

    Song, Tianqiao; Ma, Zhenchuan; Shen, Danyu; Li, Qi; Li, Wanlin; Su, Liming; Ye, Tingyue; Zhang, Meixiang; Wang, Yuanchao; Dou, Daolong

    2015-12-01

    Oomycete pathogens produce a large number of CRN effectors to manipulate plant immune responses and promote infection. However, their functional mechanisms are largely unknown. Here, we identified a Phytophthora sojae CRN effector PsCRN108 which contains a putative DNA-binding helix-hairpin-helix (HhH) motif and acts in the plant cell nucleus. Silencing of the PsCRN108 gene reduced P. sojae virulence to soybean, while expression of the gene in Nicotiana benthamiana and Arabidopsis thaliana enhanced plant susceptibility to P. capsici. Moreover, PsCRN108 could inhibit expression of HSP genes in A. thaliana, N. benthamiana and soybean. Both the HhH motif and nuclear localization signal of this effector were required for its contribution to virulence and its suppression of HSP gene expression. Furthermore, we found that PsCRN108 targeted HSP promoters in an HSE- and HhH motif-dependent manner. PsCRN108 could inhibit the association of the HSE with the plant heat shock transcription factor AtHsfA1a, which initializes HSP gene expression in response to stress. Therefore, our data support a role for PsCRN108 as a nucleomodulin in down-regulating the expression of plant defense-related genes by directly targeting specific plant promoters. PMID:26714171

  16. ATP-dependent effector-like functions of RIG-I-like receptors.

    PubMed

    Yao, Hui; Dittmann, Meike; Peisley, Alys; Hoffmann, Hans-Heinrich; Gilmore, Rachel H; Schmidt, Tobias; Schmid-Burgk, Jonathan L; Hornung, Veit; Rice, Charles M; Hur, Sun

    2015-05-01

    The vertebrate antiviral innate immune system is often considered to consist of two distinct groups of proteins: pattern recognition receptors (PRRs) that detect viral infection and induce the interferon (IFN) signaling, and effectors that directly act against viral replication. Accordingly, previous studies on PRRs, such as RIG-I and MDA5, have primarily focused on their functions in viral double-stranded RNA (dsRNA) detection and consequent antiviral signaling. We report here that both RIG-I and MDA5 efficiently displace viral proteins pre-bound to dsRNA in a manner dependent on their ATP hydrolysis, and that this activity assists a dsRNA-dependent antiviral effector protein, PKR, and allows RIG-I to promote MDA5 signaling. Furthermore, truncated RIG-I/MDA5 lacking the signaling domain, and hence the IFN stimulatory activity, displaces viral proteins and suppresses replication of certain viruses in an ATP-dependent manner. Thus, this study reveals novel "effector-like" functions of RIG-I and MDA5 that challenge the conventional view of PRRs. PMID:25891073

  17. The Piriformospora indica effector PIIN_08944 promotes the mutualistic Sebacinalean symbiosis

    PubMed Central

    Akum, Fidele N.; Steinbrenner, Jens; Biedenkopf, Dagmar; Imani, Jafargholi; Kogel, Karl-Heinz

    2015-01-01

    Pathogenic and mutualistic microbes actively suppress plant defense by secreting effector proteins to manipulate the host responses for their own benefit. Current knowledge about fungal effectors has been mainly derived from biotrophic and hemibiotrophic plant pathogenic fungi and oomycetes with restricted host range. We studied colonization strategies of the root endophytic basidiomycete Piriformospora indica that colonizes a wide range of plant species thereby establishing long-term mutualistic relationships. The release of P. indica’s genome helped to identify hundreds of genes coding for candidate effectors and provides an opportunity to investigate the role of those proteins in a mutualistic symbiosis. We demonstrate that the candidate effector PIIN_08944 plays a crucial role during fungal colonization of Arabidopsis thaliana roots. PIIN_08944 expression was detected during chlamydospore germination, and fungal deletion mutants (Pi?08944) showed delayed root colonization. Constitutive over-expression of PIIN_08944 in Arabidopsis rescued the delayed colonization phenotype of the deletion mutant. PIIN_08944-expressing Arabidopsis showed a reduced expression of flg22-induced marker genes of pattern-triggered immunity (PTI) and the salicylic acid (SA) defense pathway, and expression of PIIN_08944 in barley reduced the burst of reactive oxygen species (ROS) triggered by flg22 and chitin. These data suggest that PIIN_08944 contributes to root colonization by P. indica by interfering with SA-mediated basal immune responses of the host plant. Consistent with this, PIIN_08944-expressing Arabidopsis also supported the growth of the biotrophic oomycete Hyaloperonospora arabidopsidis while growth of the necrotrophic fungi Botrytis cinerea on Arabidopsis and Fusarium graminearum on barley was not affected. PMID:26579156

  18. The Xanthomonas campestris type III effector XopJ proteolytically degrades proteasome subunit RPT6.

    PubMed

    Üstün, Suayib; Börnke, Frederik

    2015-05-01

    Many animal and plant pathogenic bacteria inject type III effector (T3E) proteins into their eukaryotic host cells to suppress immunity. The Yersinia outer protein J (YopJ) family of T3Es is a widely distributed family of effector proteins found in both animal and plant pathogens, and its members are highly diversified in virulence functions. Some members have been shown to possess acetyltransferase activity; however, whether this is a general feature of YopJ family T3Es is currently unknown. The T3E Xanthomonas outer protein J (XopJ), a YopJ family effector from the plant pathogen Xanthomonas campestris pv vesicatoria, interacts with the proteasomal subunit Regulatory Particle AAA-ATPase6 (RPT6) in planta to suppress proteasome activity, resulting in the inhibition of salicylic acid-related immune responses. Here, we show that XopJ has protease activity to specifically degrade RPT6, leading to reduced proteasome activity in the cytoplasm as well as in the nucleus. Proteolytic degradation of RPT6 was dependent on the localization of XopJ to the plasma membrane as well as on its catalytic triad. Mutation of the Walker B motif of RPT6 prevented XopJ-mediated degradation of the protein but not XopJ interaction. This indicates that the interaction of RPT6 with XopJ is dependent on the ATP-binding activity of RPT6, but proteolytic cleavage additionally requires its ATPase activity. Inhibition of the proteasome impairs the proteasomal turnover of Nonexpressor of Pathogenesis-Related1 (NPR1), the master regulator of salicylic acid responses, leading to the accumulation of ubiquitinated NPR1, which likely interferes with the full induction of NPR1 target genes. Our results show that YopJ family T3Es are not only highly diversified in virulence function but also appear to possess different biochemical activities. PMID:25739698

  19. The Xanthomonas campestris Type III Effector XopJ Proteolytically Degrades Proteasome Subunit RPT61[OPEN

    PubMed Central

    2015-01-01

    Many animal and plant pathogenic bacteria inject type III effector (T3E) proteins into their eukaryotic host cells to suppress immunity. The Yersinia outer protein J (YopJ) family of T3Es is a widely distributed family of effector proteins found in both animal and plant pathogens, and its members are highly diversified in virulence functions. Some members have been shown to possess acetyltransferase activity; however, whether this is a general feature of YopJ family T3Es is currently unknown. The T3E Xanthomonas outer protein J (XopJ), a YopJ family effector from the plant pathogen Xanthomonas campestris pv vesicatoria, interacts with the proteasomal subunit Regulatory Particle AAA-ATPase6 (RPT6) in planta to suppress proteasome activity, resulting in the inhibition of salicylic acid-related immune responses. Here, we show that XopJ has protease activity to specifically degrade RPT6, leading to reduced proteasome activity in the cytoplasm as well as in the nucleus. Proteolytic degradation of RPT6 was dependent on the localization of XopJ to the plasma membrane as well as on its catalytic triad. Mutation of the Walker B motif of RPT6 prevented XopJ-mediated degradation of the protein but not XopJ interaction. This indicates that the interaction of RPT6 with XopJ is dependent on the ATP-binding activity of RPT6, but proteolytic cleavage additionally requires its ATPase activity. Inhibition of the proteasome impairs the proteasomal turnover of Nonexpressor of Pathogenesis-Related1 (NPR1), the master regulator of salicylic acid responses, leading to the accumulation of ubiquitinated NPR1, which likely interferes with the full induction of NPR1 target genes. Our results show that YopJ family T3Es are not only highly diversified in virulence function but also appear to possess different biochemical activities. PMID:25739698

  20. In Vitro Analyses of T Cell Effector Differentiation.

    PubMed

    Wohlfert, Elizabeth A; Carpenter, Andrea C; Belkaid, Yasmine; Bosselut, Rémy

    2016-01-01

    In vitro culture is an important complement, or substitute, to in vivo approaches in order to study T cell effector differentiation. Here, we describe culture conditions that generate specific effector cell types by exposing naïve T cells to appropriate cytokine signals. PMID:26294403

  1. Disruption of Signaling by Yersinia Effector YopJ, a

    E-print Network

    pestis is the bacterial pathogen that caused the Black Death in the Middle Ages (1). Yersinia speciesDisruption of Signaling by Yersinia Effector YopJ, a Ubiquitin-Like Protein Protease Kim Orth,1 Brian Staskawicz,2 Jack E. Dixon1 * Homologs of the Yersinia virulence effector YopJ are found in both

  2. Amphiregulin Is a Critical Downstream Effector of Estrogen Signaling in ER?-Positive Breast Cancer.

    PubMed

    Peterson, Esther A; Jenkins, Edmund C; Lofgren, Kristopher A; Chandiramani, Natasha; Liu, Hui; Aranda, Evelyn; Barnett, Maryia; Kenny, Paraic A

    2015-11-15

    Estrogen stimulation promotes epithelial cell proliferation in estrogen receptor (ER?)-positive breast cancer. Many ER? target genes have been enumerated, but the identities of the key effectors mediating the estrogen signal remain obscure. During mouse mammary gland development, the estrogen growth factor receptor (EGFR) ligand amphiregulin acts as an important stage-specific effector of estrogen signaling. In this study, we investigated the role of amphiregulin in breast cancer cell proliferation using human tissue samples and tumor xenografts in mice. Amphiregulin was enriched in ER?-positive human breast tumor cells and required for estrogen-dependent growth of MCF7 tumor xenografts. Furthermore, amphiregulin levels were suppressed in patients treated with endocrine therapy. Suppression of EGF receptor signaling appeared necessary for the therapeutic response in this setting. Our findings implicate amphiregulin as a critical mediator of the estrogen response in ER?-positive breast cancer, emphasizing the importance of EGF receptor signaling in breast tumor pathogenesis and therapeutic response. Cancer Res; 75(22); 4830-8. ©2015 AACR. PMID:26527289

  3. FairyTALE: A High-Throughput TAL Effector Synthesis Platform Jing Liang,

    E-print Network

    Zhao, Huimin

    FairyTALE: A High-Throughput TAL Effector Synthesis Platform Jing Liang, Ran Chao,, Zhanar Abil, genome engineering, genome editing, TAL effector, TALEN Recombinant transcription activator-like effector

  4. Miniature Trailing Edge Effector for Aerodynamic Control

    NASA Technical Reports Server (NTRS)

    Lee, Hak-Tae (Inventor); Bieniawski, Stefan R. (Inventor); Kroo, Ilan M. (Inventor)

    2008-01-01

    Improved miniature trailing edge effectors for aerodynamic control are provided. Three types of devices having aerodynamic housings integrated to the trailing edge of an aerodynamic shape are presented, which vary in details of how the control surface can move. A bucket type device has a control surface which is the back part of a C-shaped member having two arms connected by the back section. The C-shaped section is attached to a housing at the ends of the arms, and is rotatable about an axis parallel to the wing trailing edge to provide up, down and neutral states. A flip-up type device has a control surface which rotates about an axis parallel to the wing trailing edge to provide up, down, neutral and brake states. A rotating type device has a control surface which rotates about an axis parallel to the chord line to provide up, down and neutral states.

  5. Cerato-platanins: elicitors and effectors.

    PubMed

    Pazzagli, Luigia; Seidl-Seiboth, Verena; Barsottini, Mario; Vargas, Walter A; Scala, Aniello; Mukherjee, Prasun K

    2014-11-01

    Cerato-platanins are an interesting group of small, secreted, cysteine-rich proteins that have been implicated in virulence of certain plant pathogenic fungi. The relatively recent discovery of these proteins in plant beneficial fungi like Trichoderma spp., and their positive role in induction of defense in plants against invading pathogens has raised the question as to whether these proteins are effectors or elicitor molecules. Here we present a comprehensive review on the occurrence of these conserved proteins across the fungal kingdom, their structure-function relationships, and their physiological roles in plant pathogenic and symbiotic fungi. We also discuss the usefulness of these proteins in evolving strategies for crop protection through a transgenic approach or direct application as elicitors. PMID:25438788

  6. Rack Insertion End Effector (RIEE) automation

    NASA Technical Reports Server (NTRS)

    Malladi, Narasimha

    1993-01-01

    NASA is developing a mechanism to manipulate and insert Racks into the Space Station Logistic modules. The mechanism consists of the following: a base with three motorized degrees of freedom, a 3 section motorized boom that goes from 15 to 44 feet in length, and a Rack Insertion End Effector (RIEE) with 5 hand wheels for precise alignment. The robotics section was tasked with the automation of the RIEE unit. In this report, for the automation of the RIEE unit, application of the Perceptics Vision System was conceptually developed to determine the position and orientation of the RIEE relative to the logistic module, and a MathCad program is written to display the needed displacements for precise alignment and final insertion of the Rack. The uniqueness of this report is that the whole report is in fact a MathCad program including text, derivations, and executable equations with example inputs and outputs.

  7. The Xanthomonas campestris effector protein XopDXcc8004 triggers plant disease tolerance by targeting DELLA proteins.

    PubMed

    Tan, Leitao; Rong, Wei; Luo, Hongli; Chen, Yinhua; He, Chaozu

    2014-11-01

    Plants protect themselves from the harmful effects of pathogens by resistance and tolerance. Disease resistance, which eliminates pathogens, can be modulated by bacterial type III effectors. Little is known about whether disease tolerance, which sustains host fitness with a given pathogen burden, is regulated by effectors. Here, we examined the effects of the Xanthomonas effector protein XopDXcc8004 on plant disease defenses by constructing knockout and complemented Xanthomonas strains, and performing inoculation studies in radish (Raphanus sativus L. var. radiculus XiaoJinZhong) and Arabidopsis plants. XopDXcc8004 suppresses disease symptoms without changing bacterial titers in infected leaves. In Arabidopsis, XopDXcc8004 delays the hormone gibberellin (GA)-mediated degradation of RGA (repressor of ga1-3), one of five DELLA proteins that repress GA signaling and promote plant tolerance under biotic and abiotic stresses. The ERF-associated amphiphilic repression (EAR) motif-containing region of XopDXcc8004 interacts with the DELLA domain of RGA and might interfere with the GA-induced binding of GID1, a GA receptor, to RGA. The EAR motif was found to be present in a number of plant transcriptional regulators. Thus, our data suggest that bacterial pathogens might have evolved effectors, which probably mimic host components, to initiate disease tolerance and enhance their survival. PMID:25040905

  8. Overexpression of a Phytophthora Cytoplasmic CRN Effector Confers Resistance to Disease, Salinity and Drought in Nicotiana benthamiana.

    PubMed

    Rajput, Nasir Ahmed; Zhang, Meixiang; Shen, Danyu; Liu, Tingli; Zhang, Qimeng; Ru, Yanyan; Sun, Peng; Dou, Daolong

    2015-12-01

    The Crinkler (CRN) effector family is produced by oomycete pathogens and may manipulate host physiological and biochemical events inside host cells. Here, PsCRN161 was identified from Phytophthora sojae based on its broad and strong cell death suppression activities. The effector protein contains two predicted nuclear localization signals and localized to nuclei of plant cells, indicating that it may target plant nuclei to modify host cell physiology and function. The chimeric gene GFP:PsCRN161 driven by the Cauliflower mosaic virus (CaMV) 35S promoter was introduced into Nicotiana benthamiana. The four independent PsCRN161-transgenic lines exhibited increased resistance to two oomycete pathogens (P. parasitica and P. capsici) and showed enhanced tolerance to salinity and drought stresses. Digital gene expression profiling analysis showed that defense-related genes, including ABC transporters, Cyt P450 and receptor-like kinases (RLKs), were significantly up-regulated in PsCRN161-transgenic plants compared with GFP (green fluorescent protein) lines, implying that PsCRN161 expression may protect plants from biotic and abiotic stresses by up-regulation of many defense-related genes. The results reveal previously unknown functions of the oomycete effectors, suggesting that the pathogen effectors could be directly used as functional genes for plant molecular breeding for enhancement of tolerance to biotic and abiotic stresses. PMID:26546319

  9. T-Regulatory Cells and Programmed Death 1+ T Cells Contribute to Effector T-Cell Dysfunction in Patients with Chronic Obstructive Pulmonary Disease

    PubMed Central

    Kalathil, Suresh Gopi; Lugade, Amit Anand; Pradhan, Vandana; Miller, Austin; Parameswaran, Ganapathi Iyer; Sethi, Sanjay

    2014-01-01

    Rationale: Previous studies from our laboratory have shown that peripheral blood mononuclear cells (PBMCs) from patients with chronic obstructive pulmonary disease (COPD) prone to exacerbations with nontypeable Haemophilus influenzae have impaired responses to lipoprotein P6. We hypothesized that an underlying immunosuppressive network could be responsible for the defective antibacterial immunity observed in these patients. We evaluated T regulatory cells (Tregs), myeloid-derived suppressor cells (MDSC), and exhausted T effector cells (programmed death 1 [PD-1]+) in patients with COPD, because these cells are known to play a pivotal role in suppressing immune responses. Objectives: We performed an in-depth characterization of Tregs, T effector cells, and MDSC in COPD and correlated their levels and function with disease severity. Methods: Treg, effector T cell, and MDSC frequency from patients with COPD and healthy subjects’ PBMCs were analyzed by flow cytometry. Treg immunosuppressive capacity was measured by in vitro suppression assay. The frequency of interferon-? producing T cells and T-cell proliferation were measured after blocking CTLA-4 and PD-1. Plasma proinflammatory and immunosuppressive cytokine levels were measured. Measurements and Main Results: Significantly increased levels of Tregs, MDSC, and PD-1+ exhausted effector T cells were present in patients with COPD compared with healthy subjects. Tregs from patients with COPD suppressed P6-specific T-cell proliferation to a greater extent than Tregs from healthy subjects. Plasma levels of Treg-generated cytokines, IL-10, and transforming growth factor-? were elevated. Blockade of CTLA-4 resulted in significant augmentation of T-cell IFN-? production in patients with COPD. Conclusions: Functionally suppressive Tregs, MDSCs, and exhausted PD-1+ T cells contribute to effector T-cell dysfunction in COPD. PMID:24825462

  10. Experimental approaches to investigate effector translocation into host cells in the Ustilago maydis/maize pathosystem.

    PubMed

    Tanaka, Shigeyuki; Djamei, Armin; Presti, Libera Lo; Schipper, Kerstin; Winterberg, Sarah; Amati, Simone; Becker, Dirk; Büchner, Heike; Kumlehn, Jochen; Reissmann, Stefanie; Kahmann, Regine

    2015-01-01

    The fungus Ustilago maydis is a pathogen that establishes a biotrophic interaction with Zea mays. The interaction with the plant host is largely governed by more than 300 novel, secreted protein effectors, of which only four have been functionally characterized. Prerequisite to examine effector function is to know where effectors reside after secretion. Effectors can remain in the extracellular space, i.e. the plant apoplast (apoplastic effectors), or can cross the plant plasma membrane and exert their function inside the host cell (cytoplasmic effectors). The U. maydis effectors lack conserved motifs in their primary sequences that could allow a classification of the effectome into apoplastic/cytoplasmic effectors. This represents a significant obstacle in functional effector characterization. Here we describe our attempts to establish a system for effector classification into apoplastic and cytoplasmic members, using U. maydis for effector delivery. PMID:26118724

  11. Effector biology during biotrophic invasion of plant cells

    PubMed Central

    Chaudhari, Prateek; Ahmed, Bulbul; Joly, David L; Germain, Hugo

    2014-01-01

    Several obligate biotrophic phytopathogens, namely oomycetes and fungi, invade and feed on living plant cells through specialized structures known as haustoria. Deploying an arsenal of secreted proteins called effectors, these pathogens balance their parasitic propagation by subverting plant immunity without sacrificing host cells. Such secreted proteins, which are thought to be delivered by haustoria, conceivably reprogram host cells and instigate structural modifications, in addition to the modulation of various cellular processes. As effectors represent tools to assist disease resistance breeding, this short review provides a bird’s eye view on the relationship between the virulence function of effectors and their subcellular localization in host cells. PMID:25513771

  12. System for exchanging tools and end effectors on a robot

    DOEpatents

    Burry, David B. (Westminster, CO); Williams, Paul M. (Lafayette, CO)

    1991-02-19

    A system and method for exchanging tools and end effectors on a robot permits exchange during a programmed task. The exchange mechanism is located off the robot, thus reducing the mass of the robot arm and permitting smaller robots to perform designated tasks. A simple spring/collet mechanism mounted on the robot is used which permits the engagement and disengagement of the tool or end effector without the need for a rotational orientation of the tool to the end effector/collet interface. As the tool changing system is not located on the robot arm no umbilical cords are located on robot.

  13. System for exchanging tools and end effectors on a robot

    DOEpatents

    Burry, D.B.; Williams, P.M.

    1991-02-19

    A system and method for exchanging tools and end effectors on a robot permits exchange during a programmed task. The exchange mechanism is located off the robot, thus reducing the mass of the robot arm and permitting smaller robots to perform designated tasks. A simple spring/collet mechanism mounted on the robot is used which permits the engagement and disengagement of the tool or end effector without the need for a rotational orientation of the tool to the end effector/collet interface. As the tool changing system is not located on the robot arm no umbilical cords are located on robot. 12 figures.

  14. Hippo pathway effector Yap promotes cardiac regeneration

    PubMed Central

    Xin, Mei; Kim, Yuri; Sutherland, Lillian B.; Murakami, Masao; Qi, Xiaoxia; McAnally, John; Porrello, Enzo R.; Mahmoud, Ahmed I.; Tan, Wei; Shelton, John M.; Richardson, James A.; Sadek, Hesham A.; Bassel-Duby, Rhonda; Olson, Eric N.

    2013-01-01

    The adult mammalian heart has limited potential for regeneration. Thus, after injury, cardiomyocytes are permanently lost, and contractility is diminished. In contrast, the neonatal heart can regenerate owing to sustained cardiomyocyte proliferation. Identification of critical regulators of cardiomyocyte proliferation and quiescence represents an important step toward potential regenerative therapies. Yes-associated protein (Yap), a transcriptional cofactor in the Hippo signaling pathway, promotes proliferation of embryonic cardiomyocytes by activating the insulin-like growth factor and Wnt signaling pathways. Here we report that mice bearing mutant alleles of Yap and its paralog WW domain containing transcription regulator 1 (Taz) exhibit gene dosage-dependent cardiac phenotypes, suggesting redundant roles of these Hippo pathway effectors in establishing proper myocyte number and maintaining cardiac function. Cardiac-specific deletion of Yap impedes neonatal heart regeneration, resulting in a default fibrotic response. Conversely, forced expression of a constitutively active form of Yap in the adult heart stimulates cardiac regeneration and improves contractility after myocardial infarction. The regenerative activity of Yap is correlated with its activation of embryonic and proliferative gene programs in cardiomyocytes. These findings identify Yap as an important regulator of cardiac regeneration and provide an experimental entry point to enhance this process. PMID:23918388

  15. Human Urinary Exosomes as Innate Immune Effectors

    PubMed Central

    Hiemstra, Thomas F.; Charles, Philip D.; Gracia, Tannia; Hester, Svenja S.; Gatto, Laurent; Al-Lamki, Rafia; Floto, R. Andres; Su, Ya; Skepper, Jeremy N.

    2014-01-01

    Exosomes are small extracellular vesicles, approximately 50 nm in diameter, derived from the endocytic pathway and released by a variety of cell types. Recent data indicate a spectrum of exosomal functions, including RNA transfer, antigen presentation, modulation of apoptosis, and shedding of obsolete protein. Exosomes derived from all nephron segments are also present in human urine, where their function is unknown. Although one report suggested in vitro uptake of exosomes by renal cortical collecting duct cells, most studies of human urinary exosomes have focused on biomarker discovery rather than exosome function. Here, we report results from in-depth proteomic analyses and EM showing that normal human urinary exosomes are significantly enriched for innate immune proteins that include antimicrobial proteins and peptides and bacterial and viral receptors. Urinary exosomes, but not the prevalent soluble urinary protein uromodulin (Tamm–Horsfall protein), potently inhibited growth of pathogenic and commensal Escherichia coli and induced bacterial lysis. Bacterial killing depended on exosome structural integrity and occurred optimally at the acidic pH typical of urine from omnivorous humans. Thus, exosomes are innate immune effectors that contribute to host defense within the urinary tract. PMID:24700864

  16. Effector T cell differentiation: are master regulators of effector T cells still the masters?

    PubMed

    Wang, Chao; Collins, Mary; Kuchroo, Vijay K

    2015-12-01

    Effector CD4 T cell lineages have been implicated as potent inducers of autoimmune diseases. Tbet, Gata3 and Rorgt are master transcriptional regulators of Th1, Th2 and Th17 lineages respectively and promote the distinct expression of signature cytokines. Significant progress has been made in understanding the transcriptional network that drives CD4 T cell differentiation, revealing novel points of regulation mediated by transcription factors, cell surface receptors, cytokines and chemokines. Epigenetic modifications and metabolic mediators define the transcriptional landscape in which master transcription factors operate and collaborate with a network of transcriptional modifiers to guide lineage specification, plasticity and function. PMID:26319196

  17. Robotic end-effector for rewaterproofing shuttle tiles

    NASA Technical Reports Server (NTRS)

    Manouchehri, Davoud; Hansen, Joseph M.; Wu, Cheng M.; Yamamoto, Brian S.; Graham, Todd

    1992-01-01

    This paper summarizes work by Rockwell International's Space Systems Division's Robotics Group at Downey, California. The work is part of a NASA-led team effort to automate Space Shuttle rewaterproofing in the Orbiter Processing Facility at the Kennedy Space Center and the ferry facility at the Ames-Dryden Flight Research Facility. Rockwell's effort focuses on the rewaterproofing end-effector, whose function is to inject hazardous dimethylethyloxysilane into thousands of ceramic tiles on the underside of the orbiter after each flight. The paper has five sections. First, it presents background on the present manual process. Second, end-effector requirements are presented, including safety and interface control. Third, a design is presented for the five end-effector systems: positioning, delivery, containment, data management, and command and control. Fourth, end-effector testing and integrating to the total system are described. Lastly, future applications for this technology are discussed.

  18. An intelligent end-effector for a rehabilitation robot.

    PubMed

    Gosine, R G; Harwin, W S; Furby, L J; Jackson, R D

    1989-01-01

    A UMI RTX robot, modified with limited end-effector sensors and a restricted but effective vision system, is currently used in a developmental education setting for severely physically disabled children. The low physical and cognitive abilities of the children involved in the project require a semi-autonomous robot with environmental sensing capability to operate in a task oriented mode. A variety of low-cost sensors including proximity, distance, force and slip sensors, have been investigated for integration in end-effectors for the RTX robot. The sensors employed on a modified end-effector are detailed and experimental results are presented. A design for an end-effector with integrated sensors is discussed. The integration of the sensor information into a high-level, task-oriented programming language is detailed and examples of high-level control sequences using sensor inputs are presented. Finally, the development of intelligent gripping strategies based on sensor information is discussed. PMID:2733012

  19. Plant cells under siege: plant immune system versus pathogen effectors.

    PubMed

    Asai, Shuta; Shirasu, Ken

    2015-12-01

    Pathogen-secreted effector proteins enable pathogens to manipulate plant immunity for successful infection. To penetrate host apoplastic space, pathogens reopen the stomata. Once the invasion into the apoplast occurs, pathogens deceive the host detection system by deploying apoplastic effectors. Pathogens also deliver an arsenal of cytosolic effectors into the host cells, which undermine host immunity such as salicylic acid (SA)-dependent immunity. Here we summarize recent findings that highlight the functions of the effectors from fungal, oomycete and bacterial pathogens in the key steps of infection at the stomata, in the apoplast, and inside the cell. We also discuss cell type-specific responses in the host during infection and the necessity of further investigation of plant-pathogen interactions at spatial and temporal resolution. PMID:26343014

  20. Memory versus effector immune responses in oncolytic virotherapies.

    PubMed

    Macnamara, Cicely; Eftimie, Raluca

    2015-07-21

    The main priority when designing cancer immuno-therapies has been to seek viable biological mechanisms that lead to permanent cancer eradication or cancer control. Understanding the delicate balance between the role of effector and memory cells on eliminating cancer cells remains an elusive problem in immunology. Here we make an initial investigation into this problem with the help of a mathematical model for oncolytic virotherapy; although the model can in fact be made general enough to be applied also to other immunological problems. According to this model, we find that long-term cancer control is associated with a large number of persistent effector cells (irrespective of the initial peak in effector cell numbers). However, this large number of persistent effector cells is sustained by a relatively large number of memory cells. Moreover, the results of the mathematical model suggest that cancer control from a dormant state cannot be predicted by the size of the memory population. PMID:25882747

  1. Substrate recognition by the zinc metalloprotease effector NleC from enteropathogenic Escherichia coli.

    PubMed

    Giogha, Cristina; Wong Fok Lung, Tania; Mühlen, Sabrina; Pearson, Jaclyn S; Hartland, Elizabeth L

    2015-12-01

    Upon infection of epithelial cells, enteropathogenic Escherichia?coli suppresses host cell inflammatory signalling in a type III secretion system (T3SS) dependent manner. Two key T3SS effector proteins involved in this response are NleE and NleC. NleC is a zinc metalloprotease effector that degrades the p65 subunit of NF-?B. Although the site of p65 cleavage by NleC is now well described, other areas of interaction have not been precisely defined. Here we constructed overlapping truncations of p65 to identify regions required for NleC cleavage. We determined that NleC cleaved both p65 and p50 within the Rel homology domain (RHD) and that two motifs, E22 IIE25 and P177 VLS180 , within the RHD of p65 were important for recognition and binding by NleC. Alanine substitution of one or both of these motifs protected p65 from binding and degradation by NleC. The E22 IIE25 and P177 VLS180 motifs were located within the structurally distinct N-terminal subdomain of the RHD involved in DNA binding by p65 on adjacent, parallel strands. Although these motifs have not been recognized previously, both were needed for the correct localization and function of p65. In summary, this work has identified two regions of p65 within the RHD needed for binding and cleavage by NleC and provides further insight into the molecular basis of substrate recognition by a T3SS effector. PMID:26096513

  2. Ganglioside-exposed dendritic cells inhibit T-cell effector function by promoting regulatory cell activity.

    PubMed

    Jales, Alessandra; Falahati, Rustom; Mari, Elisabeth; Stemmy, Erik J; Shen, Weiping; Southammakosane, Cathy; Herzog, Dallen; Ladisch, Stephan; Leitenberg, David

    2011-01-01

    Tumour pathogenesis is characterized by an immunosuppressive microenvironment that limits the development of effective tumour-specific immune responses. This is in part the result of tumour-dependent recruitment and activation of regulatory cells, such as myeloid-derived suppressor cells and regulatory T cells in the tumour microenvironment and draining lymph nodes. Shedding of gangliosides by tumour cells has immunomodulatory properties, suggesting that gangliosides may be a critical factor in initiating an immunosuppressive microenvironment. To better define the immunomodulatory properties of gangliosides on antigen-specific T-cell activation and development we have developed an in vitro system using ganglioside-treated murine bone-marrow-derived dendritic cells to prime and activate antigen-specific CD4(+) T cells from AND T-cell receptor transgenic mice. Using this system, ganglioside treatment promotes the development of a dendritic cell population characterized by decreased CD86 (B7-2) expression, and decreased interleukin-12 and interleukin-6 production. When these cells are used as antigen-presenting cells, CD4 T cells are primed to proliferate normally, but have a defect in T helper (Th) effector cell development. This defect in Th effector cell responses is associated with the development of regulatory T-cell activity that can suppress the activation of previously primed Th effector cells in a contact-dependent manner. In total, these data suggest that ganglioside-exposed dendritic cells promote regulatory T-cell activity that may have long-lasting effects on the development of tumour-specific immune responses. PMID:20875076

  3. Phytophthora infestans RXLR Effector AVR1 Interacts with Exocyst Component Sec5 to Manipulate Plant Immunity1[OPEN

    PubMed Central

    Du, Yu; Mpina, Mohamed H.; Birch, Paul R.J.; Bouwmeester, Klaas; Govers, Francine

    2015-01-01

    Phytophthora infestans secretes numerous RXLR effectors that modulate host defense and thereby pave the way for successful invasion. Here, we show that the RXLR effector AVR1 is a virulence factor that promotes colonization and suppresses callose deposition, a hallmark of basal defense. To identify host targets of AVR1, we performed yeast two-hybrid screens and selected Sec5 as a candidate. Sec5 is a subunit of the exocyst, a protein complex that is involved in vesicle trafficking. AVR1-like (A-L), a close homolog of AVR1, also acts as a virulence factor, but unlike AVR1, A-L does not suppress CRINKLER2 (CRN2)-induced cell death or interact with Sec5. Compared with AVR1, A-L is shorter and lacks the carboxyl-terminal tail, the T-region that is crucial for CRN2-induced cell death suppression and Sec5 interaction. In planta analyses revealed that AVR1 and Sec5 are in close proximity, and coimmunoprecipitation confirmed the interaction. Sec5 is required for secretion of the pathogenesis-related protein PR-1 and callose deposition and also plays a role in CRN2-induced cell death. Our findings show that P. infestans manipulates an exocyst subunit and thereby potentially disturbs vesicle trafficking, a cellular process that is important for basal defense. This is a novel strategy that oomycete pathogens exploit to modulate host defense. PMID:26336092

  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-?, 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 ?B?, 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

  5. A Translocated Effector Required for Bartonella Dissemination from Derma to Blood Safeguards Migratory Host Cells from Damage by Co-translocated Effectors

    E-print Network

    Okujava, Rusudan

    Numerous bacterial pathogens secrete multiple effectors to modulate host cellular functions. These effectors may interfere with each other to efficiently control the infection process. Bartonellae are Gram-negative, ...

  6. Suppression of Fc?-Receptor-Mediated Antibody Effector Function during Persistent Viral Infection

    E-print Network

    Yamada, Douglas

    2015-01-01

    The prevalence of IgE antinuclear antibodies in rheumatoidof antibody Fc components (IgM, IgD, IgG, IgE, and IgA) thatantibody-dependent cell-mediated phagocytosis (ADCP) (97, 98). The Fc portion of IgE

  7. A Novel Ras Effector Pathway Found to Play Significant Role in Tumor Suppression | Poster

    Cancer.gov

    By Nancy Parrish, Staff Writer; photo by Richard Frederickson, Staff Photographer Normal cells have mechanisms to prevent the development of cancer. Among these is a type of tumor suppressor mechanism known as oncogene-induced senescence, or OIS, which halts the uncontrolled growth of cells caused by mutations in oncogenes. The oncogene Ras plays a crucial role in inducing OIS through a specific cascade of proteins, as reported in a recent article in Molecular and Cellular Biology by Jacqueline Salotti, Ph.D., and colleagues in the Eukaryotic Transcriptional Regulation Section of the Mouse Cancer Genetics Program, Center for Cancer Research (CCR).

  8. The Pseudomonas syringae effector HopF2 suppresses Arabidopsis immunity by targeting BAK1

    E-print Network

    Sheen, Jen

    and the plasma membrane-localized receptor-like cytoplasmic kinase BIK1 and its homologs. We further show that HopF2 directly targets BAK1, a plasma membrane-localized receptor-like kinase that is involved-localized receptor-like kinases (RLKs) or receptor-like proteins (Jones and Dangl, 2006; Boller and Felix, 2009

  9. The limited capacity of malignant glioma-derived exosomes to suppress peripheral immune effectors.

    PubMed

    Iorgulescu, J Bryan; Ivan, Michael E; Safaee, Michael; Parsa, Andrew T

    2016-01-15

    Tumor-derived microvesicular exosomes permit intercellular communication both locally and systemically by delivering a snapshot of the tumor cell's constituents. We thus investigated whether exosomes mediate malignant glioma's facility for inducing peripheral immunosuppression. In Western blot and RT-PCR analyses, glioma-derived exosomes displayed exosome-specific markers, but failed to recapitulate the antigen-presentation machinery, surface co-modulatory signals, or immunosuppressive mediator status of their parent tumor cells. Treatment with glioma-derived exosomes promoted immunosuppressive HLA-DR(low) monocytic phenotypes, but failed to induce monocytic PD-L1 expression or alter the activation of cytotoxic T-cells from patients' peripheral blood by FACS and RT-PCR analyses. Our results suggest that malignant glioma-derived exosomes are restricted in their capacity to directly prime peripheral immunosuppression. PMID:26711578

  10. IL-17 suppresses immune effector functions in HPV-associated epithelial hyperplasia1,2

    PubMed Central

    Gosmann, Christina; Mattarollo, Stephen R.; Bridge, Jennifer A.; Frazer, Ian H.; Blumenthal, Antje

    2014-01-01

    Persistent infection with high-risk human papillomaviruses (HPV) causes epithelial hyperplasia that can progress to cancer, and is thought to depend on immunosuppressive mechanisms that prevent viral clearance by the host. IL-17 is a cytokine with diverse functions in host defense and in the pathology of autoimmune disorders, chronic inflammatory diseases and cancer. We analyzed biopsies from patients with HPV- associated cervical intraepithelial neoplasia (CIN) grade 2/3 and murine skin displaying HPV16 E7 protein-induced epithelial hyperplasia, which closely models hyperplasia in chronic HPV lesions. Expression of IL-17 and IL-23, a major inducer of IL-17, was elevated in both human HPV-infected and murine E7-expressing lesions. Using a skin grafting model, we demonstrated that IL-17 in HPV16 E7 transgenic skin grafts inhibited effective host immune responses against the graft. IL-17 was produced by CD3+ T cells, predominantly CD4+ T cells in human, and CD4+ and ?? T cells in mouse hyperplastic lesions. IL-23 and IL-1?, but not IL-18, induced IL-17 production in E7 transgenic skin. Together, these findings demonstrate an immunosuppressive role for IL-17 in HPV-associated epithelial hyperplasia and suggest that blocking IL-17 in persistent viral infection may promote antiviral immunity and prevent progression to cancer. PMID:25063870

  11. Planar cell polarity effector gene Intu regulates cell fate-specific differentiation of keratinocytes through the primary cilia.

    PubMed

    Dai, D; Li, L; Huebner, A; Zeng, H; Guevara, E; Claypool, D J; Liu, A; Chen, J

    2013-01-01

    Genes involved in the planar cell polarity (PCP) signaling pathway are essential for a number of developmental processes in mammals, such as convergent extension and ciliogenesis. Tissue-specific PCP effector genes of the PCP signaling pathway are believed to mediate PCP signals in a tissue- and cell type-specific manner. However, how PCP signaling controls the morphogenesis of mammalian tissues remains unclear. In this study, we investigated the role of inturned (Intu), a tissue-specific PCP effector gene, during hair follicle formation in mice. Tissue-specific disruption of Intu in embryonic epidermis resulted in hair follicle morphogenesis arrest because of the failure of follicular keratinocyte to differentiate. Targeting Intu in the epidermis resulted in almost complete loss of primary cilia in epidermal and follicular keratinocytes, and a suppressed hedgehog signaling pathway. Surprisingly, the epidermal stratification and differentiation programs and barrier function were not affected. These results demonstrate that tissue-specific PCP effector genes of the PCP signaling pathway control the differentiation of keratinocytes through the primary cilia in a cell fate- and context-dependent manner, which may be critical in orchestrating the propagation and interpretation of polarity signals established by the core PCP components. PMID:22935613

  12. The Xanthomonas effector XopJ triggers a conditional hypersensitive response upon treatment of N. benthamiana leaves with salicylic acid

    PubMed Central

    Üstün, Suayib; Bartetzko, Verena; Börnke, Frederik

    2015-01-01

    XopJ is a Xanthomonas type III effector protein that promotes bacterial virulence on susceptible pepper plants through the inhibition of the host cell proteasome and a resultant suppression of salicylic acid (SA) – dependent defense responses. We show here that Nicotiana benthamiana leaves transiently expressing XopJ display hypersensitive response (HR) –like symptoms when exogenously treated with SA. This apparent avirulence function of XopJ was further dependent on effector myristoylation as well as on an intact catalytic triad, suggesting a requirement of its enzymatic activity for HR-like symptom elicitation. The ability of XopJ to cause a HR-like symptom development upon SA treatment was lost upon silencing of SGT1 and NDR1, respectively, but was independent of EDS1 silencing, suggesting that XopJ is recognized by an R protein of the CC-NBS-LRR class. Furthermore, silencing of NPR1 abolished the elicitation of HR-like symptoms in XopJ expressing leaves after SA application. Measurement of the proteasome activity indicated that proteasome inhibition by XopJ was alleviated in the presence of SA, an effect that was not observed in NPR1 silenced plants. Our results suggest that XopJ – triggered HR-like symptoms are closely related to the virulence function of the effector and that XopJ follows a two-signal model in order to elicit a response in the non-host plant N. benthamiana. PMID:26284106

  13. African Green Monkey TRIM5? Restriction in Simian Immunodeficiency Virus-Specific Rhesus Macaque Effector CD4 T Cells Enhances Their Survival and Antiviral Function

    PubMed Central

    Jain, Sumiti; Trivett, Matthew T.; Ayala, Victor I.; Ohlen, Claes

    2015-01-01

    ABSTRACT The expression of xenogeneic TRIM5? proteins can restrict infection in various retrovirus/host cell pairings. Previously, we have shown that African green monkey TRIM5? (AgmTRIM5?) potently restricts both human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus mac239 (SIVmac239) replication in a transformed human T-cell line (L. V. Coren, et al., Retrovirology 12:11, 2015, http://dx.doi.org/10.1186/s12977-015-0137-9). To assess AgmTRIM5? restriction in primary cells, we transduced AgmTRIM5? into primary rhesus macaque CD4 T cells and infected them with SIVmac239. Experiments with T-cell clones revealed that AgmTRIM5? could reproducibly restrict SIVmac239 replication, and that this restriction synergizes with an intrinsic resistance to infection present in some CD4 T-cell clones. AgmTRIM5? transduction of virus-specific CD4 T-cell clones increased and prolonged their ability to suppress SIV spread in CD4 target cells. This increased antiviral function was strongly linked to decreased viral replication in the AgmTRIM5?-expressing effectors, consistent with restriction preventing the virus-induced cytopathogenicity that disables effector function. Taken together, our data show that AgmTRIM5? restriction, although not absolute, reduces SIV replication in primary rhesus CD4 T cells which, in turn, increases their antiviral function. These results support prior in vivo data indicating that the contribution of virus-specific CD4 T-cell effectors to viral control is limited due to infection. IMPORTANCE The potential of effector CD4 T cells to immunologically modulate SIV/HIV infection likely is limited by their susceptibility to infection and subsequent inactivation or elimination. Here, we show that AgmTRIM5? expression inhibits SIV spread in primary effector CD4 T cells in vitro. Importantly, protection of effector CD4 T cells by AgmTRIM5? markedly enhanced their antiviral function by delaying SIV infection, thereby extending their viability despite the presence of virus. Our in vitro data support prior in vivo HIV-1 studies suggesting that the antiviral CD4 effector response is impaired due to infection and subsequent cytopathogenicity. The ability of AgmTRIM5? expression to restrict SIV infection in primary rhesus effector CD4 T cells now opens an opportunity to use the SIV/rhesus macaque model to further elucidate the potential and scope of anti-AIDS virus effector CD4 T-cell function. PMID:25653448

  14. Salmonella Effectors: Important players modulating host cell function during infection

    PubMed Central

    Agbor, Terence A.; McCormick, Beth A.

    2012-01-01

    Summary Salmonella enterica serovar Typhimurium (S. Typhimurium) is a Gram-negative facultative foodborne pathogen that causes gastroenteritis in humans. This bacterium has evolved a sophisticated machinery to alter host cell function critical to its virulence capabilities. Central to S. Typhimurium pathogenesis are two Type three secretion systems (T3SS) encoded within pathogenicity islands SPI-1 and SPI-2 that are responsible for the secretion and translocation of a set of bacterial proteins termed effectors into host cells with the intention of altering host cell physiology for bacterial entry and survival. Thus, once delivered by the T3SS, the secreted effectors play critical roles in manipulating the host cell to allow for bacteria invasion, induction of inflammatory responses, and the assembly of an intracellular protective niche created for bacterial survival and replication. Emerging evidence indicates that these effectors are modular proteins consisting of distinct functional domains/motifs that are utilized by the bacteria to activate intracellular signaling pathways modifying host cell function. Also, recently reported are the dual functionality of secreted effectors and the concept of “terminal reassortment”. Herein, we highlight some of the nascent concepts regarding Salmonella effectors in the context infection. PMID:21902796

  15. Structural Analysis of Iac Repressor Bound to Allosteric Effectors

    SciTech Connect

    Daber,R.; Stayrook, S.; Rosenberg, A.; Lewis, M.

    2007-01-01

    The lac operon is a model system for understanding how effector molecules regulate transcription and are necessary for allosteric transitions. The crystal structures of the lac repressor bound to inducer and anti-inducer molecules provide a model for how these small molecules can modulate repressor function. The structures of the apo repressor and the repressor bound to effector molecules are compared in atomic detail. All effectors examined here bind to the repressor in the same location and are anchored to the repressor through hydrogen bonds to several hydroxyl groups of the sugar ring. Inducer molecules form a more extensive hydrogen-bonding network compared to anti-inducers and neutral effector molecules. The structures of these effector molecules suggest that the O6 hydroxyl on the galactoside is essential for establishing a water-mediated hydrogen bonding network that bridges the N-terminal and C-terminal sub-domains. The altered hydrogen bonding can account in part for the different structural conformations of the repressor, and is vital for the allosteric transition.

  16. Ralstonia solanacearum Requires PopS, an Ancient AvrE-Family Effector, for Virulence and To Overcome Salicylic Acid-Mediated Defenses during Tomato Pathogenesis

    PubMed Central

    Jacobs, Jonathan M.; Milling, Annett; Mitra, Raka M.; Hogan, Clifford S.; Ailloud, Florent; Prior, Philippe; Allen, Caitilyn

    2013-01-01

    ABSTRACT During bacterial wilt of tomato, the plant pathogen Ralstonia solanacearum upregulates expression of popS, which encodes a type III-secreted effector in the AvrE family. PopS is a core effector present in all sequenced strains in the R. solanacearum species complex. The phylogeny of popS mirrors that of the species complex as a whole, suggesting that this is an ancient, vertically inherited effector needed for association with plants. A popS mutant of R. solanacearum UW551 had reduced virulence on agriculturally important Solanum spp., including potato and tomato plants. However, the popS mutant had wild-type virulence on a weed host, Solanum dulcamara, suggesting that some species can avoid the effects of PopS. The popS mutant was also significantly delayed in colonization of tomato stems compared to the wild type. Some AvrE-type effectors from gammaproteobacteria suppress salicylic acid (SA)-mediated plant defenses, suggesting that PopS, a betaproteobacterial ortholog, has a similar function. Indeed, the popS mutant induced significantly higher expression of tomato SA-triggered pathogenesis-related (PR) genes than the wild type. Further, pretreatment of roots with SA exacerbated the popS mutant virulence defect. Finally, the popS mutant had no colonization defect on SA-deficient NahG transgenic tomato plants. Together, these results indicate that this conserved effector suppresses SA-mediated defenses in tomato roots and stems, which are R. solanacearum’s natural infection sites. Interestingly, PopS did not trigger necrosis when heterologously expressed in Nicotiana leaf tissue, unlike the AvrE homolog DspEPcc from the necrotroph Pectobacterium carotovorum subsp. carotovorum. This is consistent with the differing pathogenesis modes of necrosis-causing gammaproteobacteria and biotrophic R. solanacearum. PMID:24281716

  17. Hepatic effector CD8+ T-cell dynamics

    PubMed Central

    Iannacone, Matteo

    2015-01-01

    CD8+ T cells play a critical role in hepatitis B virus (HBV) pathogenesis. During acute, self-limited infections, these cells are instrumental to viral clearance; in chronic settings, they sustain repetitive cycles of hepatocellular necrosis that promote hepatocellular carcinoma development. Both CD8+ T-cell defensive and destructive functions are mediated by antigen-experienced effector cells and depend on the ability of these cells to migrate to the liver, recognize hepatocellular antigens and perform effector functions. Understanding the signals that modulate the spatiotemporal dynamics of CD8+ T cells in the liver, particularly in the context of antigen recognition, is therefore critical to gaining insight into the pathogenesis of acute and chronic HBV infection. Here, we highlight recent data on how effector CD8+ T cells traffic within the liver, and we discuss the potential for novel imaging techniques to shed light on this important aspect of HBV pathogenesis. PMID:25242274

  18. Effector-triggered defence against apoplastic fungal pathogens

    PubMed Central

    Stotz, Henrik U.; Mitrousia, Georgia K.; de Wit, Pierre J.G.M.; Fitt, Bruce D.L.

    2014-01-01

    R gene-mediated host resistance against apoplastic fungal pathogens is not adequately explained by the terms pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) or effector-triggered immunity (ETI). Therefore, it is proposed that this type of resistance is termed ‘effector-triggered defence’ (ETD). Unlike PTI and ETI, ETD is mediated by R genes encoding cell surface-localised receptor-like proteins (RLPs) that engage the receptor-like kinase SOBIR1. In contrast to this extracellular recognition, ETI is initiated by intracellular detection of pathogen effectors. ETI is usually associated with fast, hypersensitive host cell death, whereas ETD often triggers host cell death only after an elapsed period of endophytic pathogen growth. In this opinion, we focus on ETD responses against foliar fungal pathogens of crops. PMID:24856287

  19. Effector and suppressor T cells in celiac disease

    PubMed Central

    Mazzarella, Giuseppe

    2015-01-01

    Celiac disease (CD) is a T-cell mediated immune disease in which gliadin-derived peptides activate lamina propria effector CD4+ T cells. This activation leads to the release of cytokines, compatible with a Th1-like pattern, which play a crucial role in the pathogenesis of CD, controlling many aspects of the inflammatory immune response. Recent studies have shown that a novel subset of effector T cells, characterized by expression of high levels of IL-17A, termed Th17 cells, plays a pathogenic role in CD. While these effector T cell subsets produce proinflammatory cytokines, which cause substantial tissue injury in vivo in CD, recent studies have suggested the existence of additional CD4+ T cell subsets with suppressor functions. These subsets include type 1 regulatory T cells and CD25+CD4+ regulatory T cells, expressing the master transcription factor Foxp3, which have important implications for disease progression. PMID:26139981

  20. Development and testing of the cooling coil cleaning end effector

    SciTech Connect

    Johnson, K.I.; Mullen, O.D.; Powell, M.R.; Daly, D.S.; Engel, D.W.

    1997-09-30

    The Retrieval Process Development and Enhancement (KPD{ampersand}E) program has developed and tested an end effector to support the waste retrieval mission at the Idaho National Engineering and Environmental Laboratory (INEEL). The end effector was developed specifically to remove a sticky waste material from the cooling coils in the High Level Liquid Waste (HLLW) tank, and to vacuum up a sediment layer that has settled beneath the cooling coils. An extensive testing program was conducted in the hydraulic test bed (HTB) at the Pacific Northwest National Laboratory (PNNL) to evaluate the performance of the end effector under simulated in-tank conditions. A mock up of the cooling coils was installed in the test bed tank, and simulated waste materials were included to represent the sticky waste on the tubes and the particulate waste settled beneath them. The testing program focused on assessing long-duration mining strategies for cleaning the cooling coils and removing the particulate waste forms. The report describes the results of the end effector testing program at PNNL. Section 2 describes the physical characteristics of the HLLW tanks, including the layout of the cooling coils, and it also describes what is known of the waste forms in the tanks. Section 3 describes the cleaning and retrieval strategy that was used in developing the end effector design. Section 4 describes the cooling coil mockup in the hydraulic test bed. Section 5 discusses the rationale used in selecting the simulants for the tarry waste and particulate waste forms. Section 6 describes the tests that were performed to evaluate cleaning of the cooling coils and retrieval of the particulate simulant. Section 7 summarizes the cleaning and retrieval tests, assesses the relative importance of cleaning the cooling coils and retrieving the particulate waste, and suggests modifications that would simplify the end effector design.

  1. Tyrosine phosphorylation of RAS by ABL allosterically enhances effector binding.

    PubMed

    Ting, Pamela Y; Johnson, Christian W; Fang, Cong; Cao, Xiaoqing; Graeber, Thomas G; Mattos, Carla; Colicelli, John

    2015-09-01

    RAS proteins are signal transduction gatekeepers that mediate cell growth, survival, and differentiation through interactions with multiple effector proteins. The RAS effector RAS- and RAB-interacting protein 1 (RIN1) activates its own downstream effectors, the small GTPase RAB5 and the tyrosine kinase Abelson tyrosine-protein kinase (ABL), to modulate endocytosis and cytoskeleton remodeling. To identify ABL substrates downstream of RAS-to-RIN1 signaling, we examined human HEK293T cells overexpressing components of this pathway. Proteomic analysis revealed several novel phosphotyrosine peptides, including Harvey rat sarcoma oncogene (HRAS)-pTyr(137). Here we report that ABL phosphorylates tyrosine 137 of H-, K-, and NRAS. Increased RIN1 levels enhanced HRAS-Tyr(137) phosphorylation by nearly 5-fold, suggesting that RAS-stimulated RIN1 can drive ABL-mediated RAS modification in a feedback circuit. Tyr(137) is well conserved among RAS orthologs and is part of a transprotein H-bond network. Crystal structures of HRAS(Y137F) and HRAS(Y137E) revealed conformation changes radiating from the mutated residue. Although consistent with Tyr(137) participation in allosteric control of HRAS function, the mutations did not alter intrinsic GTP hydrolysis rates in vitro. HRAS-Tyr(137) phosphorylation enhanced HRAS signaling capacity in cells, however, as reflected by a 4-fold increase in the association of phosphorylated HRAS(G12V) with its effector protein RAF proto-oncogene serine/threonine protein kinase 1 (RAF1). These data suggest that RAS phosphorylation at Tyr(137) allosterically alters protein conformation and effector binding, providing a mechanism for effector-initiated modulation of RAS signaling. PMID:25999467

  2. Nanorobotic end-effectors: Design, fabrication, and in situ characterization

    NASA Astrophysics Data System (ADS)

    Fan, Zheng

    Nano-robotic end-effectors have promising applications for nano-fabrication, nano-manufacturing, nano-optics, nano-medical, and nano-sensing; however, low performances of the conventional end-effectors have prevented the widespread utilization of them in various fields. There are two major difficulties in developing the end-effectors: their nano-fabrication and their advanced characterization in the nanoscale. Here we introduce six types of end-effectors: the nanotube fountain pen (NFP), the super-fine nanoprobe, the metal-filled carbon nanotube (m CNT)-based sphere-on-pillar (SOP) nanoantennas, the tunneling nanosensor, and the nanowire-based memristor. The investigations on the NFP are focused on nano-fluidics and nano-fabrications. The NFP could direct write metallic "inks" and fabricating complex metal nanostructures from 0D to 3D with a position servo control, which is critically important to future large-scale, high-throughput nanodevice production. With the help of NFP, we could fabricate the end-effectors such as super-fine nanoprobe and m CNT-based SOP nanoantennas. Those end-effectors are able to detect local flaws or characterize the electrical/mechanical properties of the nanostructure. Moreover, using electron-energy-loss-spectroscopy (EELS) technique during the operation of the SOP optical antenna opens a new basis for the application of nano-robotic end-effectors. The technique allows advanced characterization of the physical changes, such as carrier diffusion, that are directly responsible for the device's properties. As the device was coupled with characterization techniques of scanning-trasmission-electron-microscopy (STEM), the development of tunneling nanosensor advances this field of science into quantum world. Furthermore, the combined STEM-EELS technique plays an important role in our understanding of the memristive switching performance in the nanowire-based memristor. The developments of those nano-robotic end-effectors expend the study abilities in investigating the in situ nanotechnology, providing efficient ways in in situ nanostructure fabrication and the advanced characterization of the nanomaterials.

  3. How Do Filamentous Pathogens Deliver Effector Proteins into Plant Cells?

    PubMed Central

    Petre, Benjamin; Kamoun, Sophien

    2014-01-01

    Fungal and oomycete plant parasites are among the most devastating pathogens of food crops. These microbes secrete effector proteins inside plant cells to manipulate host processes and facilitate colonization. How these effectors reach the host cytoplasm remains an unclear and debated area of plant research. In this article, we examine recent conflicting findings that have generated discussion in the field. We also highlight promising approaches based on studies of both parasite and host during infection. Ultimately, this knowledge may inform future broad spectrum strategies for protecting crops from such pathogens. PMID:24586116

  4. Visual End-Effector Position Error Compensation for Planetary Robotics

    NASA Technical Reports Server (NTRS)

    Bajracharya, Max; DiCicco, Matthew; Backes, Paul; Nickels, Kevin

    2007-01-01

    This paper describes a vision-guided manipulation algorithm that improves arm end-effector positioning to subpixel accuracy and meets the highly restrictive imaging and computational constraints of a planetary robotic flight system. Analytical, simulation-based, and experimental analyses of the algorithm's effectiveness and sensitivity to camera and arm model error is presented along with results on several prototype research systems and 'ground-in-the-loop' technology experiments on the Mars Exploration Rover (MER) vehicles. A computationally efficient and robust subpixel end-effector fiducial detector that is instrumental to the algorithm's ability to achieve high accuracy is also described along with its validation results on MER data.

  5. Phytophthora infestans RXLR-WY Effector AVR3a Associates with Dynamin-Related Protein 2 Required for Endocytosis of the Plant Pattern Recognition Receptor FLS2

    PubMed Central

    Chaparro-Garcia, Angela; Schwizer, Simon; Sklenar, Jan; Yoshida, Kentaro; Petre, Benjamin; Bos, Jorunn I. B.; Schornack, Sebastian; Jones, Alexandra M. E.; Bozkurt, Tolga O.; Kamoun, Sophien

    2015-01-01

    Pathogens utilize effectors to suppress basal plant defense known as PTI (Pathogen-associated molecular pattern-triggered immunity). However, our knowledge of PTI suppression by filamentous plant pathogens, i.e. fungi and oomycetes, remains fragmentary. Previous work revealed that the co-receptor BAK1/SERK3 contributes to basal immunity against the potato pathogen Phytophthora infestans. Moreover BAK1/SERK3 is required for the cell death induced by P. infestans elicitin INF1, a protein with characteristics of PAMPs. The P. infestans host-translocated RXLR-WY effector AVR3a is known to supress INF1-mediated cell death by binding the plant E3 ligase CMPG1. In contrast, AVR3aKI-Y147del, a deletion mutant of the C-terminal tyrosine of AVR3a, fails to bind CMPG1 and does not suppress INF1-mediated cell death. Here, we studied the extent to which AVR3a and its variants perturb additional BAK1/SERK3-dependent PTI responses in N. benthamiana using the elicitor/receptor pair flg22/FLS2 as a model. We found that all tested variants of AVR3a suppress defense responses triggered by flg22 and reduce internalization of activated FLS2. Moreover, we discovered that AVR3a associates with the Dynamin-Related Protein 2 (DRP2), a plant GTPase implicated in receptor-mediated endocytosis. Interestingly, silencing of DRP2 impaired ligand-induced FLS2 internalization but did not affect internalization of the growth receptor BRI1. Our results suggest that AVR3a associates with a key cellular trafficking and membrane-remodeling complex involved in immune receptor-mediated endocytosis. We conclude that AVR3a is a multifunctional effector that can suppress BAK1/SERK3-mediated immunity through at least two different pathways. PMID:26348328

  6. Structure Analysis Uncovers a Highly Diverse but Structurally Conserved Effector Family in Phytopathogenic Fungi.

    PubMed

    de Guillen, Karine; Ortiz-Vallejo, Diana; Gracy, Jérome; Fournier, Elisabeth; Kroj, Thomas; Padilla, André

    2015-10-01

    Phytopathogenic ascomycete fungi possess huge effector repertoires that are dominated by hundreds of sequence-unrelated small secreted proteins. The molecular function of these effectors and the evolutionary mechanisms that generate this tremendous number of singleton genes are largely unknown. To get a deeper understanding of fungal effectors, we determined by NMR spectroscopy the 3-dimensional structures of the Magnaporthe oryzae effectors AVR1-CO39 and AVR-Pia. Despite a lack of sequence similarity, both proteins have very similar 6 ?-sandwich structures that are stabilized in both cases by a disulfide bridge between 2 conserved cysteins located in similar positions of the proteins. Structural similarity searches revealed that AvrPiz-t, another effector from M. oryzae, and ToxB, an effector of the wheat tan spot pathogen Pyrenophora tritici-repentis have the same structures suggesting the existence of a family of sequence-unrelated but structurally conserved fungal effectors that we named MAX-effectors (Magnaporthe Avrs and ToxB like). Structure-informed pattern searches strengthened this hypothesis by identifying MAX-effector candidates in a broad range of ascomycete phytopathogens. Strong expansion of the MAX-effector family was detected in M. oryzae and M. grisea where they seem to be particularly important since they account for 5-10% of the effector repertoire and 50% of the cloned avirulence effectors. Expression analysis indicated that the majority of M. oryzae MAX-effectors are expressed specifically during early infection suggesting important functions during biotrophic host colonization. We hypothesize that the scenario observed for MAX-effectors can serve as a paradigm for ascomycete effector diversity and that the enormous number of sequence-unrelated ascomycete effectors may in fact belong to a restricted set of structurally conserved effector families. PMID:26506000

  7. Structure Analysis Uncovers a Highly Diverse but Structurally Conserved Effector Family in Phytopathogenic Fungi

    PubMed Central

    Gracy, Jérome; Fournier, Elisabeth; Kroj, Thomas; Padilla, André

    2015-01-01

    Phytopathogenic ascomycete fungi possess huge effector repertoires that are dominated by hundreds of sequence-unrelated small secreted proteins. The molecular function of these effectors and the evolutionary mechanisms that generate this tremendous number of singleton genes are largely unknown. To get a deeper understanding of fungal effectors, we determined by NMR spectroscopy the 3-dimensional structures of the Magnaporthe oryzae effectors AVR1-CO39 and AVR-Pia. Despite a lack of sequence similarity, both proteins have very similar 6 ?-sandwich structures that are stabilized in both cases by a disulfide bridge between 2 conserved cysteins located in similar positions of the proteins. Structural similarity searches revealed that AvrPiz-t, another effector from M. oryzae, and ToxB, an effector of the wheat tan spot pathogen Pyrenophora tritici-repentis have the same structures suggesting the existence of a family of sequence-unrelated but structurally conserved fungal effectors that we named MAX-effectors (Magnaporthe Avrs and ToxB like). Structure-informed pattern searches strengthened this hypothesis by identifying MAX-effector candidates in a broad range of ascomycete phytopathogens. Strong expansion of the MAX-effector family was detected in M. oryzae and M. grisea where they seem to be particularly important since they account for 5–10% of the effector repertoire and 50% of the cloned avirulence effectors. Expression analysis indicated that the majority of M. oryzae MAX-effectors are expressed specifically during early infection suggesting important functions during biotrophic host colonization. We hypothesize that the scenario observed for MAX-effectors can serve as a paradigm for ascomycete effector diversity and that the enormous number of sequence-unrelated ascomycete effectors may in fact belong to a restricted set of structurally conserved effector families. PMID:26506000

  8. Robotic End Effectors for Hard-Rock Climbing

    NASA Technical Reports Server (NTRS)

    Kennedy, Brett; Leger, Patrick

    2004-01-01

    Special-purpose robot hands (end effectors) now under development are intended to enable robots to traverse cliffs much as human climbers do. Potential applications for robots having this capability include scientific exploration (both on Earth and other rocky bodies in space), military reconnaissance, and outdoor search and rescue operations. Until now, enabling robots to traverse cliffs has been considered too difficult a task because of the perceived need of prohibitively sophisticated planning algorithms as well as end effectors as dexterous as human hands. The present end effectors are being designed to enable robots to attach themselves to typical rock-face features with less planning and simpler end effectors. This advance is based on the emulation of the equipment used by human climbers rather than the emulation of the human hand. Climbing-aid equipment, specifically cams, aid hooks, and cam hooks, are used by sport climbers when a quick ascent of a cliff is desired (see Figure 1). Currently two different end-effector designs have been created. The first, denoted the simple hook emulator, consists of three "fingers" arranged around a central "palm." Each finger emulates the function of a particular type of climbing hook (aid hook, wide cam hook, and a narrow cam hook). These fingers are connected to the palm via a mechanical linkage actuated with a leadscrew/nut. This mechanism allows the fingers to be extended or retracted. The second design, denoted the advanced hook emulator (see Figure 2), shares these features, but it incorporates an aid hook and a cam hook into each finger. The spring-loading of the aid hook allows the passive selection of the type of hook used. The end effectors can be used in several different modes. In the aid-hook mode, the aid hook on one of the fingers locks onto a horizontal ledge while the other two fingers act to stabilize the end effector against the cliff face. In the cam-hook mode, the broad, flat tip of the cam hook is inserted into a non-horizontal crack in the cliff face. A subsequent transfer of weight onto the end effector causes the tip to rotate within the crack, creating a passive, self-locking action of the hook relative to the crack. In the advanced hook emulator, the aid hook is pushed into its retracted position by contact with the cliff face as the cam hook tip is inserted into the crack. When a cliff face contains relatively large pockets or cracks, another type of passive self-locking can be used. Emulating the function of the piece of climbing equipment called a "cam" (note: not the same as a "cam hook"; see Figure 1), the fingers can be fully retracted and the entire end effector inserted into the feature. The fingers are then extended as far as the feature allows. Any weight then transferred to the end effector will tend to extend the fingers further due to frictional force, passively increasing the grip on the feature. In addition to the climbing modes, these end effectors can be used to walk on (either on the palm or the fingertips) and to grasp objects by fully extending the fingers.

  9. Expression of Enteropathogenic Escherichia coli Map Is Significantly Different than That of Other Type III Secreted Effectors In Vivo

    PubMed Central

    Nguyen, Mai; Rizvi, Jason

    2014-01-01

    The enteropathogenic Escherichia coli (EPEC) locus of enterocyte effacement (LEE)-encoded effectors EspF and Map are multifunctional and have an impact on the tight junction barrier while the non-LEE-encoded proteins NleH1 and NleH2 possess significant anti-inflammatory activity. In order to address the temporal expression of these important genes in vivo, their promoters were cloned upstream of the luxCDABE operon, and luciferase expression was measured in EPEC-infected mice by bioluminescence using an in vivo imaging system (IVIS). Bioluminescent images of living mice, of excised whole intestines, and of whole intestines longitudinally opened and washed were assessed. The majority of bioluminescent bacteria localized in the cecum by 3 h postinfection, indicating that the cecum is not only a major colonization site of EPEC but also a site of EPEC effector gene expression in mice. espF, nleH1, and nleH2 were abundantly expressed over the course of infection. In contrast, map expression was suppressed at 2 days postinfection, and at 4 days postinfection it was totally abolished. After 2 to 4 days postinfection, when map is suppressed, EPEC colonization is significantly reduced, indicating that map may be one of the factors required to maintain EPEC colonization. This was confirmed in a competitive colonization study and in two models of chronic infection, repeated exposure to ketamine and Citrobacter rodentium infection. Our data suggest that map expression contributes to the maintenance of EPEC colonization. PMID:25312947

  10. N-Glycosylation of Effector Proteins by an ?-1,3-Mannosyltransferase Is Required for the Rice Blast Fungus to Evade Host Innate Immunity[W][OPEN

    PubMed Central

    Chen, Xiao-Lin; Shi, Tao; Yang, Jun; Shi, Wei; Gao, Xusheng; Chen, Deng; Xu, Xiaowen; Xu, Jin-Rong; Talbot, Nicholas J.; Peng, You-Liang

    2014-01-01

    Plant pathogenic fungi deploy secreted effectors to suppress plant immunity responses. These effectors operate either in the apoplast or within host cells, so they are putatively glycosylated, but the posttranslational regulation of their activities has not been explored. In this study, the ASPARAGINE-LINKED GLYCOSYLATION3 (ALG3)-mediated N-glycosylation of the effector, Secreted LysM Protein1 (Slp1), was found to be essential for its activity in the rice blast fungus Magnaporthe oryzae. ALG3 encodes an ?-1,3-mannosyltransferase for protein N-glycosylation. Deletion of ALG3 resulted in the arrest of secondary infection hyphae and a significant reduction in virulence. We observed that ?alg3 mutants induced massive production of reactive oxygen species in host cells, in a similar manner to ?slp1 mutants, which is a key factor responsible for arresting infection hyphae of the mutants. Slp1 sequesters chitin oligosaccharides to avoid their recognition by the rice (Oryza sativa) chitin elicitor binding protein CEBiP and the induction of innate immune responses, including reactive oxygen species production. We demonstrate that Slp1 has three N-glycosylation sites and that simultaneous Alg3-mediated N-glycosylation of each site is required to maintain protein stability and the chitin binding activity of Slp1, which are essential for its effector function. These results indicate that Alg3-mediated N-glycosylation of Slp1 is required to evade host innate immunity. PMID:24642938

  11. The HrpN effector of Erwinia amylovora, which is involved in type III translocation, contributes directly or indirectly to callose elicitation on apple leaves.

    PubMed

    Boureau, Tristan; Siamer, Sabrina; Perino, Claude; Gaubert, Stéphane; Patrit, Oriane; Degrave, Alexandre; Fagard, Mathilde; Chevreau, Elisabeth; Barny, Marie-Anne

    2011-05-01

    Erwinia amylovora is responsible for fire blight of apple and pear trees. Its pathogenicity depends on a type III secretion system (T3SS) mediating the translocation of effectors into the plant cell. The DspA/E effector suppresses callose deposition on apple leaves. We found that E. amylovora and Pseudomonas syringae DC3000 tts mutants or peptide flg22 do not trigger callose deposition as strongly as the dspA/E mutant on apple leaves. This suggests that, on apple leaves, callose deposition is poorly elicited by pathogen-associated molecular patterns (PAMPs) such as flg22 or other PAMPs harbored by tts mutants and is mainly elicited by injected effectors or by the T3SS itself. Callose elicitation partly depends on HrpW because an hrpW-dspA/E mutant elicits lower callose deposition than a dspA/E mutant. Furthermore, an hrpN-dspA/E mutant does not trigger callose deposition, indicating that HrpN is required to trigger this plant defense reaction. We showed that HrpN plays a general role in the translocation process. Thus, the HrpN requirement for callose deposition may be explained by its role in translocation: HrpN could be involved in the translocation of other effectors inducing callose deposition. Furthermore, HrpN may also directly contribute to the elicitation process because we showed that purified HrpN induces callose deposition. PMID:21463207

  12. Robot End Effector To Place and Solder Solar Cells

    NASA Technical Reports Server (NTRS)

    Hagerty, J. J.

    1982-01-01

    Encapsulated in robot end effector is RF induction-heating coil for heating solar cell while in transit. Holes in encapsulant permit end of unit to act as vacuum pickup to grip solar cell. Use of RF induction heating allows cell to be heated without requiring direct mechanical and thermal contact of bonding tool such as soldering iron.

  13. Hand to Mouth: Automatic Imitation across Effector Systems

    ERIC Educational Resources Information Center

    Leighton, Jane; Heyes, Cecilia

    2010-01-01

    The effector dependence of automatic imitation was investigated using a stimulus-response compatibility (SRC) procedure during which participants were required to make an open or closed response with their hand or their mouth. The correct response for each trial was indicated by a pair of letters in Experiments 1 and 2 and by a colored square in…

  14. Type IV secretion system of Brucella spp. and its effectors

    PubMed Central

    Ke, Yuehua; Wang, Yufei; Li, Wengfeng; Chen, Zeliang

    2015-01-01

    Brucella spp. are intracellular bacterial pathogens that cause infection in domestic and wild animals. They are often used as model organisms to study intracellular bacterial infections. Brucella VirB T4SS is a key virulence factor that plays important roles in mediating intracellular survival and manipulating host immune response to infection. In this review, we discuss the roles of Brucella VirB T4SS and 15 effectors that are proposed to be crucial for Brucella pathogenesis. VirB T4SS regulates the inflammation response and manipulates vesicle trafficking inside host cells. VirB T4SS also plays crucial roles in the inhibition of the host immune response and intracellular survival during infection. Here, we list the key molecular events in the intracellular life cycle of Brucella that are potentially targeted by the VirB T4SS effectors. Elucidating the functions of these effectors will help clarify the molecular role of T4SS during infection. Furthermore, studying the effectors secreted by Brucella spp. might provide insights into the mechanisms used by the bacteria to hijack the host signaling pathways and aid in the development of better vaccines and therapies against brucellosis. PMID:26528442

  15. Plasmodium cellular effector mechanisms and the hepatic microenvironment

    PubMed Central

    Frevert, Ute; Krzych, Urszula

    2015-01-01

    Plasmodium falciparum malaria remains one of the most serious health problems globally. Immunization with attenuated parasites elicits multiple cellular effector mechanisms capable of eliminating Plasmodium liver stages. However, malaria liver stage (LS) immunity is complex and the mechanisms effector T cells use to locate the few infected hepatocytes in the large liver in order to kill the intracellular LS parasites remain a mystery to date. Here, we review our current knowledge on the behavior of CD8 effector T cells in the hepatic microvasculature, in malaria and other hepatic infections. Taking into account the unique immunological and lymphogenic properties of the liver, we discuss whether classical granule-mediated cytotoxicity might eliminate infected hepatocytes via direct cell contact or whether cytokines might operate without cell–cell contact and kill Plasmodium LSs at a distance. A thorough understanding of the cellular effector mechanisms that lead to parasite death hence sterile protection is a prerequisite for the development of a successful malaria vaccine to protect the 40% of the world’s population currently at risk of Plasmodium infection. PMID:26074888

  16. Developmental control of integrin expression regulates Th2 effector homing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Integrin CD18, a component of the LFA-1 complex that also includes CD11a, is essential for Th2, but not Th1, cell homing, but the explanation for this phenomenon remains obscure. In this study, we investigate the mechanism by which Th2 effector responses require the LFA-1 complex. CD11a-deficient T ...

  17. Electroporation of Functional Bacterial Effectors into Mammalian Cells

    SciTech Connect

    Sontag, Ryan L.; Mihai, Cosmin; Orr, Galya; Savchenko, Alexei; Skarina, Tatiana; Cui, Hong; Cort, John R.; Adkins, Joshua N.; Brown, Roslyn N.

    2015-01-01

    Electroporation was used to insert purified bacterial virulence effector proteins directly into living eukaryotic cells. Protein localization was monitored by confocal immunofluorescence microscopy. This method allows for studies on trafficking, function, and protein-protein interactions using active exogenous proteins, avoiding the need for heterologous expression in eukaryotic cells.

  18. Pseudomonas syringae Hrp type III secretion system and effector proteins

    E-print Network

    and Pseudomonas fluorescens to inject HopPsyA into tobacco cells, thereby eliciting a hypersensitive responseColloquium Pseudomonas syringae Hrp type III secretion system and effector proteins Alan Collmer; and ¶Department of Biological Sciences, University of Nevada, Las Vegas, NV 89154-4004 Pseudomonas syringae

  19. TAL effector-mediated susceptibility to bacterial blight of cotton

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacterial blight of cotton (BBC) caused by Xanthomonas campestris pv. malvacearum (Xcm) is a destructive disease that has recently re-emerged in the U.S. Xcm injects transcription activator-like (TAL) effectors that directly induce the expression of host susceptibility (S) or resistance (R) genes. ...

  20. Consequences of exposure to ionizing radiation for effector T cell function in vivo

    SciTech Connect

    Rouse, B.T.; Hartley, D.; Doherty, P.C. )

    1989-01-01

    The adoptive transfer of acutely primed and memory virus-immune CD8+ T cells causes enhanced meningitis in both cyclophosphamide (Cy) suppressed, and unsuppressed, recipients infected with lymphocytic choriomeningitis virus (LCMV). The severity of meningitis is assessed by counting cells in cerebrospinal fluid (CSF) obtained from the cisterna magna, which allows measurement of significant inflammatory process ranging from 3 to more than 300 times the background number of cells found in mice injected with virus alone. Exposure of the donor immune population to ionizing radiation prior to transfer has shown that activated T cells from mice primed 7 or 8 days previously with virus may still promote a low level of meningitis in unsuppressed recipients following as much as 800 rads, while this effect is lost totally in Cy-suppressed mice at 600 rads. Memory T cells are more susceptible and show no evidence of in vivo effector function in either recipient population subsequent to 400 rads, a dose level which also greatly reduces the efficacy of acutely-primed T cells. The results are interpreted as indicating that heavily irradiated cells that are already fully functional show evidence of primary localization to the CNS and a limited capacity to cause pathology. Secondary localization, and events that require further proliferation of the T cells in vivo, are greatly inhibited by irradiation.

  1. Distinct Pseudomonas type-III effectors use a cleavable transit peptide to target chloroplasts

    E-print Network

    -III effectors is the suppres- sion of plant immunity. The P. syringae pv. tomato DC3000 HopK1 type-III effector with effector-triggered immunity. Here we show that DC3000 hopK1 mutants are reduced in their ability to grow and the effector proteins it injects into plant cells for pathogenesis. The primary role for P. syringae type

  2. 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.

  3. Identification of divergent type VI secretion effectors using a conserved chaperone domain.

    PubMed

    Liang, Xiaoye; Moore, Richard; Wilton, Mike; Wong, Megan J Q; Lam, Linh; Dong, Tao G

    2015-07-21

    The type VI secretion system (T6SS) is a lethal weapon used by many bacteria to kill eukaryotic predators or prokaryotic competitors. Killing by the T6SS results from repetitive delivery of toxic effectors. Despite their importance in dictating bacterial fitness, systematic prediction of T6SS effectors remains challenging due to high effector diversity and the absence of a conserved signature sequence. Here, we report a class of T6SS effector chaperone (TEC) proteins that are required for effector delivery through binding to VgrG and effector proteins. The TEC proteins share a highly conserved domain (DUF4123) and are genetically encoded upstream of their cognate effector genes. Using the conserved TEC domain sequence, we identified a large family of TEC genes coupled to putative T6SS effectors in Gram-negative bacteria. We validated this approach by verifying a predicted effector TseC in Aeromonas hydrophila. We show that TseC is a T6SS-secreted antibacterial effector and that the downstream gene tsiC encodes the cognate immunity protein. Further, we demonstrate that TseC secretion requires its cognate TEC protein and an associated VgrG protein. Distinct from previous effector-dependent bioinformatic analyses, our approach using the conserved TEC domain will facilitate the discovery and functional characterization of new T6SS effectors in Gram-negative bacteria. PMID:26150500

  4. INTRODUCTION Temperature has long been recognized as a de facto effector of

    E-print Network

    Campbell, Kevin L.

    -linked, endothermic dissociation of allosteric effectors. In vertebrate Hbs, where the main effectors are protons in metabolizing, acidic tissues) that is expressed at physiological pH, vertebrate Hbs may additionally exhibit is a more potent allosteric effector than ATP (Weber, 2000). The endothermic dissociation of red cell

  5. Phytopathogenic bacteria deliver effectors of disease into plant hosts via a Type III secretion system. These Type III effectors

    E-print Network

    Dangl, Jeff

    pathogenicity island R resistance Introduction Pathogenic bacteria invading plant tissues must survive a defense73 Phytopathogenic bacteria deliver effectors of disease into plant hosts via a Type III secretion in the understanding of plant defense mechanisms, bacterial secretion systems, and pathogenicity islands reveals

  6. Caspase 2-mediated tumor suppression involves survivin gene silencing.

    PubMed

    Guha, M; Xia, F; Raskett, C M; Altieri, D C

    2010-03-01

    One of the pivotal functions of endogenous tumor suppression is to oppose aberrant cell survival, but the molecular requirements of this process are not completely understood. Here, we show that caspase 2, a death effector with largely unknown functions, represses transcription of the survivin gene, a general regulator of cell division and cytoprotection in tumors. This pathway involves caspase 2 proteolytic cleavage of the nuclear factor kappaB (NFkappaB) activator, RIP1. In turn, loss of RIP1 abolishes transcription of NFkappaB target genes, including survivin, resulting in deregulated mitotic transitions, enhanced apoptosis and suppression of tumorigenicity in vivo. Therefore, caspase 2 functions as an endogenous inhibitor of NFkappaB-dependent cell survival and this mechanism may contribute to tumor suppression in humans. PMID:19935698

  7. A genome-wide analysis of antimicrobial effector genes and their transcription patterns in Manduca sexta

    PubMed Central

    He, Yan; Cao, Xiaolong; Li, Kai; Hu, Yingxia; Chen, Yun-ru; Blissard, Gary; Kanost, Michael R.; Jiang, Haobo

    2015-01-01

    Antimicrobial proteins/peptides (AMPs) are effectors of innate immune systems against pathogen infection in multicellular organisms. Over half of the AMPs reported so far come from insects, and these effectors act in concert to suppress or kill bacteria, fungi, viruses, and parasites. In this work, we have identified 86 AMP genes in the Manduca sexta genome, most of which seem likely to be functional. They encode 15 cecropins, 6 moricins, 6 defensins, 3 gallerimycins, 4 X-tox splicing variants, 14 diapausins, 15 whey acidic protein homologs, 11 attacins, 1 gloverin, 4 lebocins, 6 lysozyme-related proteins, and 4 transferrins. Some of these genes (e.g. attacins, cecropins) constitute large clusters, likely arising after rounds of gene duplication. We compared the amino acid sequences of M. sexta AMPs with their homologs in other insects to reveal conserved structural features and phylogenetic relationships. Expression data showed that many of them are synthesized in fat body and midgut during the larval-pupal molt. Certain genes contain one or more predicted ?B binding sites and other regulatory elements in their promoter regions, which may account for the dramatic mRNA level increases in fat body and hemocytes after an immune challenge. Consistent with these strong mRNA increases, many AMPs become highly abundant in the larval plasma at 24 h after the challenge, as demonstrated in our previous peptidomic study. Taken together, these data suggest the existence of a large repertoire of AMPs in M. sexta, whose expression is up-regulated via immune signaling pathways to fight off invading pathogens in a coordinated manner. PMID:25662101

  8. A genome-wide analysis of antimicrobial effector genes and their transcription patterns in Manduca sexta.

    PubMed

    He, Yan; Cao, Xiaolong; Li, Kai; Hu, Yingxia; Chen, Yun-ru; Blissard, Gary; Kanost, Michael R; Jiang, Haobo

    2015-07-01

    Antimicrobial proteins/peptides (AMPs) are effectors of innate immune systems against pathogen infection in multicellular organisms. Over half of the AMPs reported so far come from insects, and these effectors act in concert to suppress or kill bacteria, fungi, viruses, and parasites. In this work, we have identified 86 AMP genes in the Manduca sexta genome, most of which seem likely to be functional. They encode 15 cecropins, 6 moricins, 6 defensins, 3 gallerimycins, 4 X-tox splicing variants, 14 diapausins, 15 whey acidic protein homologs, 11 attacins, 1 gloverin, 4 lebocins, 6 lysozyme-related proteins, and 4 transferrins. Some of these genes (e.g. attacins, cecropins) constitute large clusters, likely arising after rounds of gene duplication. We compared the amino acid sequences of M. sexta AMPs with their homologs in other insects to reveal conserved structural features and phylogenetic relationships. Expression data showed that many of them are synthesized in fat body and midgut during the larval-pupal molt. Certain genes contain one or more predicted ?B binding sites and other regulatory elements in their promoter regions, which may account for the dramatic mRNA level increases in fat body and hemocytes after an immune challenge. Consistent with these strong mRNA increases, many AMPs become highly abundant in the larval plasma at 24 h after the challenge, as demonstrated in our previous peptidomic study. Taken together, these data suggest the existence of a large repertoire of AMPs in M. sexta, whose expression is up-regulated via immune signaling pathways to fight off invading pathogens in a coordinated manner. PMID:25662101

  9. 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

  10. The effect of chitin metabolic effectors on the population increase of stored product mites.

    PubMed

    Stara, Jitka; Erban, Tomas; Hubert, Jan

    2010-10-01

    The study tested the effect of the chitin metabolic effectors, teflubenzuron, diflubenzuron, and calcofluor, and a combination of a chitinase and soybean trypsin inhibitor (STI) on the population growth of eight species of stored product mites under laboratory conditions. The compounds were incorporated into the diets of the mites in concentrations ranging from 0.01 to 50 mg g(-1). The final populations of mites were observed after 21 days of growth in controlled conditions. Diflubenzuron and calcofluor suppressed the growth of all the tested species more effectively than the other compounds. The doses required to suppress the mite populations to 50% (rc(50)) in comparison to the control ranged from 0.29 to 12.68 mg g(-1) for diflubenzuron and from 1.75 to 37.7 mg g(-1) for calcofluor, depending on the mite species. When tested at the highest concentration (10 mg g(-1)), teflubenzuron also suppressed all of the tested mite species in comparison to the control. The addition of chitinase/STI into the diet influenced population growth in several ways. When the highest concentration of chitinase in a cocktail of chitinase and STI (12.5 mg g(-1) of diet) was compared to the control, populations of Acarus siro, Aleuroglyphus ovatus and Aëroglyphus robustus decreased significantly, whereas populations of Tyroborus lini and Sancassania rodionovi increased significantly. The sensitivity of species to the tested compounds differed among species. The most tolerant species was S. rodionovi, the most sensitive was A. ovatus. The results confirmed that calcofluor and diflubenzuron have a toxic effect on stored product mites. PMID:20229097

  11. Dexamethasone suppression test

    MedlinePLUS

    DST; ACTH suppression test; Cortisol suppression test ... During this test, you will receive dexamethasone. This is a strong man-made (synthetic) glucocorticoid medication. Afterward, your blood is drawn ...

  12. Multiple recognition of RXLR effectors is associated with nonhost resistance of pepper against Phytophthora infestans

    PubMed Central

    Lee, Hyun-Ah; Kim, Shin-Young; Oh, Sang-Keun; Yeom, Seon-In; Kim, Saet-Byul; Kim, Myung-Shin; Kamoun, Sophien; Choi, Doil

    2014-01-01

    Nonhost resistance (NHR) is a plant immune response to resist most pathogens. The molecular basis of NHR is poorly understood, but recognition of pathogen effectors by immune receptors, a response known as effector-triggered immunity, has been proposed as a component of NHR. We performed transient expression of 54 Phytophthora infestansRXLR effectors in pepper (Capsicum annuum) accessions. We used optimized heterologous expression methods and analyzed the inheritance of effector-induced cell death in an F2 population derived from a cross between two pepper accessions. Pepper showed a localized cell death response upon inoculation with P. infestans, suggesting that recognition of effectors may contribute to NHR in this system. Pepper accessions recognized as many as 36 effectors. Among the effectors, PexRD8 and Avrblb2 induced cell death in a broad range of pepper accessions. Segregation of effector-induced cell death in an F2 population derived from a cross between two pepper accessions fit 15 : 1, 9 : 7 or 3 : 1 ratios, depending on the effector. Our genetic data suggest that a single or two independent/complementary dominant genes are involved in the recognition of RXLR effectors. Multiple loci recognizing a series of effectors may underpin NHR of pepper to P. infestans and confer resistance durability. PMID:24889686

  13. SseK3 Is a Salmonella Effector That Binds TRIM32 and Modulates the Host’s NF-?B 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-?B 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-?B dependent cytokine, IL-8, in HeLa cells even though TRIM32 knockdown suppressed TNF-induced NF-?B 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-?B activity. However, the lack of GlcNAc modification of TRIM32 suggests that SseK3 has further, as yet unidentified, host targets. PMID:26394407

  14. Interchangeable end effector tools utilized on the protoflight manipulator arm

    NASA Technical Reports Server (NTRS)

    1987-01-01

    A subset of teleoperator and effector tools was designed, fabricated, delivered and successfully demonstrated on the Marshall Space Flight Center (MSFC) protoflight manipulator arm (PFMA). The tools delivered included a rotary power tool with interchangeable collets and two fluid coupling mate/demate tools; one for a Fairchild coupling and the other for a Purolator coupling. An electrical interface connector was also provided for the rotary power tool. A tool set, from which the subset was selected, for performing on-orbit satellite maintenance was identified and conceptionally designed. Maintenance requirements were synthesized, evaluated and prioritized to develop design requirements for a set of end effector tools representative of those needed to provide on-orbit maintenance of satellites to be flown in the 1986 to 2000 timeframe.

  15. Proteomics of effector-triggered immunity (ETI) in plants

    PubMed Central

    Hurley, Brenden; Subramaniam, Rajagopal; Guttman, David S; Desveaux, Darrell

    2014-01-01

    Effector-triggered immunity (ETI) was originally termed gene-for-gene resistance and dates back to fundamental observations of flax resistance to rust fungi by Harold Henry Flor in the 1940s. Since then, genetic and biochemical approaches have defined our current understanding of how plant “resistance” proteins recognize microbial effectors. More recently, proteomic approaches have expanded our view of the protein landscape during ETI and contributed significant advances to our mechanistic understanding of ETI signaling. Here we provide an overview of proteomic techniques that have been used to study plant ETI including both global and targeted approaches. We discuss the challenges associated with ETI proteomics and highlight specific examples from the literature, which demonstrate how proteomics is advancing the ETI research field. PMID:25513776

  16. Autonomous dexterous end-effectors for space robotics

    NASA Technical Reports Server (NTRS)

    Bekey, George A.; Iberall, Thea; Liu, Huan

    1989-01-01

    The development of a knowledge-based controller is summarized for the Belgrade/USC robot hand, a five-fingered end effector, designed for maximum autonomy. The biological principles of the hand and its architecture are presented. The conceptual and software aspects of the grasp selection system are discussed, including both the effects of the geometry of the target object and the task to be performed. Some current research issues are presented.

  17. Xanthomonas and the TAL Effectors: Nature's Molecular Biologist.

    PubMed

    White, Frank

    2016-01-01

    Agrobacterium, due to the transfer of T-DNA to the host genome, is known as nature's genetic engineer. Once again, bacteria have led the way to newfound riches in biotechnology. Xanthomonas has emerged as nature's molecular biologist as the functional domains of the sequence-specific DNA transcription factors known as TAL effectors were characterized and associated with the cognate disease susceptibility and resistance genes of plants. PMID:26443209

  18. Mirror Sub-Assembly End-Effector Design

    SciTech Connect

    Butlin, B

    2007-01-08

    The Optic Assembly Building (OAB) is a facility where large optical mirror units are assembled and installed into Line Replaceable Units (LRUs) for deployment into the National Ignition Facility (NIF) laser system. The New Optics Insertion Device (NOID) is a powered jib crane specially designed to handle large optical assemblies. The NOID arm has three degrees of freedom. it can rotate about the vertical boom, travel up and down the boom, and extend away from and retract in towards the boom. The NOID is used to assist in the assembly of five types of Laser Mirror (LM) LRUs. These five LMs have been creatively named, LM4, LM5, LM6, LM7, and LM8. The LM4 and LM5 LRUs each contain four Mirror Sub-Assemblies (MSAs). The LM6, LM7, and LM8 LRUs each contain 2 MSAs. The MSAs are assembled apart from the LRU and are then installed in the LRU at the LM4-8 workstations. An MSA NOID End-Effector is required to interface with the MSAs and install them into the LRUs. The End-Effector must attach to the robo-hand on the end of the NOID arm. At the time the MSA NOID End-Effector was being designed the NOID, the LM4-5 workstation, and the LM6-8 workstation were already installed in the OAB. The LRUs and the MSAs designs were also complete. The MSA NOID End-Effector design had to work with the assembly equipment and LRU designs that were already in place.

  19. Mouse and human FcR effector functions.

    PubMed

    Bruhns, Pierre; Jönsson, Friederike

    2015-11-01

    Mouse and human FcRs have been a major focus of attention not only of the scientific community, through the cloning and characterization of novel receptors, and of the medical community, through the identification of polymorphisms and linkage to disease but also of the pharmaceutical community, through the identification of FcRs as targets for therapy or engineering of Fc domains for the generation of enhanced therapeutic antibodies. The availability of knockout mouse lines for every single mouse FcR, of multiple or cell-specific-'à la carte'-FcR knockouts and the increasing generation of hFcR transgenics enable powerful in vivo approaches for the study of mouse and human FcR biology. This review will present the landscape of the current FcR family, their effector functions and the in vivo models at hand to study them. These in vivo models were recently instrumental in re-defining the properties and effector functions of FcRs that had been overlooked or discarded from previous analyses. A particular focus will be made on the (mis)concepts on the role of high-affinity IgG receptors in vivo and on results from antibody engineering to enhance or abrogate antibody effector functions mediated by FcRs. PMID:26497511

  20. B cells as effectors and regulators of autoimmunity.

    PubMed

    Mariño, Eliana; Grey, Shane T

    2012-08-01

    A classic understanding of the interplay between B and T cell components of the immune system that drive autoimmunity, where B cells provide an effector function, is represented by systemic lupus erythematosus (SLE), an autoimmune condition characterised by the production of auto-antibodies. In SLE, CD4+T cells provide cognate help to self-reactive B cells, which in turn produce pathogenic auto-antibodies (1). Thus, B cells act as effectors by producing auto-antibody aided by T cell help such that B and T cell interactions are unidirectional. However, this paradigm of B and T cell interactions is challenged by new clinical data demonstrating that B cell depletion is effective for T cell mediated autoimmune diseases including type I diabetes mellitus (T1D) (2), rheumatoid arthritis (3), and multiple sclerosis (4). These clinical data indicate a model whereby B cells can influence the developing autoimmune T cell response, and therefore act as effectors, in ways that extend beyond the production of autoantibody (5). In this review by largely focusing on type I diabetes we will develop a hypothesis that bi-directional B and T interactions control the course of autoimmunity. PMID:22432804

  1. 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. PMID:25865799

  2. ANTIGEN SPECIFIC ANTIBODY COATED EXOSOME-LIKE NANOVESICLES DELIVER SUPPRESSOR T CELL miRNA-150 TO EFFECTOR T CELLS IN CONTACT SENSITIVITY

    PubMed Central

    Bryniarski, Krzysztof; Ptak, Wlodzimierz; Jayakumar, Asha; Püllmann, Kerstin; Caplan, Michael J.; Chairoungdua, Arthit; Lu, Jun; Adams, Brian; Sikora, Emilia; Nazimek, Katarzyna; Marquez, Susanna; Kleinstein, Steven H.; Sangwung, Panjamaporn; Iwakiri, Yasuko; Delgato, Eric; Redegeld, Frank; Blokhuis, Bart R.; Wojcikowski, Jacek; Daniel, Anna Wladyslawa; Kormelink, Tom Groot; Askenase, Philip W.

    2014-01-01

    Background T cell tolerance of allergic cutaneous contact sensitivity (CS) induced in mice by high doses of reactive hapten is mediated by suppressor cells that release antigen-specific suppressive nanovesicles. Objective To determine the mechanism(s) of immune suppression mediated by the nanovesicles. Methods T cell tolerance was induced by i.v. injections of hapten conjugated to self antigens of syngeneic erythrocytes and subsequent contact immunization with the same hapten. Lymph node and spleen cells from tolerized or control donors were harvested and cultured to produce a supernatant containing suppressive nanovesicles that were isolated for testing in active and adoptive cell transfer models of CS. Results Tolerance was shown due to exosome-like nanovesicles in the supernatant of CD8+ suppressor T cells that were not Treg. Antigen specificity of the suppressive nanovesicles was conferred by a surface coat of antibody light chains, or possibly whole antibody, allowing targeted delivery of selected inhibitory miRNA-150 to CS effector T cells. Nanovesicles also inhibited CS in actively sensitized mice after systemic injection at the peak of the responses. The role of antibody and miRNA-150 was established by tolerizing either panimmunoglobulin deficient JH-/- or miRNA-150-/- mice that produced non-suppressive nanovesicles. These nanovesicles could be made suppressive by adding antigen-specific antibody light chains or miRNA-150, respectively. Conclusions This is the first example of T cell regulation via systemic transit of exosome-like nanovesicles delivering a chosen inhibitory miRNA to target effector T cells in an antigen-specific manner by a surface coating of antibody light chains. PMID:23727037

  3. Herbivore exploits orally secreted bacteria to suppress plant defenses.

    PubMed

    Chung, Seung Ho; Rosa, Cristina; Scully, Erin D; Peiffer, Michelle; Tooker, John F; Hoover, Kelli; Luthe, Dawn S; Felton, Gary W

    2013-09-24

    Induced plant defenses in response to herbivore attack are modulated by cross-talk between jasmonic acid (JA)- and salicylic acid (SA)-signaling pathways. Oral secretions from some insect herbivores contain effectors that overcome these antiherbivore defenses. Herbivores possess diverse microbes in their digestive systems and these microbial symbionts can modify plant-insect interactions; however, the specific role of herbivore-associated microbes in manipulating plant defenses remains unclear. Here, we demonstrate that Colorado potato beetle (Leptinotarsa decemlineata) larvae exploit bacteria in their oral secretions to suppress antiherbivore defenses in tomato (Solanum lycopersicum). We found that antibiotic-untreated larvae decreased production of JA and JA-responsive antiherbivore defenses, but increased SA accumulation and SA-responsive gene expression. Beetles benefit from down-regulating plant defenses by exhibiting enhanced larval growth. In SA-deficient plants, suppression was not observed, indicating that suppression of JA-regulated defenses depends on the SA-signaling pathway. Applying bacteria isolated from larval oral secretions to wounded plants confirmed that three microbial symbionts belonging to the genera Stenotrophomonas, Pseudomonas, and Enterobacter are responsible for defense suppression. Additionally, reinoculation of these bacteria to antibiotic-treated larvae restored their ability to suppress defenses. Flagellin isolated from Pseudomonas sp. was associated with defense suppression. Our findings show that the herbivore exploits symbiotic bacteria as a decoy to deceive plants into incorrectly perceiving the threat as microbial. By interfering with the normal perception of herbivory, beetles can evade antiherbivore defenses of its host. PMID:24019469

  4. Herbivore exploits orally secreted bacteria to suppress plant defenses

    PubMed Central

    Chung, Seung Ho; Rosa, Cristina; Scully, Erin D.; Peiffer, Michelle; Tooker, John F.; Hoover, Kelli; Luthe, Dawn S.; Felton, Gary W.

    2013-01-01

    Induced plant defenses in response to herbivore attack are modulated by cross-talk between jasmonic acid (JA)- and salicylic acid (SA)-signaling pathways. Oral secretions from some insect herbivores contain effectors that overcome these antiherbivore defenses. Herbivores possess diverse microbes in their digestive systems and these microbial symbionts can modify plant–insect interactions; however, the specific role of herbivore-associated microbes in manipulating plant defenses remains unclear. Here, we demonstrate that Colorado potato beetle (Leptinotarsa decemlineata) larvae exploit bacteria in their oral secretions to suppress antiherbivore defenses in tomato (Solanum lycopersicum). We found that antibiotic-untreated larvae decreased production of JA and JA-responsive antiherbivore defenses, but increased SA accumulation and SA-responsive gene expression. Beetles benefit from down-regulating plant defenses by exhibiting enhanced larval growth. In SA-deficient plants, suppression was not observed, indicating that suppression of JA-regulated defenses depends on the SA-signaling pathway. Applying bacteria isolated from larval oral secretions to wounded plants confirmed that three microbial symbionts belonging to the genera Stenotrophomonas, Pseudomonas, and Enterobacter are responsible for defense suppression. Additionally, reinoculation of these bacteria to antibiotic-treated larvae restored their ability to suppress defenses. Flagellin isolated from Pseudomonas sp. was associated with defense suppression. Our findings show that the herbivore exploits symbiotic bacteria as a decoy to deceive plants into incorrectly perceiving the threat as microbial. By interfering with the normal perception of herbivory, beetles can evade antiherbivore defenses of its host. PMID:24019469

  5. Plant immunity directly or indirectly restricts the injection of type III effectors by the Pseudomonas syringae type III secretion system.

    PubMed

    Crabill, Emerson; Joe, Anna; Block, Anna; van Rooyen, Jennifer M; Alfano, James R

    2010-09-01

    Plants perceive microorganisms by recognizing microbial molecules known as pathogen-associated molecular patterns (PAMPs) inducing PAMP-triggered immunity (PTI) or by recognizing pathogen effectors inducing effector-triggered immunity (ETI). The hypersensitive response (HR), a programmed cell death response associated with ETI, is known to be inhibited by PTI. Here, we show that PTI-induced HR inhibition is due to direct or indirect restriction of the type III protein secretion system's (T3SS) ability to inject type III effectors (T3Es). We found that the Pseudomonas syringae T3SS was restricted in its ability to inject a T3E-adenylate cyclase (CyaA) injection reporter into PTI-induced tobacco (Nicotiana tabacum) cells. We confirmed this restriction with a direct injection assay that monitored the in planta processing of the AvrRpt2 T3E. Virulent P. syringae strains were able to overcome a PAMP pretreatment in tobacco or Arabidopsis (Arabidopsis thaliana) and continue to inject a T3E-CyaA reporter into host cells. In contrast, ETI-inducing P. syringae strains were unable to overcome PTI-induced injection restriction. A P. syringae pv tomato DC3000 mutant lacking about one-third of its T3E inventory was less capable of injecting into PTI-induced Arabidopsis plant cells, grew poorly in planta, and did not cause disease symptoms. PTI-induced transgenic Arabidopsis expressing the T3E HopAO1 or HopF2 allowed higher amounts of the T3E-CyaA reporter to be injected into plant cells compared to wild-type plants. Our results show that PTI-induced HR inhibition is due to direct or indirect restriction of T3E injection and that T3Es can relieve this restriction by suppressing PTI. PMID:20624999

  6. Influence of serum and soluble CD25 (sCD25) on regulatory and effector T cell function in Hepatocellular Carcinoma

    PubMed Central

    Cabrera, Roniel; Ararat, Miguel; Eksioglu, Erika A; Cao, Mengde; Xu, Yiling; Wasserfall, Clive; Atkinson, Mark A; Liu, Chen; Nelson, David R.

    2010-01-01

    Background Our previous studies showed that high levels of soluble CD25 (sCD25) in the serum of patients with hepatocellular carcinoma (HCC) correlated with blunted effector T cells (Teff) responses, tumor burden and poor survival. Understanding the interactions between Teff, CD4+CD25+ regulatory T cells (Treg) and soluble factors can identify novel therapeutic targets. Aims In this study, we characterize the mechanisms by which HCC serum and sCD25 mediate suppression of Teff. We also evaluate the effect of sCD25 on the suppression assays with normal healthy control cells (NHC) at a 1:1 Treg to Teff cell ratio to determine if sCD25 has any impact on Treg suppression. Results HCC serum and sCD25 suppressed Teff proliferation and down regulated CD25 expression on HCC Teffs in a dose dependent fashion with sCD25 doses above 3,000 pg/ml. Tregs from HCC and cirrhosis patients suppressed proliferation of target CD4+CD25? Teff in serum free medium (SFM). HCC Tregs showed a higher degree of suppression than cirrhosis derived Tregs. In contrast, Tregs from NHC did not suppress target Teff in SFM. However, isolated Tregs from all three study subjects (HCC, cirrhosis, NHC) suppressed CD4+CD25? Teff in serum conditions or in the presence of sCD25 in the range 6,000–12,000 pg/mL. Conclusion Down regulation of CD25 cell surface expression on Teffs is part of the overall suppressive mechanism of sCD25 and HCC serum on Teff responses. The observed sCD25 and HCC serum mediated suppression is further influenced via novel immune-inhibitory interaction between CD4+CD25+ Tregs and sCD25. PMID:20883314

  7. Influence of serum and soluble CD25 (sCD25) on regulatory and effector T-cell function in hepatocellular carcinoma.

    PubMed

    Cabrera, R; Ararat, M; Eksioglu, E A; Cao, M; Xu, Y; Wasserfall, C; Atkinson, M A; Liu, C; Nelson, D R

    2010-10-01

    Our previous studies showed that high levels of soluble CD25 (sCD25) in the serum of patients with hepatocellular carcinoma (HCC) correlated with blunted effector T-cells (Teff) responses, tumour burden and poor survival. Understanding the interactions between Teff, CD4+CD25+ regulatory T cells (Treg) and soluble factors can identify novel therapeutic targets. In this study, we characterize the mechanisms by which HCC serum and sCD25 mediate suppression of Teff and evaluate the effect of sCD25 on the suppression assays with normal healthy control cells (NHC) at a 1:1 Treg to Teff cell ratio to determine whether sCD25 has any impact on Treg suppression. HCC serum and sCD25 suppressed Teff proliferation and downregulated CD25 expression on HCC Teff in a dose-dependent fashion with sCD25 doses above 3000 pg/ml. Treg from HCC and cirrhosis patients suppressed proliferation of target CD4+CD25- Teff in serum-free medium (SFM). HCC Treg showed a higher degree of suppression than cirrhosis-derived Treg. In contrast, Treg from NHC did not suppress target Teff in SFM. However, isolated Treg from all three study subjects (HCC, cirrhosis and NHC) suppressed CD4+CD25- Teff in serum conditions or in the presence of sCD25 in the range 6000-12,000 pg/ml. In conclusion, downregulation of CD25 cell surface expression on Teff is part of the overall suppressive mechanism of sCD25 and HCC serum on Teff responses. The observed sCD25 and HCC serum-mediated suppression is further influenced via novel immune-inhibitory interaction between CD4+CD25+ Treg and sCD25. PMID:20883314

  8. Deletions in the repertoire of Pseudomonas syringae pv. tomato DC3000 type III secretion effector genes reveal functional overlap among effectors

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Many bacterial pathogens of plants and animals disarm and remodel host cells by injecting large repertoires of effectors via the type III secretion system (T3SS). The repertoires of individual strains appear to function as robust systems that can tolerate loss of individual effectors with little or ...

  9. A survey of the Pseudomonas syringae pv. tomato DC3000 type III secretion system effector repertoire reveals several effectors that are deleterious when expressed in Saccharomyces cerevisiae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The injection of nearly 30 effector proteins by the type III secretion system underlies the ability of Pseudomonas syringae pv. tomato strain DC3000 to cause disease in tomato and other host plants. The search for effector functions is complicated by redundancy within the repertoire and by plant R-g...

  10. Suppression of polymorphonuclear (PMN) and monocyte-mediated inhibition of Candida albicans growth by delta-9-tetrahydrocannabinol

    SciTech Connect

    Djeu, J.Y.; Parapanios, A.; Halkias, D.; Friedman, H.

    1986-03-05

    This study was an in vitro attempt to identify the effector cells responsible for growth inhibition of the opportunistic fungus, candida albicans, and to determine if THC or another marijuana derivatives, 11-hydroxyTHC, would adversely affect their function. Using a 24h radiolabel assay, the authors found that growth inhibition of C. albicans was primarily mediated by PMN and monocytes that could be isolated normal human peripheral blood. Both effector cell types caused almost complete inhibition of Candida growth at effector/target ratio of 300/1 and inhibition was often still seen at 30/1-. Incubation of PMN, PBL, or monocytes for 1 hr at 37C with THC or 11-hydroxyTHC caused a marked suppression of function in all 3 cell populations. Maximal suppression was obtained with 7.5-10..mu..g/ml of the drugs in medium containing 10% fetal bovine serum (FBS) or with 2-4..mu..g/ml in 1% FBS. These drug concentrations did not affect lymphoid cell viability or candida growth in the absence of lymphoid effector cells. Marijuana derivatives, therefore, are doubly dangerous in that opportunistic fungi such as C. albicans can grow in their presence while the effector cells that control fungal growth are readily inactivated.

  11. Pseudomonas syringae pv. tomato DC3000 Type III Secretion Effector Polymutants Reveal an Interplay between HopAD1 and AvrPtoB.

    PubMed

    Wei, Hai-Lei; Chakravarthy, Suma; Mathieu, Johannes; Helmann, Tyler C; Stodghill, Paul; Swingle, Bryan; Martin, Gregory B; Collmer, Alan

    2015-06-10

    The bacterial pathogen Pseudomonas syringae pv. tomato DC3000 suppresses the two-tiered plant innate immune system by injecting a complex repertoire of type III secretion effector (T3E) proteins. Beyond redundancy and interplay, individual T3Es may interact with multiple immunity-associated proteins, rendering their analysis challenging. We constructed a Pst DC3000 polymutant lacking all 36 T3Es and restored individual T3Es or their mutants to explore the interplay among T3Es. The weakly expressed T3E HopAD1 was sufficient to elicit immunity-associated cell death in Nicotiana benthamiana. HopAD1-induced cell death was suppressed partially by native AvrPtoB and completely by AvrPtoBM3, which has mutations disrupting its E3 ubiquitin ligase domain and two known domains for interacting with immunity-associated kinases. AvrPtoBM3 also gained the ability to interact with the immunity-kinase MKK2, which is required for HopAD1-dependent cell death. Thus, AvrPtoB has alternative, competing mechanisms for suppressing effector-triggered plant immunity. This approach allows the deconvolution of individual T3E activities. PMID:26067603

  12. Identification and Characterisation CRN Effectors in Phytophthora capsici Shows Modularity and Functional Diversity

    PubMed Central

    Stam, Remco; Jupe, Julietta; Howden, Andrew J. M.; Morris, Jenny A.; Boevink, Petra C.; Hedley, Pete E.; Huitema, Edgar

    2013-01-01

    Phytophthora species secrete a large array of effectors during infection of their host plants. The Crinkler (CRN) gene family encodes a ubiquitous but understudied class of effectors with possible but as of yet unknown roles in infection. To appreciate CRN effector function in Phytophthora, we devised a simple Crn gene identification and annotation pipeline to improve effector prediction rates. We predicted 84 full-length CRN coding genes and assessed CRN effector domain diversity in sequenced Oomycete genomes. These analyses revealed evidence of CRN domain innovation in Phytophthora and expansion in the Peronosporales. We performed gene expression analyses to validate and define two classes of CRN effectors, each possibly contributing to infection at different stages. CRN localisation studies revealed that P. capsici CRN effector domains target the nucleus and accumulate in specific sub-nuclear compartments. Phenotypic analyses showed that few CRN domains induce necrosis when expressed in planta and that one cell death inducing effector, enhances P. capsici virulence on Nicotiana benthamiana. These results suggest that the CRN protein family form an important class of intracellular effectors that target the host nucleus during infection. These results combined with domain expansion in hemi-biotrophic and necrotrophic pathogens, suggests specific contributions to pathogen lifestyles. This work will bolster CRN identification efforts in other sequenced oomycete species and set the stage for future functional studies towards understanding CRN effector functions. PMID:23536880

  13. Flavonols modulate the effector functions of healthy individuals' immune complex-stimulated neutrophils: a therapeutic perspective for rheumatoid arthritis.

    PubMed

    Santos, Everton O L; Kabeya, Luciana M; Figueiredo-Rinhel, Andréa S G; Marchi, Larissa F; Andrade, Micássio F; Piatesi, Fabiana; Paoliello-Paschoalato, Adriana B; Azzolini, Ana Elisa C S; Lucisano-Valim, Yara M

    2014-07-01

    Rheumatoid arthritis (RA) patients usually exhibit immune complex (IC) deposition and increased neutrophil activation in the joint. In this study, we assessed how four flavonols (galangin, kaempferol, quercetin, and myricetin) modulate the effector functions of healthy individuals' and active RA patients' IC-stimulated neutrophils. We measured superoxide anion and total reactive oxygen species production using lucigenin (CL-luc)- and luminol (CL-lum)-enhanced chemiluminescence assays, respectively. Galangin, kaempferol, and quercetin inhibited CL-lum to the same degree (mean IC50=2.5 ?M). At 2.5 ?M, quercetin and galangin suppressed nearly 65% CL-lum of active RA patients' neutrophils. Quercetin inhibited CL-luc the most effectively (IC50=1.71±0.36 ?M). The four flavonols diminished myeloperoxidase activity, but they did not decrease NADPH oxidase activity, phagocytosis, microbial killing, or cell viability of neutrophils. The ability of the flavonols to scavenge hypochlorous acid and chloramines, but not H2O2, depended on the hydroxylation degree of the flavonol B-ring. Therefore, at physiologically relevant concentrations, the flavonols partially inhibited the oxidative metabolism of IC-stimulated neutrophils without affecting the other investigated effector functions. Using these compounds to modulate IC-mediated neutrophil activation is a promising safe therapeutic strategy to control inflammation in active RA patients. PMID:24797916

  14. Fire Suppression and Response

    NASA Technical Reports Server (NTRS)

    Ruff, Gary A.

    2004-01-01

    This report is concerned with the following topics regarding fire suppression:What is the relative effectiveness of candidate suppressants to extinguish a representative fire in reduced gravity, including high-O2 mole fraction, low -pressure environments? What are the relative advantages and disadvantages of physically acting and chemically-acting agents in spacecraft fire suppression? What are the O2 mole fraction and absolute pressure below which a fire cannot exist? What effect does gas-phase radiation play in the overall fire and post-fire environments? Are the candidate suppressants effective to extinguish fires on practical solid fuels? What is required to suppress non-flaming fires (smoldering and deep seated fires) in reduced gravity? How can idealized space experiment results be applied to a practical fire scenario? What is the optimal agent deployment strategy for space fire suppression?

  15. Single-cell quantification of IL-2 response by effector and regulatory T cells reveals critical plasticity in immune response

    PubMed Central

    Feinerman, Ofer; Jentsch, Garrit; Tkach, Karen E; Coward, Jesse W; Hathorn, Matthew M; Sneddon, Michael W; Emonet, Thierry; Smith, Kendall A; Altan-Bonnet, Grégoire

    2010-01-01

    Understanding how the immune system decides between tolerance and activation by antigens requires addressing cytokine regulation as a highly dynamic process. We quantified the dynamics of interleukin-2 (IL-2) signaling in a population of T cells during an immune response by combining in silico modeling and single-cell measurements in vitro. We demonstrate that IL-2 receptor expression levels vary widely among T cells creating a large variability in the ability of the individual cells to consume, produce and participate in IL-2 signaling within the population. Our model reveals that at the population level, these heterogeneous cells are engaged in a tug-of-war for IL-2 between regulatory (Treg) and effector (Teff) T cells, whereby access to IL-2 can either increase the survival of Teff cells or the suppressive capacity of Treg cells. This tug-of-war is the mechanism enforcing, at the systems level, a core function of Treg cells, namely the specific suppression of survival signals for weakly activated Teff cells but not for strongly activated cells. Our integrated model yields quantitative, experimentally validated predictions for the manipulation of Treg suppression. PMID:21119631

  16. Investigation of a bio-inspired lift-enhancing effector on a 2D airfoil.

    PubMed

    Johnston, Joe; Gopalarathnam, Ashok

    2012-09-01

    A flap mounted on the upper surface of an airfoil, called a 'lift-enhancing effector', has been shown in wind tunnel tests to have a similar function to a bird's covert feathers, which rise off the wing's surface in response to separated flows. The effector, fabricated from a thin Mylar sheet, is allowed to rotate freely about its leading edge. The tests were performed in the NCSU subsonic wind tunnel at a chord Reynolds number of 4 × 10(5). The maximum lift coefficient with the effector was the same as that for the clean airfoil, but was maintained over an angle-of-attack range from 12° to almost 20°, resulting in a very gentle stall behavior. To better understand the aerodynamics and to estimate the deployment angle of the free-moving effector, fixed-angle effectors fabricated out of stiff wood were also tested. A progressive increase in the stall angle of attack with increasing effector angle was observed, with diminishing returns beyond the effector angle of 60°. Drag tests on both the free-moving and fixed effectors showed a marked improvement in drag at high angles of attack. Oil flow visualization on the airfoil with and without the fixed-angle effectors proved that the effector causes the separation point to move aft on the airfoil, as compared to the clean airfoil. This is thought to be the main mechanism by which an effector improves both lift and drag. A comparison of the fixed-effector results with those from the free-effector tests shows that the free effector's deployment angle is between 30° and 45°. When operating at and beyond the clean airfoil's stall angle, the free effector automatically deploys to progressively higher angles with increasing angles of attack. This slows down the rapid upstream movement of the separation point and avoids the severe reduction in the lift coefficient and an increase in the drag coefficient that are seen on the clean airfoil at the onset of stall. Thus, the effector postpones the stall by 4-8° and makes the stall behavior more gentle. The benefits of using the effector could include care-free operations at high angles of attack during perching and maneuvering flight, especially in gusty conditions. PMID:22498691

  17. Transgenic Plants That Express the Phytoplasma Effector SAP11 Show Altered Phosphate Starvation and Defense Responses1[W][OPEN

    PubMed Central

    Lu, Yen-Ting; Li, Meng-Ying; Cheng, Kai-Tan; Tan, Choon Meng; Su, Li-Wen; Lin, Wei-Yi; Shih, Hsien-Tzung; Chiou, Tzyy-Jen; Yang, Jun-Yi

    2014-01-01

    Phytoplasmas have the smallest genome among bacteria and lack many essential genes required for biosynthetic and metabolic functions, making them unculturable, phloem-limited plant pathogens. In this study, we observed that transgenic Arabidopsis (Arabidopsis thaliana) expressing the secreted Aster Yellows phytoplasma strain Witches’ Broom protein11 shows an altered root architecture, similarly to the disease symptoms of phytoplasma-infected plants, by forming hairy roots. This morphological change is paralleled by an accumulation of cellular phosphate (Pi) and an increase in the expression levels of Pi starvation-induced genes and microRNAs. In addition to the Pi starvation responses, we found that secreted Aster Yellows phytoplasma strain Witches’ Broom protein11 suppresses salicylic acid-mediated defense responses and enhances the growth of a bacterial pathogen. These results contribute to an improved understanding of the role of phytoplasma effector SAP11 and provide new insights for understanding the molecular basis of plant-pathogen interactions. PMID:24464367

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

    PubMed

    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

  19. PKC-Theta in Regulatory and Effector T-cell Functions.

    PubMed

    Brezar, Vedran; Tu, Wen Juan; Seddiki, Nabila

    2015-01-01

    One of the major goals in immunology research is to understand the regulatory mechanisms that underpin the rapid switch on/off of robust and efficient effector (Teffs) or regulatory (Tregs) T-cell responses. Understanding the molecular mechanisms underlying the regulation of such responses is critical for the development of effective therapies. T-cell activation involves the engagement of T-cell receptor and co-stimulatory signals, but the subsequent recruitment of serine/threonine-specific protein Kinase C-theta (PKC-?) to the immunological synapse (IS) is instrumental for the formation of signaling complexes, which ultimately lead to a transcriptional network in T cells. Recent studies demonstrated that major differences between Teffs and Tregs occurred at the IS where its formation induces altered signaling pathways in Tregs. These pathways are characterized by reduced recruitment of PKC-?, suggesting that PKC-? inhibits Tregs suppressive function in a negative feedback loop. As the balance of Teffs and Tregs has been shown to be central in several diseases, it was not surprising that some studies revealed that PKC-? plays a major role in the regulation of this balance. This review will examine recent knowledge on the role of PKC-? in T-cell transcriptional responses and how this protein can impact on the function of both Tregs and Teffs. PMID:26528291

  20. PKC-Theta in Regulatory and Effector T-cell Functions

    PubMed Central

    Brezar, Vedran; Tu, Wen Juan; Seddiki, Nabila

    2015-01-01

    One of the major goals in immunology research is to understand the regulatory mechanisms that underpin the rapid switch on/off of robust and efficient effector (Teffs) or regulatory (Tregs) T-cell responses. Understanding the molecular mechanisms underlying the regulation of such responses is critical for the development of effective therapies. T-cell activation involves the engagement of T-cell receptor and co-stimulatory signals, but the subsequent recruitment of serine/threonine-specific protein Kinase C-theta (PKC-?) to the immunological synapse (IS) is instrumental for the formation of signaling complexes, which ultimately lead to a transcriptional network in T cells. Recent studies demonstrated that major differences between Teffs and Tregs occurred at the IS where its formation induces altered signaling pathways in Tregs. These pathways are characterized by reduced recruitment of PKC-?, suggesting that PKC-? inhibits Tregs suppressive function in a negative feedback loop. As the balance of Teffs and Tregs has been shown to be central in several diseases, it was not surprising that some studies revealed that PKC-? plays a major role in the regulation of this balance. This review will examine recent knowledge on the role of PKC-? in T-cell transcriptional responses and how this protein can impact on the function of both Tregs and Teffs. PMID:26528291

  1. Effector candidates in the secretome of Piriformospora indica, a ubiquitous plant-associated fungus

    PubMed Central

    Rafiqi, Maryam; Jelonek, Lukas; Akum, Ndifor F.; Zhang, Feng; Kogel, Karl-Heinz

    2013-01-01

    One of the emerging systems in plant–microbe interaction is the study of proteins, referred to as effectors, secreted by microbes in order to modulate host cells function and structure and to promote microbial growth on plant tissue. Current knowledge on fungal effectors derives mainly from biotrophic and hemibiotrophic plant fungal pathogens that have a limited host range. Here, we focus on effectors of Piriformospora indica, a soil borne endophyte forming intimate associations with roots of a wide range of plant species. Complete genome sequencing provides an opportunity to investigate the role of effectors during the interaction of this mutualistic fungus with plants. We describe in silico analyses to predict effectors of P. indica and we explore effector features considered here to mine a high priority protein list for functional analysis. PMID:23874344

  2. Effector candidates in the secretome of Piriformospora indica, a ubiquitous plant-associated fungus.

    PubMed

    Rafiqi, Maryam; Jelonek, Lukas; Akum, Ndifor F; Zhang, Feng; Kogel, Karl-Heinz

    2013-01-01

    One of the emerging systems in plant-microbe interaction is the study of proteins, referred to as effectors, secreted by microbes in order to modulate host cells function and structure and to promote microbial growth on plant tissue. Current knowledge on fungal effectors derives mainly from biotrophic and hemibiotrophic plant fungal pathogens that have a limited host range. Here, we focus on effectors of Piriformospora indica, a soil borne endophyte forming intimate associations with roots of a wide range of plant species. Complete genome sequencing provides an opportunity to investigate the role of effectors during the interaction of this mutualistic fungus with plants. We describe in silico analyses to predict effectors of P. indica and we explore effector features considered here to mine a high priority protein list for functional analysis. PMID:23874344

  3. Induction and suppression of PEN3 focal accumulation during Pseudomonas syringae pv. tomato DC3000 infection of Arabidopsis.

    PubMed

    Xin, Xiu-Fang; Nomura, Kinya; Underwood, William; He, Sheng Yang

    2013-08-01

    The pleiotropic drug resistance (PDR) proteins belong to the super-family of ATP-binding cassette (ABC) transporters. AtPDR8, also called PEN3, is required for penetration resistance of Arabidopsis to nonadapted powdery mildew fungi. During fungal infection, plasma-membrane-localized PEN3 is concentrated at fungal entry sites, as part of the plant's focal immune response. Here, we show that the pen3 mutant is compromised in resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000. P. syringae pv. tomato DC3000 infection or treatment with a flagellin-derived peptide, flg22, induced strong focal accumulation of PEN3-green fluorescent protein. Interestingly, after an initial induction of PEN3 accumulation, P. syringae pv. tomato DC3000 but not the type-III-secretion-deficient mutant hrcC could suppress PEN3 accumulation. Moreover, transgenic overexpression of the P. syringae pv. tomato DC3000 effector AvrPto was sufficient to suppress PEN3 focal accumulation in response to flg22. Analyses of P. syringae pv. tomato DC3000 effector deletion mutants showed that individual effectors, including AvrPto, appear to be insufficient to suppress PEN3 accumulation when delivered by bacteria, suggesting a requirement for a combined action of multiple effectors. Collectively, our results indicate that PEN3 plays a positive role in plant resistance to a bacterial pathogen and show that focal accumulation of PEN3 protein may be a useful cellular response marker for the Arabidopsis-P. syringae interaction. PMID:23815470

  4. Perturbation of Maize Phenylpropanoid Metabolism by an AvrE Family Type III Effector from Pantoea stewartii1[OPEN

    PubMed Central

    Asselin, Jo Ann E.; Lin, Jinshan; Perez-Quintero, Alvaro L.; Gentzel, Irene; Majerczak, Doris; Opiyo, Stephen O.; Zhao, Wanying; Paek, Seung-Mann; Kim, Min Gab; Coplin, David L.; Blakeslee, Joshua J.; Mackey, David

    2015-01-01

    AvrE family type III effector proteins share the ability to suppress host defenses, induce disease-associated cell death, and promote bacterial growth. However, despite widespread contributions to numerous bacterial diseases in agriculturally important plants, the mode of action of these effectors remains largely unknown. WtsE is an AvrE family member required for the ability of Pantoea stewartii ssp. stewartii (Pnss) to proliferate efficiently and cause wilt and leaf blight symptoms in maize (Zea mays) plants. Notably, when WtsE is delivered by a heterologous system into the leaf cells of susceptible maize seedlings, it alone produces water-soaked disease symptoms reminiscent of those produced by Pnss. Thus, WtsE is a pathogenicity and virulence factor in maize, and an Escherichia coli heterologous delivery system can be used to study the activity of WtsE in isolation from other factors produced by Pnss. Transcriptional profiling of maize revealed the effects of WtsE, including induction of genes involved in secondary metabolism and suppression of genes involved in photosynthesis. Targeted metabolite quantification revealed that WtsE perturbs maize metabolism, including the induction of coumaroyl tyramine. The ability of mutant WtsE derivatives to elicit transcriptional and metabolic changes in susceptible maize seedlings correlated with their ability to promote disease. Furthermore, chemical inhibitors that block metabolic flux into the phenylpropanoid pathways targeted by WtsE also disrupted the pathogenicity and virulence activity of WtsE. While numerous metabolites produced downstream of the shikimate pathway are known to promote plant defense, our results indicate that misregulated induction of phenylpropanoid metabolism also can be used to promote pathogen virulence. PMID:25635112

  5. Leptin's effect on hyperactivity: potential downstream effector mechanisms.

    PubMed

    Hillebrand, J J G; Kas, M J H; van Elburg, A A; Hoek, H W; Adan, R A H

    2008-08-01

    Up to 80% of patients with Anorexia Nervosa (AN) demonstrate hyperactivity. Hyperactivity counteracts weight gain during treatment and is associated with poor outcome of the disease. We hypothesized that hyperactivity in AN patients has a neurobiological basis and used an animal model-based translational approach to gain insight in mechanisms underlying this hyperactivity. Previously we and others showed that leptin treatment attenuates hyperactivity in the rat activity-based anorexia (ABA) model. The mechanisms involved in this process are, however, unknown. Here we describe potential downstream effector mechanisms involved in the attenuation of hyperactivity by leptin treatment in ABA rats. PMID:18495181

  6. Transcription Activator-like Effectors: A Toolkit for Synthetic Biology

    PubMed Central

    2015-01-01

    Transcription activator-like effectors (TALEs) are proteins secreted by Xanthomonas bacteria to aid the infection of plant species. TALEs assist infections by binding to specific DNA sequences and activating the expression of host genes. Recent results show that TALE proteins consist of a central repeat domain, which determines the DNA targeting specificity and can be rapidly synthesized de novo. Considering the highly modular nature of TALEs, their versatility, and the ease of constructing these proteins, this technology can have important implications for synthetic biology applications. Here, we review developments in the area with a particular focus on modifications for custom and controllable gene regulation. PMID:24933470

  7. Functions and requirements for the INEL light duty utility arm gripper end effector

    SciTech Connect

    Pace, D.P.; Barnes, G.E.

    1995-02-01

    This gripper end effector system functions and requirements document defines the system functions that the end effector must perform as well as the requirements the design must meet. Safety, quality assurance, operations, environmental conditions, and regulatory requirements have been considered. The main purpose of this document is to provide a basis for the end effector engineering, design, and fabrication activities. The document shall be the living reference document to initiate the development activities and will be updated as system technologies are finalized.

  8. The Exo70 Subunit of the Exocyst Is an Effector for Both Cdc42 and Rho3 Function in Polarized Exocytosis

    PubMed Central

    Wu, Hao; Turner, Courtney; Gardner, Jimmy; Temple, Brenda

    2010-01-01

    The Rho3 and Cdc42 members of the Rho GTPase family are important regulators of exocytosis in yeast. However, the precise mechanism by which they regulate this process is controversial. Here, we present evidence that the Exo70 component of the exocyst complex is a direct effector of both Rho3 and Cdc42. We identify gain-of-function mutants in EXO70 that potently suppress mutants in RHO3 and CDC42 defective for exocytic function. We show that Exo70 has the biochemical properties expected of a direct effector for both Rho3 and Cdc42. Surprisingly, we find that C-terminal prenylation of these GTPases both promotes the interaction and influences the sites of binding within Exo70. Finally, we demonstrate that the phenotypes associated with novel loss-of-function mutants in EXO70, are entirely consistent with Exo70 as an effector for both Rho3 and Cdc42 function in secretion. These data suggest that interaction with the Exo70 component of the exocyst is a key event in spatial regulation of exocytosis by Rho GTPases. PMID:19955214

  9. Immune activation mediated by the late blight resistance protein R1 requires nuclear localization of R1 and the effector AVR1.

    PubMed

    Du, Yu; Berg, Jeroen; Govers, Francine; Bouwmeester, Klaas

    2015-08-01

    Resistance against oomycete pathogens is mainly governed by intracellular nucleotide-binding leucine-rich repeat (NLR) receptors that recognize matching avirulence (AVR) proteins from the pathogen, RXLR effectors that are delivered inside host cells. Detailed molecular understanding of how and where NLR proteins and RXLR effectors interact is essential to inform the deployment of durable resistance (R) genes. Fluorescent tags, nuclear localization signals (NLSs) and nuclear export signals (NESs) were exploited to determine the subcellular localization of the potato late blight protein R1 and the Phytophthora infestans RXLR effector AVR1, and to target these proteins to the nucleus or cytoplasm. Microscopic imaging revealed that both R1 and AVR1 occurred in the nucleus and cytoplasm, and were in close proximity. Transient expression of NLS- or NES-tagged R1 and AVR1 in Nicotiana benthamiana showed that activation of the R1-mediated hypersensitive response and resistance required localization of the R1/AVR1 pair in the nucleus. However, AVR1-mediated suppression of cell death in the absence of R1 was dependent on localization of AVR1 in the cytoplasm. Balanced nucleocytoplasmic partitioning of AVR1 seems to be a prerequisite. Our results show that R1-mediated immunity is activated inside the nucleus with AVR1 in close proximity and suggest that nucleocytoplasmic transport of R1 and AVR1 is tightly regulated. PMID:25760731

  10. Metabolic plasticity of human T cells: Preserved cytokine production under glucose deprivation or mitochondrial restriction, but 2-deoxy-glucose affects effector functions.

    PubMed

    Renner, Kathrin; Geiselhöringer, Anna-Lena; Fante, Matthias; Bruss, Christina; Färber, Stephanie; Schönhammer, Gabriele; Peter, Katrin; Singer, Katrin; Andreesen, Reinhard; Hoffmann, Petra; Oefner, Peter; Herr, Wolfgang; Kreutz, Marina

    2015-09-01

    The strong link between T-cell metabolism and effector functions is well characterized in the murine system but hardly investigated in human T cells. Therefore, we analyzed glycolytic and mitochondrial activity in correlation to function in activated human CD4 and CD8 T cells. Glycolysis was barely detectable upon stimulation but accelerated beyond 24 h, whereas mitochondrial activity was elevated immediately in both T-cell populations. Glucose deprivation or mitochondrial restriction reduced proliferation, had only a transient impact on "on-blast formation" and no impact on viability, IFN-?, IL-2, IL-4, and IL-10 production, whereas TNF was reduced. Similar results were obtained in bulk T cells and T-cell subsets. Elevated respiration under glucose restriction demonstrated metabolic flexibility. Administration of the glycolytic inhibitor 2-deoxy-glucose suppressed both glycolysis and respiration and exerted a strong impact on cytokine production that persisted for IFN-? after removal of 2-deoxy-glucose. Taken together, glycolytic or mitochondrial restriction alone compromised proliferation of human T cells, but barely affected their effector functions. In contrast, effector functions were severely affected by 2-deoxy-glucose treatment. PMID:26114249

  11. A Plethora of Virulence Strategies Hidden Behind Nuclear Targeting of Microbial Effectors

    PubMed Central

    Rivas, Susana; Genin, Stéphane

    2011-01-01

    Plant immune responses depend on the ability to couple rapid recognition of the invading microbe to an efficient response. During evolution, plant pathogens have acquired the ability to deliver effector molecules inside host cells in order to manipulate cellular and molecular processes and establish pathogenicity. Following translocation into plant cells, microbial effectors may be addressed to different subcellular compartments. Intriguingly, a significant number of effector proteins from different pathogenic microorganisms, including viruses, oomycetes, fungi, nematodes, and bacteria, is targeted to the nucleus of host cells. In agreement with this observation, increasing evidence highlights the crucial role played by nuclear dynamics, and nucleocytoplasmic protein trafficking during a great variety of analyzed plant–pathogen interactions. Once in the nucleus, effector proteins are able to manipulate host transcription or directly subvert essential host components to promote virulence. Along these lines, it has been suggested that some effectors may affect histone packing and, thereby, chromatin configuration. In addition, microbial effectors may either directly activate transcription or target host transcription factors to alter their regular molecular functions. Alternatively, nuclear translocation of effectors may affect subcellular localization of their cognate resistance proteins in a process that is essential for resistance protein-mediated plant immunity. Here, we review recent progress in our field on the identification of microbial effectors that are targeted to the nucleus of host plant cells. In addition, we discuss different virulence strategies deployed by microbes, which have been uncovered through examination of the mechanisms that guide nuclear localization of effector proteins. PMID:22639625

  12. Candidate Effector Proteins of the Rust Pathogen Melampsora larici-populina Target Diverse Plant Cell Compartments.

    PubMed

    Petre, Benjamin; Saunders, Diane G O; Sklenar, Jan; Lorrain, Cécile; Win, Joe; Duplessis, Sébastien; Kamoun, Sophien

    2015-06-01

    Rust fungi are devastating crop pathogens that deliver effector proteins into infected tissues to modulate plant functions and promote parasitic growth. The genome of the poplar leaf rust fungus Melampsora larici-populina revealed a large catalog of secreted proteins, some of which have been considered candidate effectors. Unraveling how these proteins function in host cells is a key to understanding pathogenicity mechanisms and developing resistant plants. In this study, we used an effectoromics pipeline to select, clone, and express 20 candidate effectors in Nicotiana benthamiana leaf cells to determine their subcellular localization and identify the plant proteins they interact with. Confocal microscopy revealed that six candidate effectors target the nucleus, nucleoli, chloroplasts, mitochondria, and discrete cellular bodies. We also used coimmunoprecipitation (coIP) and mass spectrometry to identify 606 N. benthamiana proteins that associate with the candidate effectors. Five candidate effectors specifically associated with a small set of plant proteins that may represent biologically relevant interactors. We confirmed the interaction between the candidate effector MLP124017 and TOPLESS-related protein 4 from poplar by in planta coIP. Altogether, our data enable us to validate effector proteins from M. larici-populina and reveal that these proteins may target multiple compartments and processes in plant cells. It also shows that N. benthamiana can be a powerful heterologous system to study effectors of obligate biotrophic pathogens. PMID:25650830

  13. Deletions in the repertoire of Pseudomonas syringae pv. tomato DC3000 type III secretion effector genes reveal functional overlap among effectors.

    PubMed

    Kvitko, Brian H; Park, Duck Hwan; Velásquez, André C; Wei, Chia-Fong; Russell, Alistair B; Martin, Gregory B; Schneider, David J; Collmer, Alan

    2009-04-01

    The gamma-proteobacterial plant pathogen Pseudomonas syringae pv. tomato DC3000 uses the type III secretion system to inject ca. 28 Avr/Hop effector proteins into plants, which enables the bacterium to grow from low inoculum levels to produce bacterial speck symptoms in tomato, Arabidopsis thaliana, and (when lacking hopQ1-1) Nicotiana benthamiana. The effectors are collectively essential but individually dispensable for the ability of the bacteria to defeat defenses, grow, and produce symptoms in plants. Eighteen of the effector genes are clustered in six genomic islands/islets. Combinatorial deletions involving these clusters and two of the remaining effector genes revealed a redundancy-based structure in the effector repertoire, such that some deletions diminished growth in N. benthamiana only in combination with other deletions. Much of the ability of DC3000 to grow in N. benthamiana was found to be due to five effectors in two redundant-effector groups (REGs), which appear to separately target two high-level processes in plant defense: perception of external pathogen signals (AvrPto and AvrPtoB) and deployment of antimicrobial factors (AvrE, HopM1, HopR1). Further support for the membership of HopR1 in the same REG as AvrE was gained through bioinformatic analysis, revealing the existence of an AvrE/DspA/E/HopR effector superfamily, which has representatives in virtually all groups of proteobacterial plant pathogens that deploy type III effectors. PMID:19381254

  14. Regulation of Cell Wall-Bound Invertase in Pepper Leaves by Xanthomonas campestris pv. vesicatoria Type Three Effectors

    PubMed Central

    Sonnewald, Sophia; Priller, Johannes P. R.; Schuster, Julia; Glickmann, Eric; Hajirezaei, Mohammed-Reza; Siebig, Stefan; Mudgett, Mary Beth; Sonnewald, Uwe

    2012-01-01

    Xanthomonas campestris pv. vesicatoria (Xcv) possess a type 3 secretion system (T3SS) to deliver effector proteins into its Solanaceous host plants. These proteins are involved in suppression of plant defense and in reprogramming of plant metabolism to favour bacterial propagation. There is increasing evidence that hexoses contribute to defense responses. They act as substrates for metabolic processes and as metabolic semaphores to regulate gene expression. Especially an increase in the apoplastic hexose-to-sucrose ratio has been suggested to strengthen plant defense. This shift is brought about by the activity of cell wall-bound invertase (cw-Inv). We examined the possibility that Xcv may employ type 3 effector (T3E) proteins to suppress cw-Inv activity during infection. Indeed, pepper leaves infected with a T3SS-deficient Xcv strain showed a higher level of cw-Inv mRNA and enzyme activity relative to Xcv wild type infected leaves. Higher cw-Inv activity was paralleled by an increase in hexoses and mRNA abundance for the pathogenesis-related gene PRQ. These results suggest that Xcv suppresses cw-Inv activity in a T3SS-dependent manner, most likely to prevent sugar-mediated defense signals. To identify Xcv T3Es that regulate cw-Inv activity, a screen was performed with eighteen Xcv strains, each deficient in an individual T3E. Seven Xcv T3E deletion strains caused a significant change in cw-Inv activity compared to Xcv wild type. Among them, Xcv lacking the xopB gene (Xcv ?xopB) caused the most prominent increase in cw-Inv activity. Deletion of xopB increased the mRNA abundance of PRQ in Xcv ?xopB-infected pepper leaves, but not of Pti5 and Acre31, two PAMP-triggered immunity markers. Inducible expression of XopB in transgenic tobacco inhibited Xcv-mediated induction of cw-Inv activity observed in wild type plants and resulted in severe developmental phenotypes. Together, these data suggest that XopB interferes with cw-Inv activity in planta to suppress sugar-enhanced defense responses during Xcv infection. PMID:23272161

  15. Flight Control Using Distributed Shape-Change Effector Arrays

    NASA Technical Reports Server (NTRS)

    Raney, David L.; Montgomery, Raymond C.; Green, Lawrence I.; Park, Michael A.

    2000-01-01

    Recent discoveries in material science and fluidics have been used to create a variety of novel effector devices that offer great potential to enable new approaches to aerospace vehicle flight control. Examples include small inflatable blisters, shape-memory alloy diaphragms, and piezoelectric patches that may be used to produce distortions or bumps on the surface of an airfoil to generate control moments. Small jets have also been used to produce a virtual shape-change through fluidic means by creating a recirculation bubble on the surface of an airfoil. An advanced aerospace vehicle might use distributed arrays of hundreds of such devices to generate moments for stabilization and maneuver control, either augmenting or replacing conventional ailerons, flaps or rudders. This research demonstrates the design and use of shape-change device arrays for a tailless aircraft in a low-rate maneuvering application. A methodology for assessing the control authority of the device arrays is described, and a suite of arrays is used in a dynamic simulation to illustrate allocation and deployment methodologies. Although the authority of the preliminary shape-change array designs studied in this paper appeared quite low, the simulation results indicate that the effector suite possessed sufficient authority to stabilize and maneuver the vehicle in mild turbulence.

  16. Armet is an effector protein mediating aphid-plant interactions.

    PubMed

    Wang, Wei; Dai, Huaien; Zhang, Yi; Chandrasekar, Raman; Luo, Lan; Hiromasa, Yasuaki; Sheng, Changzhong; Peng, Gongxin; Chen, Shaoliang; Tomich, John M; Reese, John; Edwards, Owain; Kang, Le; Reeck, Gerald; Cui, Feng

    2015-05-01

    Aphid saliva is predicted to contain proteins that modulate plant defenses and facilitate feeding. Armet is a well-characterized bifunctional protein in mammalian systems. Here we report a new role of Armet, namely as an effector protein in the pea aphid, Acyrthosiphon pisum. Pea aphid Armet's physical and chemical properties and its intracellular role are comparable to those reported for mammalian Armets. Uniquely, we detected Armet in aphid watery saliva and in the phloem sap of fava beans fed on by aphids. Armet's transcript level is several times higher in the salivary gland when aphids feed on bean plants than when they feed on an artificial diet. Knockdown of the Armet transcript by RNA interference disturbs aphid feeding behavior on fava beans measured by the electrical penetration graph technique and leads to a shortened life span. Inoculation of pea aphid Armet protein into tobacco leaves induced a transcriptional response that included pathogen-responsive genes. The data suggest that Armet is an effector protein mediating aphid-plant interactions. PMID:25678626

  17. Local sensory control of a dexterous end effector

    NASA Technical Reports Server (NTRS)

    Pinto, Victor H.; Everett, Louis J.; Driels, Morris

    1990-01-01

    A numerical scheme was developed to solve the inverse kinematics for a user-defined manipulator. The scheme was based on a nonlinear least-squares technique which determines the joint variables by minimizing the difference between the target end effector pose and the actual end effector pose. The scheme was adapted to a dexterous hand in which the joints are either prismatic or revolute and the fingers are considered open kinematic chains. Feasible solutions were obtained using a three-fingered dexterous hand. An algorithm to estimate the position and orientation of a pre-grasped object was also developed. The algorithm was based on triangulation using an ideal sensor and a spherical object model. By choosing the object to be a sphere, only the position of the object frame was important. Based on these simplifications, a minimum of three sensors are needed to find the position of a sphere. A two dimensional example to determine the position of a circle coordinate frame using a two-fingered dexterous hand was presented.

  18. Interleukin 10 and transforming growth factor ? contribute to the development of experimentally induced allergic conjunctivitis in mice during the effector phase

    PubMed Central

    Fukushima, A; Sumi, T; Fukuda, K; Kumagai, N; Nishida, T; Yagita, H; Ueno, H

    2006-01-01

    Aim To investigate the involvement of interleukin (IL)10 and transforming growth factor (TGF) ? in the development of experimentally induced allergic conjunctivitis in mice. Methods Balb/c mice were actively sensitised with ragweed in alum, and then challenged with ragweed in eye drops after 10?days. 24?h later, the conjunctivas, spleens and blood were collected for histological and cytokine expression analyses, proliferation and cytokine production assays and measurement of immunoglobulin (Ig) levels. Mice developing experimentally induced allergic conjunctivitis were injected intraperitoneally with 200??g of anti?IL10 or anti?TGF ? antibodies at 0, 2, 4, 6 and 8?days (induction phase treatment) or 500??g of antibodies 2?h before ragweed challenge (effector phase treatment). Normal rat IgG was used for control injections. Results Treatment with either anti?IL10 or anti?TGF ? antibodies during the induction phase did not affect eosinophil infiltration into the conjunctiva. By contrast, treatment with either antibody during the effector phase suppressed infiltration. During the effector phase, treatment with anti?TGF ? antibody, but not the anti?IL10 antibody, markedly up regulated proliferation and Th2 cytokine production by splenocytes. IL1? levels in the conjunctiva were reduced after treatment with either antibody; in addition, eotaxin and tumour necrosis factor ? levels were reduced after treatment with antibody to TGF ?. Conclusions IL10 and TGF ? do not have immunosuppressive roles in the development of experimentally induced allergic conjunctivitis. Rather, they augment the infiltration of eosinophils into the conjunctiva during the effector phase of experimentally induced allergic conjunctivitis. PMID:16914468

  19. T3SEdb: data warehousing of virulence effectors secreted by the bacterial Type III Secretion System

    PubMed Central

    2010-01-01

    Background Effectors of Type III Secretion System (T3SS) play a pivotal role in establishing and maintaining pathogenicity in the host and therefore the identification of these effectors is important in understanding virulence. However, the effectors display high level of sequence diversity, therefore making the identification a difficult process. There is a need to collate and annotate existing effector sequences in public databases to enable systematic analyses of these sequences for development of models for screening and selection of putative novel effectors from bacterial genomes that can be validated by a smaller number of key experiments. Results Herein, we present T3SEdb http://effectors.bic.nus.edu.sg/T3SEdb, a specialized database of annotated T3SS effector (T3SE) sequences containing 1089 records from 46 bacterial species compiled from the literature and public protein databases. Procedures have been defined for i) comprehensive annotation of experimental status of effectors, ii) submission and curation review of records by users of the database, and iii) the regular update of T3SEdb existing and new records. Keyword fielded and sequence searches (BLAST, regular expression) are supported for both experimentally verified and hypothetical T3SEs. More than 171 clusters of T3SEs were detected based on sequence identity comparisons (intra-cluster difference up to ~60%). Owing to this high level of sequence diversity of T3SEs, the T3SEdb provides a large number of experimentally known effector sequences with wide species representation for creation of effector predictors. We created a reliable effector prediction tool, integrated into the database, to demonstrate the application of the database for such endeavours. Conclusions T3SEdb is the first specialised database reported for T3SS effectors, enriched with manual annotations that facilitated systematic construction of a reliable prediction model for identification of novel effectors. The T3SEdb represents a platform for inclusion of additional annotations of metadata for future developments of sophisticated effector prediction models for screening and selection of putative novel effectors from bacterial genomes/proteomes that can be validated by a small number of key experiments. PMID:21106126

  20. Two Fis Regulators Directly Repress the Expression of Numerous Effector-Encoding Genes in Legionella pneumophila

    PubMed Central

    Zusman, Tal; Speiser, Yariv

    2014-01-01

    Legionella pneumophila is an intracellular human pathogen that utilizes the Icm/Dot type IVB secretion system to translocate a large repertoire of effectors into host cells. For most of these effectors, there is no information regarding their regulation. Therefore, the aim of this study was to examine the involvement of the three L. pneumophila Fis homologs in the regulation of effector-encoding genes. Deletion mutants constructed in the genes encoding the three Fis regulators revealed that Fis1 (lpg0542 gene) and Fis3 (lpg1743) but not Fis2 (lpg1370) are partially required for intracellular growth of L. pneumophila in Acanthamoeba castellanii. To identify pathogenesis-related genes directly regulated by Fis, we established a novel in vivo system which resulted in the discovery of numerous effector-encoding genes directly regulated by Fis. Further examination of these genes revealed that Fis1 and Fis3 repress the level of expression of effector-encoding genes during exponential phase. Three groups of effector-encoding genes were identified: (i) effectors regulated mainly by Fis1, (ii) effectors regulated mainly by Fis3, and (iii) effectors regulated by both Fis1 and Fis3. Examination of the upstream regulatory region of all of these effector-encoding genes revealed multiple putative Fis regulatory elements, and site-directed mutagenesis confirmed that a few of these sites constitute part of a repressor binding element. Furthermore, gel mobility shift assays demonstrated the direct relation between the Fis1 and Fis3 regulators and these regulatory elements. Collectively, our results demonstrate for the first time that two of the three L. pneumophila Fis regulators directly repress the expression of Icm/Dot effector-encoding genes. PMID:25225276

  1. The Salmonella type III secretion system virulence effector forms a new hexameric chaperone assembly for export of effector/chaperone complexes

    DOE PAGESBeta

    Tsai, Chi -Lin; Burkinshaw, Brianne J.; Strynadka, Natalie C. J.; Tainer, John A.

    2014-12-08

    Bacteria hijack eukaryotic cells by injecting virulence effectors into host cytosol with a type III secretion system (T3SS). Effectors are targeted with their cognate chaperones to hexameric T3SS ATPase at the bacterial membrane's cytosolic face. In this issue of the Journal of Bacteriology, Roblin et al. (P. Roblin, F. Dewitte, V. Villeret, E. G. Biondi, and C. Bompard, J Bacteriol 197:688–698, 2015, http://dx.doi.org/10.1128/JB.02294-14) show that the T3SS chaperone SigE of Salmonella can form hexameric rings rather than dimers when bound to its cognate effector, SopB, implying a novel multimeric association for chaperone/effector complexes with their ATPase.

  2. The Salmonella type III secretion system virulence effector forms a new hexameric chaperone assembly for export of effector/chaperone complexes

    SciTech Connect

    Tsai, Chi -Lin; Burkinshaw, Brianne J.; Strynadka, Natalie C. J.; Tainer, John A.

    2014-12-08

    Bacteria hijack eukaryotic cells by injecting virulence effectors into host cytosol with a type III secretion system (T3SS). Effectors are targeted with their cognate chaperones to hexameric T3SS ATPase at the bacterial membrane's cytosolic face. In this issue of the Journal of Bacteriology, Roblin et al. (P. Roblin, F. Dewitte, V. Villeret, E. G. Biondi, and C. Bompard, J Bacteriol 197:688–698, 2015, http://dx.doi.org/10.1128/JB.02294-14) show that the T3SS chaperone SigE of Salmonella can form hexameric rings rather than dimers when bound to its cognate effector, SopB, implying a novel multimeric association for chaperone/effector complexes with their ATPase.

  3. Comparative reactivity of human IgE to cynomolgus monkey and human effector cells and effects on IgE effector cell potency

    PubMed Central

    Saul, Louise; Saul, Louise; Josephs, Debra H; Josephs, Debra H; Cutler, Keith; Cutler, Keith; Bradwell, Andrew; Bradwell, Andrew; Karagiannis, Panagiotis; Karagiannis, Panagiotis; Selkirk, Chris; Selkirk, Chris; Gould, Hannah J; Gould, Hannah J; Jones, Paul; Jones, Paul; Spicer, James F; Spicer, James F; Karagiannis, Sophia N; Karagiannis, Sophia N

    2014-01-01

    Background: Due to genetic similarities with humans, primates of the macaque genus such as the cynomolgus monkey are often chosen as models for toxicology studies of antibody therapies. IgE therapeutics in development depend upon engagement with the Fc?RI and Fc?RII receptors on immune effector cells for their function. Only limited knowledge of the primate IgE immune system is available to inform the choice of models for mechanistic and safety evaluations.   Methods: The recognition of human IgE by peripheral blood lymphocytes from cynomolgus monkey and man was compared. We used effector cells from each species in ex vivo affinity, dose-response, antibody-receptor dissociation and potency assays. Results: We report cross-reactivity of human IgE Fc with cynomolgus monkey cells, and comparable binding kinetics to peripheral blood lymphocytes from both species. In competition and dissociation assays, however, human IgE dissociated faster from cynomolgus monkey compared with human effector cells. Differences in association and dissociation kinetics were reflected in effector cell potency assays of IgE-mediated target cell killing, with higher concentrations of human IgE needed to elicit effector response in the cynomolgus monkey system. Additionally, human IgE binding on immune effector cells yielded significantly different cytokine release profiles in each species. Conclusion: These data suggest that human IgE binds with different characteristics to human and cynomolgus monkey IgE effector cells. This is likely to affect the potency of IgE effector functions in these two species, and so has relevance for the selection of biologically-relevant model systems when designing pre-clinical toxicology and functional studies. PMID:24492303

  4. A survey of the Pseudomonas syringae pv. tomato DC3000 type III secretion system effector repertoire reveals several effectors that are deleterious when expressed in Saccharomyces cerevisiae.

    PubMed

    Munkvold, Kathy R; Martin, Michael E; Bronstein, Philip A; Collmer, Alan

    2008-04-01

    The injection of nearly 30 effector proteins by the type III secretion system underlies the ability of Pseudomonas syringae pv. tomato DC3000 to cause disease in tomato and other host plants. The search for effector functions is complicated by redundancy within the repertoire and by plant resistance (R)-gene sentinels, which may convert effector virulence activities into a monolithic defense response. On the premise that some effectors target universal eukaryotic processes and that yeast (Saccharomyces cerevisiae) lacks R genes, the DC3000 effector repertoire was expressed in yeast. Of 27 effectors tested, HopAD1, HopAO1, HopD1, HopN1, and HopU1 were found to inhibit growth when expressed from a galactose-inducible GAL1 promoter, and HopAA1-1 and HopAM1 were found to cause cell death. Catalytic site mutations affecting the tyrosine phosphatase activity of HopAO1 and the cysteine protease activity of HopN1 prevented these effectors from inhibiting yeast growth. Expression of HopAA1-1, HopAM1, HopAD1, and HopAO1 impaired respiration in yeast, as indicated by tests with ethanol glycerol selective media. HopAA1-1 colocalized with porin to yeast mitochondria and was shown to cause cell death in yeast and plants in a domain-dependent manner. These results support the use of yeast for the study of plant-pathogen effector repertoires. PMID:18321194

  5. Targeting Human MicroRNA Genes Using Engineered Tal-Effector Nucleases (TALENs)

    E-print Network

    Shaw, Janet M.

    Targeting Human MicroRNA Genes Using Engineered Tal-Effector Nucleases (TALENs) Ruozhen Hu1 , Jared. Here, we report a novel approach to disrupting human miRNA genes ex vivo using engineered TAL Engineered Tal-Effector Nucleases (TALENs). PLoS ONE 8(5): e63074. doi:10.1371/journal.pone.0063074 Editor

  6. Micro-Review Roadmap for future research on plant pathogen effectors

    E-print Network

    in agriculture.mpp_588 805..814 INTRODUCTION Plant pathogens, including bacteria, fungi, oomycetes and nemaMicro-Review Roadmap for future research on plant pathogen effectors JAMES R. ALFANO* The Center 68588-0660, USA SUMMARY Bacterial and eukaryotic plant pathogens deliver effector pro- teins into plant

  7. How filamentous pathogens co-opt plants; the ins and outs of eukaryotic effectors

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Research on effectors secreted by pathogens during host attack has dominated the field of molecular plant-microbe interactions over recent years. Functional analysis of type III secreted effectors that are injected by pathogenic bacteria into host cells has significantly advanced the field and demon...

  8. Differential expression of candidate salivary effector proteins in field collections of Hessian fly, Mayetiola destructor

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Evidence is emerging that proteins secreted by gall forming plant-parasites are the effectors responsible for systemic changes in the host plant, such as galling and nutrient tissue formation. A large number of secreted salivary gland proteins (SSGPs) that are hypothesized to be the effectors respon...

  9. Transcription Activator-Like Effector Nucleases (TALENs): A Highly Efficient and Versatile Tool for

    E-print Network

    Zhao, Huimin

    REVIEW Transcription Activator-Like Effector Nucleases (TALENs): A Highly Efficient and Versatile.edu ABSTRACT: Transcription activator-like effector (TALE) nucleases (TALENs) have recently emerged double-strand breaks introduced by TALENs significantly increase the efficiency of genomic modification

  10. Pseudomonas syringae type III effector AvrRpt2 alters Arabidopsis thaliana auxin physiology

    E-print Network

    Sheen, Jen

    Pseudomonas syringae type III effector AvrRpt2 alters Arabidopsis thaliana auxin physiology type III effector AvrRpt2 promotes bacterial virulence on Arabidopsis thaliana plants lacking a func resistance on Arabidopsis thaliana plants carrying the resistance gene RPS2 (11, 12). AvrRpt2 also promotes

  11. Plant Immunity Directly or Indirectly Restricts the Injection of Type III Effectors by the Pseudomonas

    E-print Network

    -induced injection restriction. A P. syringae pv tomato DC3000 mutant lacking about one-third of its T3E inventory proteins. The response induced by this recogni- tion is termed effector-triggered immunity (ETI; Jones plant pathogen is generally due to the recognition of bacte- rial type III effector (T3E) proteins

  12. Integration of Target and Effector Information in Human Posterior Parietal Cortex for the Planning of Action

    E-print Network

    Vilis, Tutis

    in this decision-making process is some assessment of the cost of the movement by each effector to the selected. To generate movements, this spatial information must be integrated with the selected effector. We now tested-related functional MRI (fMRI), we located a bilateral region in the human posterior parietal cortex (ret

  13. Actin polymerization as a key innate immune effector mechanism to control Salmonella infection

    E-print Network

    Cicuta, Pietro

    Actin polymerization as a key innate immune effector mechanism to control Salmonella infection Si inflammasome effector func- tions and play a crucial role in controlling Salmonella infection. Inflammasome to directly kill intracellular salmonellae within macrophages. The mecha- nism, therefore, governing how

  14. NORE1A is a Ras senescence effector that controls the apoptotic/senescent balance of p53 via HIPK2

    PubMed Central

    Donninger, Howard; Calvisi, Diego F.; Barnoud, Thibaut; Clark, Jennifer; Schmidt, M. Lee; Vos, Michele D.

    2015-01-01

    The Ras oncoprotein is a key driver of cancer. However, Ras also provokes senescence, which serves as a major barrier to Ras-driven transformation. Ras senescence pathways remain poorly characterized. NORE1A is a novel Ras effector that serves as a tumor suppressor. It is frequently inactivated in tumors. We show that NORE1A is a powerful Ras senescence effector and that down-regulation of NORE1A suppresses senescence induction by Ras and enhances Ras transformation. We show that Ras induces the formation of a complex between NORE1A and the kinase HIPK2, enhancing HIPK2 association with p53. HIPK2 is a tumor suppressor that can induce either proapoptotic or prosenescent posttranslational modifications of p53. NORE1A acts to suppress its proapoptotic phosphorylation of p53 but enhance its prosenescent acetylation of p53. Thus, we identify a major new Ras signaling pathway that links Ras to the control of specific protein acetylation and show how NORE1A allows Ras to qualitatively modify p53 function to promote senescence. PMID:25778922

  15. Crystal Structure of Xanthomonas AvrRxo1-ORF1, a Type III Effector with a Polynucleotide Kinase Domain, and Its Interactor AvrRxo1-ORF2.

    PubMed

    Han, Qian; Zhou, Changhe; Wu, Shuchi; Liu, Yi; Triplett, Lindsay; Miao, Jiamin; Tokuhisa, James; Deblais, Loïc; Robinson, Howard; Leach, Jan E; Li, Jianyong; Zhao, Bingyu

    2015-10-01

    Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak (BLS) disease on rice plants. Xoc delivers a type III effector AvrRxo1-ORF1 into rice plant cells that can be recognized by disease resistance (R) protein Rxo1, and triggers resistance to BLS disease. However, the mechanism and virulence role of AvrRxo1 is not known. In the genome of Xoc, AvrRxo1-ORF1 is adjacent to another gene AvrRxo1-ORF2, which was predicted to encode a molecular chaperone of AvrRxo1-ORF1. We report the co-purification and crystallization of the AvrRxo1-ORF1:AvrRxo1-ORF2 tetramer complex at 1.64 Å resolution. AvrRxo1-ORF1 has a T4 polynucleotide kinase domain, and expression of AvrRxo1-ORF1 suppresses bacterial growth in a manner dependent on the kinase motif. Although AvrRxo1-ORF2 binds AvrRxo1-ORF1, it is structurally different from typical effector-binding chaperones, in that it has a distinct fold containing a novel kinase-binding domain. AvrRxo1-ORF2 functions to suppress the bacteriostatic activity of AvrRxo1-ORF1 in bacterial cells. PMID:26344722

  16. Growth hormone suppression test

    MedlinePLUS

    GH suppression test; Glucose loading test ... right away. The lab measures the glucose and GH levels in each sample. ... is because exercise or increased activity can change GH levels. If your child is to have this ...

  17. Molecular weaponry: diverse effectors delivered by the Type VI secretion system.

    PubMed

    Alcoforado Diniz, Juliana; Liu, Yi-Chia; Coulthurst, Sarah J

    2015-12-01

    The Type VI secretion system is a widespread bacterial nanomachine, used to deliver toxins directly into eukaryotic or prokaryotic target cells. These secreted toxins, or effectors, act on diverse cellular targets, and their action provides the attacking bacterial cell with a significant fitness advantage, either against rival bacteria or eukaryotic host organisms. In this review, we discuss the delivery of diverse effectors by the Type VI secretion system, the modes of action of the so-called 'anti-bacterial' and 'anti-eukaryotic' effectors, the mechanism of self-resistance against anti-bacterial effectors and the evolutionary implications of horizontal transfer of Type VI secretion system-associated toxins. Whilst it is likely that many more effectors remain to be identified, it is already clear that toxins delivered by this secretion system represent efficient weapons against both bacteria and eukaryotes. PMID:26432982

  18. MorTAL Kombat: the story of defense against TAL effectors through loss-of-susceptibility

    PubMed Central

    Hutin, Mathilde; Pérez-Quintero, Alvaro L.; Lopez, Camilo; Szurek, Boris

    2015-01-01

    Many plant-pathogenic xanthomonads rely on Transcription Activator-Like (TAL) effectors to colonize their host. This particular family of type III effectors functions as specific plant transcription factors via a programmable DNA-binding domain. Upon binding to the promoters of plant disease susceptibility genes in a sequence-specific manner, the expression of these host genes is induced. However, plants have evolved specific strategies to counter the action of TAL effectors and confer resistance. One mechanism is to avoid the binding of TAL effectors by mutations of their DNA binding sites, resulting in resistance by loss-of-susceptibility. This article reviews our current knowledge of the susceptibility hubs targeted by Xanthomonas TAL effectors, possible evolutionary scenarios for plants to combat the pathogen with loss-of-function alleles, and how this knowledge can be used overall to develop new pathogen-informed breeding strategies and improve crop resistance. PMID:26236326

  19. Independently Evolved Virulence Effectors Converge onto Hubs in a Plant Immune System Network

    PubMed Central

    Mukhtar, M. Shahid; Carvunis, Anne-Ruxandra; Dreze, Matija; Epple, Petra; Steinbrenner, Jens; Moore, Jonathan; Tasan, Murat; Galli, Mary; Hao, Tong; Nishimura, Marc T.; Pevzner, Samuel J.; Donovan, Susan E.; Ghamsari, Lila; Santhanam, Balaji; Romero, Viviana; Poulin, Matthew M.; Gebreab, Fana; Gutierrez, Bryan J.; Tam, Stanley; Monachello, Dario; Boxem, Mike; Harbort, Christopher J.; McDonald, Nathan; Gai, Lantian; Chen, Huaming; He, Yijian; Vandenhaute, Jean; Roth, Frederick P.; Hill, David E.; Ecker, Joseph R.; Vidal, Marc; Beynon, Jim; Braun, Pascal; Dangl, Jeffery L.

    2011-01-01

    Plants generate effective responses to infection by recognizing both conserved and variable pathogen-encoded molecules. Pathogens deploy virulence effector proteins into host cells, where they interact physically with host proteins to modulate defense. We generated a plant-pathogen immune system protein interaction network using effectors from two pathogens spanning the eukaryote-eubacteria divergence, three classes of Arabidopsis immune system proteins and ~8,000 other Arabidopsis proteins. We noted convergence of effectors onto highly interconnected host proteins, and indirect, rather than direct, connections between effectors and plant immune receptors. We demonstrated plant immune system functions for 15 of 17 tested host proteins that interact with effectors from both pathogens. Thus, pathogens from different kingdoms deploy independently evolved virulence proteins that interact with a limited set of highly connected cellular hubs to facilitate their diverse life cycle strategies. PMID:21798943

  20. Intrathymic programming of effector fates in three molecularly distinct ?? T cell subtypes

    PubMed Central

    Narayan, Kavitha; Sylvia, Katelyn E.; Malhotra, Nidhi; Yin, Catherine C.; Martens, Gregory; Vallerskog, Therese; Kornfeld, Hardy; Xiong, Na; Cohen, Nadia R.; Brenner, Michael B.; Berg, Leslie J.; Kang, Joonsoo

    2012-01-01

    ?? T cells function in the early phase of immune responses. Although innate ?? T cells have primarily been studied as one homogenous population, they can be functionally classified into effector subsets based on the production of signature cytokines, analogous to adaptive T helper subsets. Unlike adaptive T cells, however, ?? T effector function correlates with genomically encoded TCR chains, suggesting that clonal TCR selection is not the primary determinant of ?? effector differentiation. A high resolution transcriptome analysis of all emergent ?? thymocyte subsets segregated based on TCR?/? chain usage indicates the existence of three separate subtypes of ?? effectors in the thymus. The immature ?? subsets are distinguished by unique transcription factor modules that program effector function. PMID:22473038

  1. Structure of NS1A effector domain from the influenza A/Udorn/72 virus

    SciTech Connect

    Xia, Shuangluo; Monzingo, Arthur F.; Robertus, Jon D.

    2009-01-01

    The structure of the effector domain of the influenza protein NS1, a validated antiviral drug target, has been solved in two space groups. The nonstructural protein NS1A from influenza virus is a multifunctional virulence factor and a potent inhibitor of host immunity. It has two functional domains: an N-terminal 73-amino-acid RNA-binding domain and a C-terminal effector domain. Here, the crystallographic structure of the NS1A effector domain of influenza A/Udorn/72 virus is presented. Structure comparison with the NS1 effector domain from mouse-adapted influenza A/Puerto Rico/8/34 (PR8) virus strain reveals a similar monomer conformation but a different dimer interface. Further analysis and evaluation shows that the dimer interface observed in the structure of the PR8 NS1 effector domain is likely to be a crystallographic packing effect. A hypothetical model of the intact NS1 dimer is presented.

  2. Apparatus for adapting an end effector device remotely controlled manipulator arm

    NASA Technical Reports Server (NTRS)

    Clark, K. H. (inventor)

    1985-01-01

    Apparatus for adapting a general purpose and effector device to a special purpose and effector is disclosed which includes an adapter bracket assembly which provides a mechanical and electrical interface between the end effector devices. The adapter bracket assembly includes an adapter connector post which interlocks with a diamond shaped gripping channel formed in closed jaws of the general purpose end effector. The angularly intersecting surfaces of the connector post and gripping channel prevent any relative movement there between. Containment webs constrain the outer finger plates of the general purpose jaws to prevent pitch motion. Electrical interface is provided by conical, self aligning electrical connector components carried by respective ones of said end effectors.

  3. TALEs from a spring -- superelasticity of Tal effector protein structures

    E-print Network

    Holger Flechsig

    2014-06-30

    A simple force-probe setup is employed to study the mechanical properties of transcription activator-like effector (TALE) proteins in computer experiments. It is shown that their spring-like arrangement benefits superelastic behaviour which is manifested by large-scale global conformational changes along the helical axis, thus linking structure and dynamics in TALE proteins. As evidenced from the measured force-extension curves the dHax3 and PthXo1 TALEs behave like linear springs, obeying Hooke's law, for moderate global structural changes. For larger deformations, however, the proteins exhibit nonlinearities and the structures become stiffer the more they are stretched. Flexibility is not homogeneously distributed over TALE structure, but instead soft spots which correspond to the RVD loop residues and present key agents in the transmission of conformational motions are identified.

  4. TAL Effector DNA-Binding Principles and Specificity.

    PubMed

    Richter, Annekatrin; Streubel, Jana; Boch, Jens

    2016-01-01

    Transcription activator-like effectors (TALEs) are proteins with a unique DNA-binding domain that confers both a predictable and programmable specificity. The DNA-binding domain consists typically of 34-amino acid near-identical repeats. The repeats form a right-handed superhelical structure that wraps around the DNA double helix and exposes the variable amino acids at position 13 of each repeat to the sense strand DNA bases. Each repeat binds one base in a highly specific, non-overlapping, and comma-free fashion. Although TALE specificities are encoded in a simple way, sophisticated rules can be taken into account to build highly efficient DNA-binding modules for biotechnological use. PMID:26443210

  5. Intervention of Phytohormone Pathways by Pathogen Effectors[OPEN

    PubMed Central

    Kazan, Kemal; Lyons, Rebecca

    2014-01-01

    The constant struggle between plants and microbes has driven the evolution of multiple defense strategies in the host as well as offense strategies in the pathogen. To defend themselves from pathogen attack, plants often rely on elaborate signaling networks regulated by phytohormones. In turn, pathogens have adopted innovative strategies to manipulate phytohormone-regulated defenses. Tactics frequently employed by plant pathogens involve hijacking, evading, or disrupting hormone signaling pathways and/or crosstalk. As reviewed here, this is achieved mechanistically via pathogen-derived molecules known as effectors, which target phytohormone receptors, transcriptional activators and repressors, and other components of phytohormone signaling in the host plant. Herbivores and sap-sucking insects employ obligate pathogens such as viruses, phytoplasma, or symbiotic bacteria to intervene with phytohormone-regulated defenses. Overall, an improved understanding of phytohormone intervention strategies employed by pests and pathogens during their interactions with plants will ultimately lead to the development of new crop protection strategies. PMID:24920334

  6. Cell-Autonomous Effector Mechanisms against Mycobacterium tuberculosis

    PubMed Central

    MacMicking, John D.

    2014-01-01

    Few pathogens run the gauntlet of sterilizing immunity like Mycobacterium tuberculosis (Mtb). This organism infects mononuclear phagocytes and is also ingested by neutrophils, both of which possess an arsenal of cell-intrinsic effector mechanisms capable of eliminating it. Here Mtb encounters acid, oxidants, nitrosylating agents, and redox congeners, often exuberantly delivered under low oxygen tension. Further pressure is applied by withholding divalent Fe2+, Mn2+, Cu2+, and Zn2+, as well as by metabolic privation in the form of carbon needed for anaplerosis and aromatic amino acids for growth. Finally, host E3 ligases ubiquinate, cationic peptides disrupt, and lysosomal enzymes digest Mtb as part of the autophagic response to this particular pathogen. It is a testament to the evolutionary fitness of Mtb that sterilization is rarely complete, although sufficient to ensure most people infected with this airborne bacterium remain disease-free. PMID:25081628

  7. Type VI secretion effectors: poisons with a purpose

    PubMed Central

    Russell, Alistair B.; Peterson, S. Brook; Mougous, Joseph D.

    2014-01-01

    The type VI secretion system (T6SS) mediates interactions between a diverse range of Gram-negative bacterial species. Recent studies have led to a drastic increase in the number of characterized T6SS effector proteins and produced a more complete and nuanced view of the adaptive significance of the system. While the system is most often implicated in antagonism, in this review we consider the case for its involvement in both antagonistic and non-antagonistic behaviors. Clarifying the roles that T6S plays in microbial communities will contribute to broader efforts to understand the importance of microbial interactions in maintaining human and environmental health, and will inform efforts to manipulate these interactions for therapeutic or environmental benefit. PMID:24384601

  8. End effectors and attachments for buried waste excavation equipment

    SciTech Connect

    King, R.H.

    1993-09-01

    The Buried Waste Integrated Demonstration (BWID) supports the applied research, development, demonstration, and evaluation of a suite of advanced technologies that form a comprehensive remediation system for the effective and efficient remediation of buried waste. Their efforts are identified and coordinated in support of the U.S. Department of Energy (DOE), Environmental Restoration and Waste Management (ER&WM) Department`s needs and objectives. The present focus of BWID is to support retrieval and ex-situ treatment configuration options. Future activities will explore and support containment, and stabilization efforts in addition to the retrieval/ex situ treatment options. This report presents a literature search on the state-of-the-art in end effectors and attachments in support of excavator of buried transuranic waste. Included in the report are excavator platforms and a discussion of the various attachments. Also included is it list of vendors and specifications.

  9. Gibberellin Perception by the Gibberellin Receptor and its Effector Recognition

    NASA Astrophysics Data System (ADS)

    Hakoshima, Toshio; Murase, Kohji; Hirano, Yoshinori; Sun, Tai-Ping

    Gibberellins control a diverse range of growth and developmental processes in higher plants and have been widely utilized in the agricultural industry. By binding to a nuclear receptor GIBBERELLIN INSENSITIVE DWARF1 (GID1), gibberellins regulate gene expression by promoting degradation of the transcriptional regulator DELLA proteins. The precise manner in which GID1 discriminates and becomes activated by bioactive gibberellins for specific binding to DELLA proteins remains unclear. We present the crystal structure of a ternary complex of Arabidopsis thaliana GID1A, a bioactive gibberellin and the N-terminal DELLA domain of GAI. In this complex, GID1a occludes gibberellin in a deep binding pocket covered by its N-terminal helical switch region, which in turn interacts with the DELLA domain containing DELLA, VHYNP and LExLE motifs. Our results establish a structural model of a plant hormone receptor which is distinct from the hormone-perception mechanism and effector recognition of the known auxin receptors.

  10. Molecular regulation of effector and memory T cell differentiation

    PubMed Central

    Chang, John T; Wherry, E John; Goldrath, Ananda W

    2015-01-01

    Immunological memory is a cardinal feature of adaptive immunity and an important goal of vaccination strategies. Here we highlight advances in the understanding of the diverse T lymphocyte subsets that provide acute and long-term protection from infection. These include new insights into the transcription factors, and the upstream ‘pioneering’ factors that regulate their accessibility to key sites of gene regulation, as well as metabolic regulators that contribute to the differentiation of effector and memory subsets; ontogeny and defining characteristics of tissue-resident memory lymphocytes; and origins of the remarkable heterogeneity exhibited by activated T cells. Collectively, these findings underscore progress in delineating the underlying pathways that control diversification in T cell responses but also reveal gaps in the knowledge, as well as the challenges that arise in the application of this knowledge to rationally elicit desired T cell responses through vaccination and immunotherapy. PMID:25396352

  11. Enhancement of phototransduction g protein-effector interactions by phosphoinositides.

    PubMed

    He, Feng; Mao, Muling; Wensel, Theodore G

    2004-03-01

    Light responses in photoreceptor cells are mediated by the action of the G protein transducin (G(t)) on the effector enzyme cGMP phosphodiesterase (PDE6) at the surface of disk membranes. The enzymatic components needed for phosphoinositide-based signaling are known to be present in rod cells, but it has remained uncertain what role phosphoinositides play in vertebrate phototransduction. Reconstitution of PDE6 and activated G(alphat), on the surface of large unilamellar vesicles containing d-myo-phosphatidylinositol-4,5-bisphosphate (PI(4,5)P(2)), stimulated PDE activity nearly 4-fold above the level observed with membranes containing no phosphoinositides, whereas G protein-independent activation by trypsin was unaffected by the presence of phosphoinositides. PDE activity was similarly stimulated by d-myo-phosphatidylinositol-3,4-bisphosphate and d-myo-phosphatidylinositol-4-phosphate (PI(4)P), but much less by d-myo-phosphatidylinositol-5-phosphate (PI(5)P) or d-myo-phosphatidylinositol-3,5-bisphosphate. Incubation of rod outer segment membranes with phosphoinositide-specific phospholipase C decreased G protein-stimulated activation of endogenous PDE6, but not trypsin-stimulated PDE activity. Binding experiments using phosphoinositide-containing vesicles revealed patterns of PDE6 binding and PDE6-enhanced G(alphat)-GTPgammaS binding, consistent with the activation profile PI(4,5)P(2) > PI(4)P > PI(5)P approximately control vesicles. These results suggest that enhancement of effector-G protein interactions represents a possible mechanism for modulation of phototransduction gain by changes in phosphoinositide levels, perhaps occurring in response to longterm changes in illumination or other environmental cues. PMID:14699118

  12. Protection after stroke: cellular effectors of neurovascular unit integrity

    PubMed Central

    Posada-Duque, Rafael Andres; Barreto, George E.; Cardona-Gomez, Gloria Patricia

    2014-01-01

    Neurological disorders are prevalent worldwide. Cerebrovascular diseases (CVDs), which account for 55% of all neurological diseases, are the leading cause of permanent disability, cognitive and motor disorders and dementia. Stroke affects the function and structure of blood-brain barrier, the loss of cerebral blood flow regulation, oxidative stress, inflammation and the loss of neural connections. Currently, no gold standard treatments are available outside the acute therapeutic window to improve outcome in stroke patients. Some promising candidate targets have been identified for the improvement of long-term recovery after stroke, such as Rho GTPases, cell adhesion proteins, kinases, and phosphatases. Previous studies by our lab indicated that Rho GTPases (Rac and RhoA) are involved in both tissue damage and survival, as these proteins are essential for the morphology and movement of neurons, astrocytes and endothelial cells, thus playing a critical role in the balance between cell survival and death. Treatment with a pharmacological inhibitor of RhoA/ROCK blocks the activation of the neurodegeneration cascade. In addition, Rac and synaptic adhesion proteins (p120 catenin and N-catenin) play critical roles in protection against cerebral infarction and in recovery by supporting the neurovascular unit and cytoskeletal remodeling activity to maintain the integrity of the brain parenchyma. Interestingly, neuroprotective agents, such as atorvastatin, and CDK5 silencing after cerebral ischemia and in a glutamate-induced excitotoxicity model may act on the same cellular effectors to recover neurovascular unit integrity. Therefore, future efforts must focus on individually targeting the structural and functional roles of each effector of neurovascular unit and the interactions in neural and non-neural cells in the post-ischemic brain and address how to promote the recovery or prevent the loss of homeostasis in the short, medium and long term. PMID:25177270

  13. Wake of the flood: ascribing functions to the wave of type III effector proteins of phytopathogenic bacteria

    E-print Network

    Dangl, Jeff

    Wake of the flood: ascribing functions to the wave of type III effector proteins of phytopathogenic cell for the pathogen's benefit. This is evidenced by the flood of effector genes that have recently

  14. Suppression of developmental anomalies by maternal macrophages in mice

    SciTech Connect

    Nomura, T.; Hata, S.; Kusafuka, T. )

    1990-11-01

    We tested whether nonspecific tumoricidal immune cells can suppress congenital malformations by killing precursor cells destined to cause such defects. Pretreatment of pregnant ICR mice with synthetic (Pyran copolymer) and biological (Bacillus Calmette-Guerin) agents significantly suppressed radiation- and chemical-induced congenital malformations (cleft palate, digit anomalies, tail anomalies, etc.). Such suppressive effects were associated with the activation of maternal macrophages by these agents, but were lost either after the disruption of activated macrophages by supersonic waves or by inhibition of their lysosomal enzyme activity with trypan blue. These results indicate that a live activated macrophage with active lysosomal enzymes can be an effector cell to suppress maldevelopment. A similar reduction by activated macrophages was observed in strain CL/Fr, which has a high spontaneous frequency of cleft lips and palates. Furthermore, Pyran-activated maternal macrophages could pass through the placenta, and enhanced urethane-induced cell killing (but not somatic mutation) in the embryo. It is likely that a maternal immunosurveillance system eliminating preteratogenic cells allows for the replacement with normal totipotent blast cells during the pregnancy to protect abnormal development.

  15. The genome sequence and effector complement of the flax rust pathogen Melampsora lini.

    PubMed

    Nemri, Adnane; Saunders, Diane G O; Anderson, Claire; Upadhyaya, Narayana M; Win, Joe; Lawrence, Gregory J; Jones, David A; Kamoun, Sophien; Ellis, Jeffrey G; Dodds, Peter N

    2014-01-01

    Rust fungi cause serious yield reductions on crops, including wheat, barley, soybean, coffee, and represent real threats to global food security. Of these fungi, the flax rust pathogen Melampsora lini has been developed most extensively over the past 80 years as a model to understand the molecular mechanisms that underpin pathogenesis. During infection, M. lini secretes virulence effectors to promote disease. The number of these effectors, their function and their degree of conservation across rust fungal species is unknown. To assess this, we sequenced and assembled de novo the genome of M. lini isolate CH5 into 21,130 scaffolds spanning 189 Mbp (scaffold N50 of 31 kbp). Global analysis of the DNA sequence revealed that repetitive elements, primarily retrotransposons, make up at least 45% of the genome. Using ab initio predictions, transcriptome data and homology searches, we identified 16,271 putative protein-coding genes. An analysis pipeline was then implemented to predict the effector complement of M. lini and compare it to that of the poplar rust, wheat stem rust and wheat stripe rust pathogens to identify conserved and species-specific effector candidates. Previous knowledge of four cloned M. lini avirulence effector proteins and two basidiomycete effectors was used to optimize parameters of the effector prediction pipeline. Markov clustering based on sequence similarity was performed to group effector candidates from all four rust pathogens. Clusters containing at least one member from M. lini were further analyzed and prioritized based on features including expression in isolated haustoria and infected leaf tissue and conservation across rust species. Herein, we describe 200 of 940 clusters that ranked highest on our priority list, representing 725 flax rust candidate effectors. Our findings on this important model rust species provide insight into how effectors of rust fungi are conserved across species and how they may act to promote infection on their hosts. PMID:24715894

  16. QueTAL: a suite of tools to classify and compare TAL effectors functionally and phylogenetically

    PubMed Central

    Pérez-Quintero, Alvaro L.; Lamy, Léo; Gordon, Jonathan L.; Escalon, Aline; Cunnac, Sébastien; Szurek, Boris; Gagnevin, Lionel

    2015-01-01

    Transcription Activator-Like (TAL) effectors from Xanthomonas plant pathogenic bacteria can bind to the promoter region of plant genes and induce their expression. DNA-binding specificity is governed by a central domain made of nearly identical repeats, each determining the recognition of one base pair via two amino acid residues (a.k.a. Repeat Variable Di-residue, or RVD). Knowing how TAL effectors differ from each other within and between strains would be useful to infer functional and evolutionary relationships, but their repetitive nature precludes reliable use of traditional alignment methods. The suite QueTAL was therefore developed to offer tailored tools for comparison of TAL effector genes. The program DisTAL considers each repeat as a unit, transforms a TAL effector sequence into a sequence of coded repeats and makes pair-wise alignments between these coded sequences to construct trees. The program FuncTAL is aimed at finding TAL effectors with similar DNA-binding capabilities. It calculates correlations between position weight matrices of potential target DNA sequence predicted from the RVD sequence, and builds trees based on these correlations. The programs accurately represented phylogenetic and functional relationships between TAL effectors using either simulated or literature-curated data. When using the programs on a large set of TAL effector sequences, the DisTAL tree largely reflected the expected species phylogeny. In contrast, FuncTAL showed that TAL effectors with similar binding capabilities can be found between phylogenetically distant taxa. This suite will help users to rapidly analyse any TAL effector genes of interest and compare them to other available TAL genes and should improve our understanding of TAL effectors evolution. It is available at http://bioinfo-web.mpl.ird.fr/cgi-bin2/quetal/quetal.cgi. PMID:26284082

  17. The genome sequence and effector complement of the flax rust pathogen Melampsora lini

    PubMed Central

    Nemri, Adnane; Saunders, Diane G. O.; Anderson, Claire; Upadhyaya, Narayana M.; Win, Joe; Lawrence, Gregory J.; Jones, David A.; Kamoun, Sophien; Ellis, Jeffrey G.; Dodds, Peter N.

    2014-01-01

    Rust fungi cause serious yield reductions on crops, including wheat, barley, soybean, coffee, and represent real threats to global food security. Of these fungi, the flax rust pathogen Melampsora lini has been developed most extensively over the past 80 years as a model to understand the molecular mechanisms that underpin pathogenesis. During infection, M. lini secretes virulence effectors to promote disease. The number of these effectors, their function and their degree of conservation across rust fungal species is unknown. To assess this, we sequenced and assembled de novo the genome of M. lini isolate CH5 into 21,130 scaffolds spanning 189 Mbp (scaffold N50 of 31 kbp). Global analysis of the DNA sequence revealed that repetitive elements, primarily retrotransposons, make up at least 45% of the genome. Using ab initio predictions, transcriptome data and homology searches, we identified 16,271 putative protein-coding genes. An analysis pipeline was then implemented to predict the effector complement of M. lini and compare it to that of the poplar rust, wheat stem rust and wheat stripe rust pathogens to identify conserved and species-specific effector candidates. Previous knowledge of four cloned M. lini avirulence effector proteins and two basidiomycete effectors was used to optimize parameters of the effector prediction pipeline. Markov clustering based on sequence similarity was performed to group effector candidates from all four rust pathogens. Clusters containing at least one member from M. lini were further analyzed and prioritized based on features including expression in isolated haustoria and infected leaf tissue and conservation across rust species. Herein, we describe 200 of 940 clusters that ranked highest on our priority list, representing 725 flax rust candidate effectors. Our findings on this important model rust species provide insight into how effectors of rust fungi are conserved across species and how they may act to promote infection on their hosts. PMID:24715894

  18. MITEs in the promoters of effector genes allow prediction of novel virulence genes in Fusarium oxysporum

    PubMed Central

    2013-01-01

    Background The plant-pathogenic fungus Fusarium oxysporum f.sp.lycopersici (Fol) has accessory, lineage-specific (LS) chromosomes that can be transferred horizontally between strains. A single LS chromosome in the Fol4287 reference strain harbors all known Fol effector genes. Transfer of this pathogenicity chromosome confers virulence to a previously non-pathogenic recipient strain. We hypothesize that expression and evolution of effector genes is influenced by their genomic context. Results To gain a better understanding of the genomic context of the effector genes, we manually curated the annotated genes on the pathogenicity chromosome and identified and classified transposable elements. Both retro- and DNA transposons are present with no particular overrepresented class. Retrotransposons appear evenly distributed over the chromosome, while DNA transposons tend to concentrate in large chromosomal subregions. In general, genes on the pathogenicity chromosome are dispersed within the repeat landscape. Effector genes are present within subregions enriched for DNA transposons. A miniature Impala (mimp) is always present in their promoters. Although promoter deletion studies of two effector gene loci did not reveal a direct function of the mimp for gene expression, we were able to use proximity to a mimp as a criterion to identify new effector gene candidates. Through xylem sap proteomics we confirmed that several of these candidates encode proteins secreted during plant infection. Conclusions Effector genes in Fol reside in characteristic subregions on a pathogenicity chromosome. Their genomic context allowed us to develop a method for the successful identification of novel effector genes. Since our approach is not based on effector gene similarity, but on unique genomic features, it can easily be extended to identify effector genes in Fo strains with different host specificities. PMID:23432788

  19. Diverse Secreted Effectors Are Required for Salmonella Persistence in a Mouse Infection Model

    SciTech Connect

    Kidwai, Afshan S.; Mushamiri, Ivy T.; Niemann, George; Brown, Roslyn N.; Adkins, Joshua N.; Heffron, Fred

    2013-08-12

    Salmonella enterica serovar Typhimurium causes typhoid-like disease in mice and is a model of typhoid fever in humans. One of the hallmarks of typhoid is persistence, the ability of the bacteria to survive in the host weeks after infection. Virulence factors called effectors facilitate this process by direct transfer to the cytoplasm of infected cells thereby subverting cellular processes. Secretion of effectors to the cell cytoplasm takes place through multiple routes, including two separate type III secretion (T3SS) apparati as well as outer membrane vesicles. The two T3SS are encoded on separate pathogenicity islands, SPI-1 and -2, with SPI-1 more strongly associated with the intestinal phase of infection, and SPI-2 with the systemic phase. Both T3SS are required for persistence, but the effectors required have not been systematically evaluated. In this study, mutations in 48 described effectors were tested for persistence. We replaced each effector with a specific DNA barcode sequence by allelic exchange and co-infected with a wild-type reference to calculate the ratio of wild-type parent to mutant at different times after infection. The competitive index (CI) was determined by quantitative PCR in which primers that correspond to the barcode were used for amplification. Mutations in all but seven effectors reduced persistence demonstrating that most effectors were required. One exception was CigR, a recently discovered effector that is widely conserved throughout enteric bacteria. Deletion of cigR increased lethality, suggesting that it may be an anti-virulence factor. The fact that almost all Salmonella effectors are required for persistence argues against redundant functions. This is different from effector repertoires in other intracellular pathogens such as Legionella.

  20. A rapid one-generation genetic screen in a Drosophila model to capture rhabdomyosarcoma effectors and therapeutic targets.

    PubMed

    Galindo, Kathleen A; Endicott, Tiana R; Avirneni-Vadlamudi, Usha; Galindo, Rene L

    2015-02-01

    Rhabdomyosarcoma (RMS) is an aggressive childhood malignancy of neoplastic muscle-lineage precursors that fail to terminally differentiate into syncytial muscle. The most aggressive form of RMS, alveolar-RMS, is driven by misexpression of the PAX-FOXO1 oncoprotein, which is generated by recurrent chromosomal translocations that fuse either the PAX3 or PAX7 gene to FOXO1. The molecular underpinnings of PAX-FOXO1-mediated RMS pathogenesis remain unclear, however, and clinical outcomes poor. Here, we report a new approach to dissect RMS, exploiting a highly efficient Drosophila PAX7-FOXO1 model uniquely configured to uncover PAX-FOXO1 RMS genetic effectors in only one generation. With this system, we have performed a comprehensive deletion screen against the Drosophila autosomes and demonstrate that mutation of Mef2, a myogenesis lynchpin in both flies and mammals, dominantly suppresses PAX7-FOXO1 pathogenicity and acts as a PAX7-FOXO1 gene target. Additionally, we reveal that mutation of mastermind, a gene encoding a MEF2 transcriptional coactivator, similarly suppresses PAX7-FOXO1, further pointing toward MEF2 transcriptional activity as a PAX-FOXO1 underpinning. These studies show the utility of the PAX-FOXO1 Drosophila system as a robust one-generation (F1) RMS gene discovery platform and demonstrate how Drosophila transgenic conditional expression models can be configured for the rapid dissection of human disease. PMID:25491943

  1. A Rapid One-Generation Genetic Screen in a Drosophila Model to Capture Rhabdomyosarcoma Effectors and Therapeutic Targets

    PubMed Central

    Galindo, Kathleen A.; Endicott, Tiana R.; Avirneni-Vadlamudi, Usha; Galindo, Rene L.

    2014-01-01

    Rhabdomyosarcoma (RMS) is an aggressive childhood malignancy of neoplastic muscle-lineage precursors that fail to terminally differentiate into syncytial muscle. The most aggressive form of RMS, alveolar-RMS, is driven by misexpression of the PAX-FOXO1 oncoprotein, which is generated by recurrent chromosomal translocations that fuse either the PAX3 or PAX7 gene to FOXO1. The molecular underpinnings of PAX-FOXO1?mediated RMS pathogenesis remain unclear, however, and clinical outcomes poor. Here, we report a new approach to dissect RMS, exploiting a highly efficient Drosophila PAX7-FOXO1 model uniquely configured to uncover PAX-FOXO1 RMS genetic effectors in only one generation. With this system, we have performed a comprehensive deletion screen against the Drosophila autosomes and demonstrate that mutation of Mef2, a myogenesis lynchpin in both flies and mammals, dominantly suppresses PAX7-FOXO1 pathogenicity and acts as a PAX7-FOXO1 gene target. Additionally, we reveal that mutation of mastermind, a gene encoding a MEF2 transcriptional coactivator, similarly suppresses PAX7-FOXO1, further pointing toward MEF2 transcriptional activity as a PAX-FOXO1 underpinning. These studies show the utility of the PAX-FOXO1 Drosophila system as a robust one-generation (F1) RMS gene discovery platform and demonstrate how Drosophila transgenic conditional expression models can be configured for the rapid dissection of human disease. PMID:25491943

  2. Auto-acetylation on K289 is not essential for HopZ1a-mediated plant defense suppression

    PubMed Central

    Rufián, José S.; Lucía, Ainhoa; Macho, Alberto P.; Orozco-Navarrete, Begoña; Arroyo-Mateos, Manuel; Bejarano, Eduardo R.; Beuzón, Carmen R.; Ruiz-Albert, Javier

    2015-01-01

    The Pseudomonas syringae type III-secreted effector HopZ1a is a member of the HopZ/YopJ superfamily of effectors that triggers immunity in Arabidopsis. We have previously shown that HopZ1a suppresses both local [effector-triggered immunity (ETI)] and systemic immunity [systemic acquired resistance (SAR)] triggered by the heterologous effector AvrRpt2. HopZ1a has been shown to possess acetyltransferase activity, and this activity is essential to trigger immunity in Arabidopsis. HopZ1a acetyltransferase activity has been reported to require the auto-acetylation of the effector on a specific lysine (K289) residue. In this paper we analyze the relevance of autoacetylation of lysine residue 289 in HopZ1a ability to suppress plant defenses, and on the light of the results obtained, we also revise its relevance for HopZ1a avirulence activity. Our results indicate that, while the HopZ1aK289R mutant is impaired to some degree in its virulence and avirulence activities, is by no means phenotypically equivalent to the catalytically inactive HopZ1aC216A, since it is still able to trigger a defense response that induces detectable macroscopic HR and effectively protects Arabidopsis from infection, reducing growth of P. syringae within the plant. We also present evidence that the HopZ1aK289R mutant still displays virulence activities, partially suppressing both ETI and SAR. PMID:26217317

  3. Design schemes and comparison research of the end-effector of large space manipulator

    NASA Astrophysics Data System (ADS)

    Feng, Fei; Liu, Yiwei; Liu, Hong; Cai, Hegao

    2012-07-01

    The end-effector of the large space manipulator is employed to assist the manipulator in handling and manipulating large payloads on orbit. Currently, there are few researches about the end-effector, and the existing end-effectors have some disadvantages, such as poor misalignment tolerance capability and complex mechanical components. According to the end positioning errors and the residual vibration characters of the large space manipulators, two basic performance requirements of the end-effector which include the capabilities of misalignment tolerance and soft capture are proposed. And the end-effector should accommodate the following misalignments of the mechanical interface. The translation misalignments in axial and radial directions and the angular misalignments in roll, pitch and yaw are ±100 mm, 100 mm, ±10o, ±15o, ±15o, respectively. Seven end-effector schemes are presented and the capabilities of misalignment tolerance and soft capture are analyzed elementarily. The three fingers-three petals end-effector and the steel cable-snared end-effector are the most feasible schemes among the seven schemes, and they are designed in detail. The capabilities of misalignment tolerance and soft capture are validated and evaluated, through the experiment on the micro-gravity simulating device and the dynamic analysis in ADAMS software. The results show that the misalignment tolerance capabilities of these two schemes could satisfy the requirement. And the translation misalignment tolerances in axial and radial directions and the angular misalignment tolerances in roll, pitch and yaw of the steel cable-snared end-effector are 30mm, 15mm, 6o, 3o and 3o larger than those of the three fingers-three petals end-effector, respectively. And the contact force of the steel cable-snared end-effector is smaller and smoother than that of the three fingers-three petals end-effector. The end-effector schemes and research methods are beneficial to the developments of the large space manipulator end-effctor and the space docking mechanism.

  4. siRNA-mediated silencing of PD-1 ligands enhances tumor-specific human T-cell effector functions.

    PubMed

    Iwamura, K; Kato, T; Miyahara, Y; Naota, H; Mineno, J; Ikeda, H; Shiku, H

    2012-10-01

    Adoptive cell therapy using tumor-specific T cells is a promising strategy for treating patients with malignancy. However, accumulating evidences have demonstrated that optimal function of tumor-reactive T cells is often attenuated by negative regulatory signal(s) delivered through receptors, such as cytotoxic T-lymphocyte antigen 4 (CTLA-4), programmed death 1 (PD-1), and their cognate ligands. Although systemic blocking of these molecules needs careful attention on the risk of uncontrolled immune activation, selective inhibition of negative signals in tumor-specific T cells by their genetic modification is an attractive approach to overcome immunological suppression in cancer patients. Here, we demonstrate the improved effector functions of tumor-specific CD4(+) and CD8(+) human T cells by small interfering RNA (siRNA) -mediated silencing of PD-1 ligands, PD-L1 or PD-L2. Tumor antigen MAGE-A4-specific human T-cell clones upregulated the expression of PD-1 ligands upon activation. siRNA-mediated knockdown of PD-L1 or -L2 enhanced the interferon-? production and antigen-specific cytotoxicity of these cells. Peripheral blood mononuclear cells transduced with a retroviral vector encoding MAGE-A4-specific T-cell receptor ?/? chains also increased their effector functions by this modification. These results suggest that siRNA-mediated knockdown of PD-1 ligands is an attractive strategy to inhibit a negative regulatory mechanism of tumor-specific T cells resulting in enhanced efficacy of adoptive T-cell therapy of cancer using genetically modified autologous lymphocytes. PMID:22113316

  5. NopC Is a Rhizobium-Specific Type 3 Secretion System Effector Secreted by Sinorhizobium (Ensifer) fredii HH103

    PubMed Central

    Medina, Carlos; Ollero, Francisco Javier; López-Baena, Francisco Javier

    2015-01-01

    Sinorhizobium (Ensifer) fredii HH103 is a broad host-range nitrogen-fixing bacterium able to nodulate many legumes, including soybean. In several rhizobia, root nodulation is influenced by proteins secreted through the type 3 secretion system (T3SS). This specialized secretion apparatus is a common virulence mechanism of many plant and animal pathogenic bacteria that delivers proteins, called effectors, directly into the eukaryotic host cells where they interfere with signal transduction pathways and promote infection by suppressing host defenses. In rhizobia, secreted proteins, called nodulation outer proteins (Nops), are involved in host-range determination and symbiotic efficiency. S. fredii HH103 secretes at least eight Nops through the T3SS. Interestingly, there are Rhizobium-specific Nops, such as NopC, which do not have homologues in pathogenic bacteria. In this work we studied the S. fredii HH103 nopC gene and confirmed that its expression was regulated in a flavonoid-, NodD1- and TtsI-dependent manner. Besides, in vivo bioluminescent studies indicated that the S. fredii HH103 T3SS was expressed in young soybean nodules and adenylate cyclase assays confirmed that NopC was delivered directly into soybean root cells by means of the T3SS machinery. Finally, nodulation assays showed that NopC exerted a positive effect on symbiosis with Glycine max cv. Williams 82 and Vigna unguiculata. All these results indicate that NopC can be considered a Rhizobium-specific effector secreted by S. fredii HH103. PMID:26569401

  6. Development of a Mild Viral Expression System for Gain-Of-Function Study of Phytoplasma Effector In Planta

    PubMed Central

    Liu, Li-Yu Daisy; Hong, Syuan-Fei; Yang, Chiao-Yin; Lo, Hsiao-Feng; Tseng, Ting-Yu; Chen, Wei-Yao; Lin, Shih-Shun

    2015-01-01

    PHYL1 and SAP54 are orthologs of pathogenic effectors of Aster yellow witches’-broom (AYWB) phytoplasma and Peanut witches’-broom (PnWB) phytoplasma, respectively. These effectors cause virescence and phyllody symptoms (hereafter leafy flower) in phytoplasma-infected plants. T0 lines of transgenic Arabidopsis expressing the PHYL1 or SAP54 genes (PHYL1 or SAP54 plants) show a leafy flower phenotype and result in seedless, suggesting that PHYL1 and SAP54 interfere with reproduction stage that restrict gain-of-function studies in the next generation of transgenic plants. Turnip mosaic virus (TuMV) mild strain (TuGK) has an Arg182Lys mutation in the helper-component proteinase (HC-ProR182K) that blocks suppression of the miRNA pathway and prevents symptom development in TuGK-infected plants. We exploited TuGK as a viral vector for gain-of-function studies of PHYL1 and SAP54 in Arabidopsis plants. TuGK-PHYL1- and TuGK-SAP54-infected Arabidopsis plants produced identical leafy flower phenotypes and similar gene expression profiles as PHYL1 and SAP54 plants. In addition, the leafy flower formation rate was enhanced in TuGK-PHYL1- or TuGK-SAP54-infected Arabidopsis plants that compared with the T0 lines of PHYL1 plants. These results provide more evidence and novel directions for further studying the mechanism of PHYL1/SAP54-mediated leafy flower development. In addition, the TuGK vector is a good alternative in transgenic plant approaches for rapid gene expression in gain-of-function studies. PMID:26076458

  7. The Pseudomonas syringae effector protein, AvrRPS4, requires in planta processing and the KRVY domain to function.

    PubMed

    Sohn, Kee Hoon; Zhang, Yan; Jones, Jonathan D G

    2009-03-01

    A Pseudomonas syringae pv. pisi effector protein, AvrRPS4, triggers RPS4-dependent immunity in Arabidopsis. We characterized biochemical and genetic aspects of AvrRPS4 function. Secretion of AvrRPS4 from Pst DC3000 is type III secretion-dependent, and AvrRPS4 is processed into a smaller form in plant cells but not in bacteria or yeast. Agrobacterium-mediated transient expression analysis of N-terminally truncated AvrRPS4 mutants revealed that the C-terminal 88 amino acids are sufficient to trigger the hypersensitive response in turnip. N-terminal sequencing of the processed AvrRPS4 showed that processing occurs between G133 and G134. The processing-deficient mutant, R112L, still triggers RPS4-dependent immunity, suggesting that the processing is not required for the AvrRPS4 avirulence function. AvrRPS4 enhances bacterial growth when delivered by Pta 6606 into Nicotiana benthamiana in which AvrRPS4 is not recognized. Transgenic expression of AvrRPS4 in the Arabidopsis rps4 mutant enhances the growth of Pst DC3000 and suppresses PTI (PAMP-triggered immunity), showing that AvrRPS4 promotes virulence in two distinct host plants. Furthermore, full virulence activity of AvrRPS4 requires both proteolytic processing and the KRVY motif at the N-terminus of processed AvrRPS4. XopO, an Xcv effector, shares the amino acids required for AvrRPS4 processing and the KRVY motif. XopO is also processed into a smaller form in N. benthamiana, similar to AvrRPS4, suggesting that a common mechanism is involved in activation of the virulence activities of both AvrRPS4 and XopO. PMID:19054367

  8. The effector T cell response to ryegrass pollen is counterregulated by simultaneous induction of regulatory T cells.

    PubMed

    Mittag, Diana; Scholzen, Anja; Varese, Nirupama; Baxter, Lorraine; Paukovics, Geza; Harrison, Leonard C; Rolland, Jennifer M; O'Hehir, Robyn E

    2010-05-01

    Allergy is associated with pathological Th2 responses to otherwise harmless environmental Ags. In contrast, nonallergic individuals mount nonpathological immune responses to allergens, partly attributed to regulatory T cell (Treg) activity. Although thymus-derived natural Tregs have been shown to maintain tolerance to self-Ags and prevent autoimmunity, the generation of Tregs specific to non-self-Ags is less well understood. We investigated the potential for induction of Tregs from PBMCs of ryegrass pollen-allergic or healthy subjects by stimulation in vitro with ryegrass pollen extract in the absence of additional exogenous stimuli. We found that two subsets of proliferating CD4(+) T cells were induced, one expressing intermediate levels of Foxp3 (and IFN-gamma, IL-4, IL-17, or IL-2) and the other expressing high levels of Foxp3 (and no effector cytokines). After enrichment based on CD39 expression, the Foxp3(hi) subset suppressed CD4(+) T cell proliferation and IFN-gamma production. The Foxp3(hi) Treg originated from both conversion of dividing non-Tregs (CD4(+)CD25(-)CD127(hi)) and expansion of natural Tregs (CD4(+)CD25(+)CD127(lo)). Stable functional Tregs expressing high levels of Foxp3 were induced simultaneously with effector T cells by allergen stimulation. Induction of Foxp3(hi) Tregs was reduced in allergic subjects. These results indicate that the cogeneration of Foxp3(hi) Tregs in response to allergen may be a mechanism for controlling allergic reactions in healthy individuals, which is impaired in those with allergies. PMID:20308632

  9. The potential of effector-target genes in breeding for plant innate immunity

    PubMed Central

    Gawehns, Fleur; Cornelissen, Ben J C; Takken, Frank L W

    2013-01-01

    Increasing numbers of infectious crop diseases that are caused by fungi and oomycetes urge the need to develop alternative strategies for resistance breeding. As an alternative for the use of resistance (R) genes, the application of mutant susceptibility (S) genes has been proposed as a potentially more durable type of resistance. Identification of S genes is hampered by their recessive nature. Here we explore the use of pathogen-derived effectors as molecular probes to identify S genes. Effectors manipulate specific host processes thereby contributing to disease. Effector targets might therefore represent S genes. Indeed, the Pseudomonas syringae effector HopZ2 was found to target MLO2, an Arabidopsis thaliana homologue of the barley S gene Mlo. Unfortunately, most effector targets identified so far are not applicable as S genes due to detrimental effects they have on other traits. However, some effector targets such as Mlo are successfully used, and with the increase in numbers of effector targets being identified, the numbers of S genes that can be used in resistance breeding will rise as well. PMID:23279965

  10. The potential of effector-target genes in breeding for plant innate immunity.

    PubMed

    Gawehns, Fleur; Cornelissen, Ben J C; Takken, Frank L W

    2013-05-01

    Increasing numbers of infectious crop diseases that are caused by fungi and oomycetes urge the need to develop alternative strategies for resistance breeding. As an alternative for the use of resistance (R) genes, the application of mutant susceptibility (S) genes has been proposed as a potentially more durable type of resistance. Identification of S genes is hampered by their recessive nature. Here we explore the use of pathogen-derived effectors as molecular probes to identify S genes. Effectors manipulate specific host processes thereby contributing to disease. Effector targets might therefore represent S genes. Indeed, the Pseudomonas syringae effector HopZ2 was found to target MLO2, an Arabidopsis thaliana homologue of the barley S gene Mlo. Unfortunately, most effector targets identified so far are not applicable as S genes due to detrimental effects they have on other traits. However, some effector targets such as Mlo are successfully used, and with the increase in numbers of effector targets being identified, the numbers of S genes that can be used in resistance breeding will rise as well. PMID:23279965

  11. Chimeric adaptor proteins translocate diverse type VI secretion system effectors in Vibrio cholerae

    PubMed Central

    Unterweger, Daniel; Kostiuk, Benjamin; Ötjengerdes, Rina; Wilton, Ashley; Diaz-Satizabal, Laura; Pukatzki, Stefan

    2015-01-01

    Vibrio cholerae is a diverse species of Gram-negative bacteria, commonly found in the aquatic environment and the causative agent of the potentially deadly disease cholera. These bacteria employ a type VI secretion system (T6SS) when they encounter prokaryotic and eukaryotic competitors. This contractile puncturing device translocates a set of effector proteins into neighboring cells. Translocated effectors are toxic unless the targeted cell produces immunity proteins that bind and deactivate incoming effectors. Comparison of multiple V. cholerae strains indicates that effectors are encoded in T6SS effector modules on mobile genetic elements. We identified a diverse group of chimeric T6SS adaptor proteins required for the translocation of diverse effectors encoded in modules. An example for a T6SS effector that requires T6SS adaptor protein 1 (Tap-1) is TseL found in pandemic V. cholerae O1 serogroup strains and other clinical isolates. We propose a model in which Tap-1 is required for loading TseL onto the secretion apparatus. After T6SS-mediated TseL export is completed, Tap-1 is retained in the bacterial cell to load other T6SS machines. PMID:26194724

  12. Chimeric adaptor proteins translocate diverse type VI secretion system effectors in Vibrio cholerae.

    PubMed

    Unterweger, Daniel; Kostiuk, Benjamin; Ötjengerdes, Rina; Wilton, Ashley; Diaz-Satizabal, Laura; Pukatzki, Stefan

    2015-08-13

    Vibrio cholerae is a diverse species of Gram-negative bacteria, commonly found in the aquatic environment and the causative agent of the potentially deadly disease cholera. These bacteria employ a type VI secretion system (T6SS) when they encounter prokaryotic and eukaryotic competitors. This contractile puncturing device translocates a set of effector proteins into neighboring cells. Translocated effectors are toxic unless the targeted cell produces immunity proteins that bind and deactivate incoming effectors. Comparison of multiple V. cholerae strains indicates that effectors are encoded in T6SS effector modules on mobile genetic elements. We identified a diverse group of chimeric T6SS adaptor proteins required for the translocation of diverse effectors encoded in modules. An example for a T6SS effector that requires T6SS adaptor protein 1 (Tap-1) is TseL found in pandemic V. cholerae O1 serogroup strains and other clinical isolates. We propose a model in which Tap-1 is required for loading TseL onto the secretion apparatus. After T6SS-mediated TseL export is completed, Tap-1 is retained in the bacterial cell to load other T6SS machines. PMID:26194724

  13. Space-based multifunctional end effector systems functional requirements and proposed designs

    NASA Technical Reports Server (NTRS)

    Mishkin, A. H.; Jau, B. M.

    1988-01-01

    The end effector is an essential element of teleoperator and telerobot systems to be employed in space in the next decade. The report defines functional requirements for end effector systems to perform operations that are currently only feasible through Extra-Vehicular Activity (EVA). Specific tasks and functions that the end effectors must be capable of performing are delineated. Required capabilities for forces and torques, clearances, compliance, and sensing are described, using current EVA requirements as guidelines where feasible. The implications of these functional requirements on the elements of potential end effector systems are discussed. The systems issues that must be considered in the design of space-based manipulator systems are identified; including impacts on subsystems tightly coupled to the end effector, i.e., control station, information processing, manipulator arm, tool and equipment stowage. Possible end effector designs are divided into three categories: single degree-of-freedom end effectors, multiple degree of freedom end effectors, and anthropomorphic hands. Specific design alternatives are suggested and analyzed within the individual categories. Two evaluations are performed: the first considers how well the individual end effectors could substitute for EVA; the second compares how manipulator systems composed of the top performers from the first evaluation would improve the space shuttle Remote Manipulator System (RMS) capabilities. The analysis concludes that the anthropomorphic hand is best-suited for EVA tasks. A left- and right-handed anthropomorphic manipulator arm configuration is suggested as appropriate to be affixed to the RMS, but could also be used as part of the Smart Front End for the Orbital Maneuvering Vehicle (OMV). The technical feasibility of the anthropomorphic hand and its control are demonstrated. An evolutionary development approach is proposed and approximate scheduling provided for implementing the suggested manipulator systems in time for space stations operations in the early 1990s.

  14. Explosion suppression system

    DOEpatents

    Sapko, Michael J. (Finleyville, PA); Cortese, Robert A. (Pittsburgh, PA)

    1992-01-01

    An explosion suppression system and triggering apparatus therefor are provided for quenching gas and dust explosions. An electrically actuated suppression mechanism which dispenses an extinguishing agent into the path ahead of the propagating flame is actuated by a triggering device which is light powered. This triggering device is located upstream of the propagating flame and converts light from the flame to an electrical actuation signal. A pressure arming device electrically connects the triggering device to the suppression device only when the explosion is sensed by a further characteristic thereof beside the flame such as the pioneer pressure wave. The light powered triggering device includes a solar panel which is disposed in the path of the explosion and oriented between horizontally downward and vertical. Testing mechanisms are also preferably provided to test the operation of the solar panel and detonator as well as the pressure arming mechanism.

  15. Immune modulation of effector CD4+ and regulatory T cell function by sorafenib in patients with hepatocellular carcinoma

    PubMed Central

    Ararat, Miguel; Xu, Yiling; Brusko, Todd; Wasserfall, Clive; Atkinson, Mark A.; Chang, Lung Ji; Liu, Chen; Nelson, David R.

    2013-01-01

    Hepatocellular carcinoma (HCC) is a difficult to treat cancer characterized by poor tumor immunity with only one approved systemic drug, sorafenib. If novel combination treatments are to be developed with immunological agents, the effects of sorafenib on tumor immunity are important to understand. In this study, we investigate the impact of sorafenib on the CD4+CD25? effector T cells (Teff) and CD4+CD25+ regulatory T cells (Tregs) from patients with HCC. We isolated Teff and Treg from peripheral mononuclear cells of HCC patients to determineimmune reactivity by thymidine incorporation, ELISA and flow cytometry. Teff cultured alone or with Treg were supplemented with different concentrations of sorafenib. The effects of sorafenib on Teff responses were dose-dependent. Pharmacologic doses of sorafenib decreased Teff activation by down regulating CD25 surface expression. In contrast, sub-pharmacologic concentrations of sorafenib resulted in Teff activation. These low doses of sorafenib in the Teff cultures led to a significant increase in Teff proliferation, IL2 secretion and up-regulation of CD25 expression on the cell surface. In addition, low doses of sorafenib in the suppression Teff/Treg cocultures restored Teff responses by eliminating Treg suppression. The loss of Treg suppressive function correlated with an increase in IL2 and IL6 secretion. Our findings showthat sub-pharmacologic doses of sorafenib impact subsets of T cells differently, selectively increasing Teff activation while blocking Treg function. In conclusion, this study describes novel immune activating properties of low doses of sorafenib by promoting immune responsiveness in patients with HCC. PMID:23223899

  16. Immune modulation of effector CD4+ and regulatory T cell function by sorafenib in patients with hepatocellular carcinoma.

    PubMed

    Cabrera, Roniel; Ararat, Miguel; Xu, Yiling; Brusko, Todd; Wasserfall, Clive; Atkinson, Mark A; Chang, Lung Ji; Liu, Chen; Nelson, David R

    2013-04-01

    Hepatocellular carcinoma (HCC) is a difficult to treat cancer characterized by poor tumor immunity with only one approved systemic drug, sorafenib. If novel combination treatments are to be developed with immunological agents, the effects of sorafenib on tumor immunity are important to understand. In this study, we investigate the impact of sorafenib on the CD4+CD25- effector T cells (Teff) and CD4+CD25+ regulatory T cells (Tregs) from patients with HCC. We isolated Teff and Treg from peripheral mononuclear cells of HCC patients to determine immune reactivity by thymidine incorporation, ELISA and flow cytometry. Teff cultured alone or with Treg were supplemented with different concentrations of sorafenib. The effects of sorafenib on Teff responses were dose-dependent. Pharmacologic doses of sorafenib decreased Teff activation by down regulating CD25 surface expression. In contrast, sub-pharmacologic concentrations of sorafenib resulted in Teff activation. These low doses of sorafenib in the Teff cultures led to a significant increase in Teff proliferation, IL2 secretion and up-regulation of CD25 expression on the cell surface. In addition, low doses of sorafenib in the suppression Teff/Treg cocultures restored Teff responses by eliminating Treg suppression. The loss of Treg suppressive function correlated with an increase in IL2 and IL6 secretion. Our findings show that sub-pharmacologic doses of sorafenib impact subsets of T cells differently, selectively increasing Teff activation while blocking Treg function. In conclusion, this study describes novel immune activating properties of low doses of sorafenib by promoting immune responsiveness in patients with HCC. PMID:23223899

  17. The Xanthomonas campestris pv. vesicatoria type III effector protein XopJ inhibits protein secretion: evidence for interference with cell wall-associated defense responses.

    PubMed

    Bartetzko, Verena; Sonnewald, Sophia; Vogel, Florian; Hartner, Kristina; Stadler, Ruth; Hammes, Ulrich Z; Börnke, Frederik

    2009-06-01

    The phytopathogenic bacterium Xanthomonas campestris pv. vesicatoria uses the type III secretion system (T3SS) to inject effector proteins into cells of its Solanaceous host plants. It is generally assumed that these effectors manipulate host pathways to favor bacterial replication and survival. However, the molecular mechanisms by which type III effectors suppress host defense responses are far from being understood. Based on sequence similarity, Xanthomonas outer protein J (XopJ) is a member of the YopJ/AvrRxv family of SUMO peptidases and acetyltranferases, although its biochemical activity has not yet been demonstrated. Confocal laser scanning microscopy revealed that green fluorescent protein (GFP) fusions of XopJ are targeted to the plasma membrane when expressed in plant cells, which most likely involves N-myristoylation. In contrast to a XopJ(C235A) mutant disrupted in the catalytic triad sequence, the wild-type effector GFP fusion protein was also localized in vesicle-like structures colocalizing together with a Golgi marker protein, suggesting an effect of XopJ on vesicle trafficking. To explore an effect of XopJ on protein secretion, we used a GFP-based secretion assay. When a secreted (sec)GFP marker was coexpressed with XopJ in leaves of Nicotiana benthamiana, GFP fluorescence was retained in reticulate structures. In contrast, in plant cells expressing secGFP alone or along with the XopJ(C235A) mutant, no GFP fluorescence accumulated within the cells. Moreover, coexpressing secGFP together with XopJ led to a reduced accumulation of secGFP within the apoplastic fluid of N. benthamiana leaves, further showing that XopJ affects protein secretion. Transgenic expression of XopJ in Arabidopsis suppressed callose deposition elicited by a T3SS-negative mutant of Pseudomonas syringae pv. tomato DC3000. A role of XopJ in the inhibition of cell wall-based defense responses is discussed. PMID:19445590

  18. Using the Kinect to limit abnormal kinematics and compensation strategies during therapy with end effector robots.

    PubMed

    Brokaw, Elizabeth B; Lum, Peter S; Cooper, Rory A; Brewer, Bambi R

    2013-06-01

    Abnormal kinematics and the use of compensation strategies during training limit functional improvement from therapy. The Kinect is a low cost ($100) sensor that does not require any markers to be placed on the user. Integration of this sensor into currently used therapy systems can provide feedback about the user's movement quality, and the use of compensatory strategies to complete tasks. This paper presents a novel technique of adding the Kinect to an end effector robot to limit compensation strategies and to train normal joint coordination during movements with an end effector robot. This methodology has wider implications for other robotic and passively actuated end effector rehabilitation devices. PMID:24187203

  19. Lessons from Anaplasma phagocytophilum: Chromatin Remodeling by Bacterial Effectors

    PubMed Central

    Rennoll-Bankert, Kristen E; Dumler, J Stephen

    2012-01-01

    Bacterial pathogens can alter global host gene expression via histone modifications and chromatin remodeling in order to subvert host responses, including those involved with innate immunity, allowing for bacterial survival. Shigella flexneri, Listeria monocytogenes, Chlamydia trachomatis, and Anaplasma phagocytophilum express effector proteins that modify host histones and chromatin structure. A. phagocytophilum modulates granulocyte respiratory burst in part by dampening transcription of several key phagocyte oxidase genes. The A. phagocytophilum protein AnkA localizes to the myeloid cell nucleus where it binds AT-rich regions in the CYBB promoter and decreases its transcription. AT-rich regions of DNA are characteristic of matrix attachment regions (MARs) which are critical for chromatin structure and transcription. MAR-binding proteins, such as SATB1, interact with histone modifying enzymes resulting in altered gene expression. With A. phagocytophilum infection, histone deacetylase 1 (HDAC1) expression is increased and histone H3 acetylation is decreased at the CYBB promoter, suggesting a role for AnkA in altering host epigenetics and modulating gene transcription, at this, and perhaps other loci. This review will focus on how bacterial pathogens alter host epigenetics, by specifically examining A. phagocytophilum AnkA cis-regulation of CYBB transcription and epigenetic changes associated with infection. PMID:23082961

  20. Receptor-coupled effector systems and their interactions

    SciTech Connect

    Wiener, E.C.

    1988-01-01

    We investigated the modulation of intracellular signal generation by receptor-coupled effector systems in B lymphocytes, and whether these alterations are consistent with the effects of prostaglandins. TPA (12-O-tetradecanoyl phorbol-13-acetate) and sn-1,2,-dioctanoylglycerol (diC{sub 8}) substitute for lipid derived signals which activate protein kinase C. Pretreating splenocytes from athymic nude mice with 100nM TPA or 5 {mu}M diC{sub 8} potentiated the forskolin-induced increased in cAMP (measured by radioimmunoassay) 2.5 and 3.0 times (respectively), but they decreased the PGE{sub 1}-induced cAMP rise 48% and 35% (respectively). Goat anti-mouse IgM, which activates diacylglycerol production, potentiated the forskolin-induced cAMP increase by 76%, but reduced that of PGE{sub 1} by 30%. Rabbit anti-mouse IgG, its F(ab{prime}){sub 2} fragment, or goat anti-mouse IGM induced increases in the cytosolic free (Ca{sup 2+}), (Ca{sup 2+}){sub i}, which TPA inhibited. In contrast, TPA potential antibody-induced {sup 3}H-thymidine (85x) and {sup 3}H-uridine (30x) uptake in B lymphocytes.

  1. Plasma Aerodynamic Control Effectors for Improved Wind Turbine Performance

    SciTech Connect

    Mehul P. Patel; Srikanth Vasudevan; Robert C. Nelson; Thomas C. Corke

    2008-08-01

    Orbital Research Inc is developing an innovative Plasma Aerodynamic Control Effectors (PACE) technology for improved performance of wind turbines. The PACE system is aimed towards the design of "smart" rotor blades to enhance energy capture and reduce aerodynamic loading and noise using flow-control. The PACE system will provide ability to change aerodynamic loads and pitch distribution across the wind turbine blade without any moving surfaces. Additional benefits of the PACE system include reduced blade structure weight and complexity that should translate into a substantially reduced initial cost. During the Phase I program, the ORI-UND Team demonstrated (proof-of-concept) performance improvements on select rotor blade designs using PACE concepts. Control of both 2-D and 3-D flows were demonstrated. An analytical study was conducted to estimate control requirements for the PACE system to maintain control during wind gusts. Finally, independent laboratory experiments were conducted to identify promising dielectric materials for the plasma actuator, and to examine environmental effects (water and dust) on the plasma actuator operation. The proposed PACE system will be capable of capturing additional energy, and reducing aerodynamic loading and noise on wind turbines. Supplementary benefits from the PACE system include reduced blade structure weight and complexity that translates into reduced initial capital costs.

  2. Mast cells as "tunable" effector and immunoregulatory cells: recent advances.

    PubMed

    Galli, Stephen J; Kalesnikoff, Janet; Grimbaldeston, Michele A; Piliponsky, Adrian M; Williams, Cara M M; Tsai, Mindy

    2005-01-01

    This review focuses on recent progress in our understanding of how mast cells can contribute to the initiation, development, expression, and regulation of acquired immune responses, both those associated with IgE and those that are apparently expressed independently of this class of Ig. We emphasize findings derived from in vivo studies in mice, particularly those employing genetic approaches to influence mast cell numbers and/or to alter or delete components of pathways that can regulate mast cell development, signaling, or function. We advance the hypothesis that mast cells not only can function as proinflammatory effector cells and drivers of tissue remodeling in established acquired immune responses, but also may contribute to the initiation and regulation of such responses. That is, we propose that mast cells can also function as immunoregulatory cells. Finally, we show that the notion that mast cells have primarily two functional configurations, off (or resting) or on (or activated for extensive mediator release), markedly oversimplifies reality. Instead, we propose that mast cells are "tunable," by both genetic and environmental factors, such that, depending on the circumstances, the cell can be positioned phenotypically to express a wide spectrum of variation in the types, kinetics, and/or magnitude of its secretory functions. PMID:15771585

  3. Molecular diversity of antimicrobial effectors in the oyster Crassostrea gigas

    PubMed Central

    2010-01-01

    Background To gain insight into the molecular diversity of antimicrobial peptides and proteins in the oyster Crassostrea gigas, we characterized and compared the sequence polymorphism of the antimicrobial peptides (AMPs), Cg-Defensins (Cg-Defs) and Cg-Proline Rich peptide (Cg-Prp), and of the bactericidal permeability increasing protein, Cg-BPI. For that, we analyzed genomic and transcript sequences obtained by specific PCR amplification and in silico searches. Results High diversification among the three antimicrobial effectors was evidenced by this polymorphism survey. On the basis of sequence phylogenies, each AMP aggregates into clearly defined groups of variants and is the product of a multigenic family displaying a variety of gene structures. In contrast, Cg-bpi forms a single group and is encoded by a single gene copy. Moreover, we identified for both AMPs several genetic mechanisms of diversification such as recombination, parallel mutations leading to phylogenetic homoplasy and indel events. In addition, the non synonymous to synonymous substitutions ratio by codon (dN/dS) revealed several negatively and positively selected sites for both AMPs, suggesting that directional selection pressures have shaped their sequence variations. Conclusions This study shows for the first time in a mollusc that antimicrobial peptides and proteins have been subject to distinct patterns of diversification and we evidence the existence of different evolutionary routes leading to such sequence variability. PMID:20100329

  4. Plasma suppression of beamstrahlung

    SciTech Connect

    Whittum, D.H.; Sessler, A.M.; Stewart, J.J.; Yu, S.S.

    1988-06-01

    We investigate the use of a plasma at the interaction point of two colliding beams to suppress beamsstrahlung and related phenomena. We derive conditions for good current cancellation via plasma return currents and report on numerical simulations conducted to confirm our analytic results. 10 refs., 5 figs., 4 tabs.

  5. Parasitic suppressing circuit

    NASA Technical Reports Server (NTRS)

    Fowler, J. T.; Raposa, F. L. (inventors)

    1973-01-01

    A circuit for suppressing parasitic oscillations across an inductor operating in a resonant mode is described. The circuit includes a switch means and resistive means connected serially across the inductor. A unidirectional resistive-capacitive network is also connected across the inductor and to the switch means to automatically render the switch means conducting when inductive current through the inductor ceases to flow.

  6. Legionella suppresses the host unfolded protein response via multiple mechanisms

    PubMed Central

    Treacy-Abarca, Sean; Mukherjee, Shaeri

    2015-01-01

    The intracellular pathogen, Legionella pneumophila, secretes ?300 effector proteins to modulate the host environment. Given the intimate interaction between L. pneumophila and the endoplasmic reticulum, we investigated the role of the host unfolded protein response (UPR) during L. pneumophila infection. Interestingly, we show that the host identifies L. pneumophila infection as a form of endoplasmic reticulum stress and the sensor pATF6 is processed to generate pATF6(N), a transcriptional activator of downstream UPR genes. However, L. pneumophila is able to suppress the UPR and block the translation of prototypical UPR genes, BiP and CHOP. Furthermore, biochemical studies reveal that L. pneumophila uses two effectors (Lgt1 and Lgt2) to inhibit the splicing of XBP1u mRNA to spliced XBP1 (XBP1s), an UPR response regulator. Thus, we demonstrate that L. pneumophila is able to inhibit the UPR by multiple mechanisms including blocking XBP1u splicing and causing translational repression. This observation highlights the utility of L. pneumophila as a powerful tool for studying a critical protein homeostasis regulator. PMID:26219498

  7. Regulation of vesicular trafficking and leukocyte function by Rab27 GTPases and their effectors

    PubMed Central

    Catz, Sergio Daniel

    2013-01-01

    The Rab27 family of GTPases regulates the efficiency and specificity of exocytosis in hematopoietic cells, including neutrophils, CTLs, NK cells, and mast cells. However, the mechanisms regulated by Rab27 GTPases are cell-specific, as they depend on the differential expression and function of particular effector molecules that are recruited by the GTPases. In addition, Rab27 GTPases participate in multiple steps of the regulation of the secretory process, including priming, tethering, docking, and fusion through sequential interaction with multiple effector molecules. Finally, recent reports suggest that Rab27 GTPases and their effectors regulate vesicular trafficking mechanisms other than exocytosis, including endocytosis and phagocytosis. This review focuses on the latest discoveries on the function of Rab27 GTPases and their effectors Munc13-4 and Slp1 in neutrophil function comparatively to their functions in other leukocytes. PMID:23378593

  8. Programmable Sequence-Specific Transcriptional Regulation of Mammalian Genome Using Designer TAL Effectors

    E-print Network

    Zhang, Feng

    The ability to direct functional proteins to specific DNA sequences is a long-sought goal in the study and engineering of biological processes. Transcription activator–like effectors (TALEs) from Xanthomonas sp. are ...

  9. From GFP to ?-lactamase: advancing intact cell imaging for toxins and effectors.

    PubMed

    Zuverink, Madison; Barbieri, Joseph T

    2015-12-01

    Canonical reporters such as green fluorescent protein (GFP) and luciferase have assisted researchers in probing cellular pathways and processes. Prior research in pathogenesis depended on sensitivity of biochemical and biophysical techniques to identify effectors and elucidate entry mechanisms. Recently, the ?-lactamase (?lac) reporter system has advanced toxin and effector reporting by permitting measurement of ?lac delivery into the cytosol or host ?lac expression in intact cells. ?lac measurement in cells was facilitated by the development of the fluorogenic substrate, CCF2-AM, to identify novel effectors, target cells, and domains involved in bacterial pathogenesis. The assay is also adaptable for high-throughput screening of small molecule inhibitors against toxins, providing information on mechanism and potential therapeutic agents. The versatility and limitations of the ?lac reporter system as applied to toxins and effectors are discussed in this review. PMID:26500183

  10. End-effector: Joint conjugates for robotic assembly of large truss structures in space: Extended concepts

    NASA Technical Reports Server (NTRS)

    Brewer, W. V.; Rasis, E. P.; Shih, H. R.

    1993-01-01

    Results from NASA/HBCU Grant No. NAG-1-1125 are summarized. Designs developed for model fabrication, exploratory concepts drafted, interface of computer with robot and end-effector, and capability enhancement are discussed.

  11. THE ENTEROHEMORRHAGIC ESCHERICHIA COLI EFFECTOR PROTEIN NLEF BINDS MAMMALIAN HOST PROTEINS

    E-print Network

    Olsen, Rachel Lee

    2012-12-31

    The extracellular human pathogens enterohemorrhagic and enteropathogenic Escherichia coli (EHEC and EPEC) and the related mouse pathogen Citrobacter rodentium inject type III secretion system (T3SS) effector proteins to promote their replication...

  12. Elucidating the role of effector caspases in immune development using lentiviral RNAi

    E-print Network

    Dillon, Christopher P

    2006-01-01

    Caspases play an important role in apoptosis, or programmed cell death. In particular, three highly related effector caspases, caspases-3, -6, and -7, translate upstream death signals into the physical manifestations of ...

  13. Intrathymic programming of effector fates in three molecularly distinct ?? T cell subtypes

    E-print Network

    Narayan, Kavitha

    Innate ?? T cells function in the early phase of immune responses. Although innate ?? T cells have often been studied as one homogenous population, they can be functionally classified into effector subsets on the basis of ...

  14. Analysis of secreted proteins of Magnaporthe grisea and the search for protein effectors 

    E-print Network

    Shang, Yue

    2007-09-17

    Magnaporthe grisea is a notorious pathogenic fungus that causes rice blast disease worldwide. Proteins secreted by the fungus are likely candidates for being effectors that are potentially recognized by determinants of ...

  15. A Chlamydia effector recruits CEP170 to reprogram host microtubule organization.

    PubMed

    Dumoux, Maud; Menny, Anais; Delacour, Delphine; Hayward, Richard D

    2015-09-15

    The obligate intracellular bacterial pathogen Chlamydia trachomatis deploys virulence effectors to subvert host cell functions enabling its replication within a specialized membrane-bound compartment termed an inclusion. The control of the host cytoskeleton is crucial for Chlamydia uptake, inclusion biogenesis and cell exit. Here, we demonstrate how a Chlamydia effector rearranges the microtubule (MT) network by initiating organization of the MTs at the inclusion surface. We identified an inclusion-localized effector that is sufficient to interfere with MT assembly, which we named inclusion protein acting on MTs (IPAM). We established that IPAM recruits and stimulates the centrosomal protein 170?kDa (CEP170) to hijack the MT organizing functions of the host cell. We show that CEP170 is essential for chlamydial control of host MT assembly, and is required for inclusion morphogenesis and bacterial infectivity. Together, we demonstrate how a pathogen effector reprograms the host MT network to support its intracellular development. PMID:26220855

  16. A Chlamydia effector recruits CEP170 to reprogram host microtubule organization

    PubMed Central

    Dumoux, Maud; Menny, Anais; Delacour, Delphine; Hayward, Richard D.

    2015-01-01

    ABSTRACT The obligate intracellular bacterial pathogen Chlamydia trachomatis deploys virulence effectors to subvert host cell functions enabling its replication within a specialized membrane-bound compartment termed an inclusion. The control of the host cytoskeleton is crucial for Chlamydia uptake, inclusion biogenesis and cell exit. Here, we demonstrate how a Chlamydia effector rearranges the microtubule (MT) network by initiating organization of the MTs at the inclusion surface. We identified an inclusion-localized effector that is sufficient to interfere with MT assembly, which we named inclusion protein acting on MTs (IPAM). We established that IPAM recruits and stimulates the centrosomal protein 170?kDa (CEP170) to hijack the MT organizing functions of the host cell. We show that CEP170 is essential for chlamydial control of host MT assembly, and is required for inclusion morphogenesis and bacterial infectivity. Together, we demonstrate how a pathogen effector reprograms the host MT network to support its intracellular development. PMID:26220855

  17. Structural consequences of effector protein complex formation in a diiron hydroxylase

    SciTech Connect

    Bailey, Lucas J.; McCoy, Jason G.; Phillips, Jr., George N.; Fox, Brian G.

    2009-06-12

    Carboxylate-bridged diiron hydroxylases are multicomponent enzyme complexes responsible for the catabolism of a wide range of hydrocarbons and as such have drawn attention for their mechanism of action and potential uses in bioremediation and enzymatic synthesis. These enzyme complexes use a small molecular weight effector protein to modulate the function of the hydroxylase. However, the origin of these functional changes is poorly understood. Here, we report the structures of the biologically relevant effector protein-hydroxylase complex of toluene 4-monooxygenase in 2 redox states. The structures reveal a number of coordinated changes that occur up to 25 {angstrom} from the active site and poise the diiron center for catalysis. The results provide a structural basis for the changes observed in a number of the measurable properties associated with effector protein binding. This description provides insight into the functional role of effector protein binding in all carboxylate-bridged diiron hydroxylases.

  18. Lifestyles of the effector-rich: genome-enabled characterization of bacterial plant pathogens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genome sequencing of bacterial plant pathogens is providing transformative insights into the complex network of molecular plant-microbe interactions mediated by extracellular effectors during pathogenesis. Bacterial pathogens sequenced to completion are phylogenetically diverse and vary significant...

  19. Intraspecies Competition in Serratia marcescens Is Mediated by Type VI-Secreted Rhs Effectors and a Conserved Effector-Associated Accessory Protein

    PubMed Central

    Alcoforado Diniz, Juliana

    2015-01-01

    ABSTRACT The type VI secretion system (T6SS) is widespread in Gram-negative bacteria and can deliver toxic effector proteins into eukaryotic cells or competitor bacteria. Antibacterial T6SSs are increasingly recognized as key mediators of interbacterial competition and may contribute to the outcome of many polymicrobial infections. Multiple antibacterial effectors can be delivered by these systems, with diverse activities against target cells and distinct modes of secretion. Polymorphic toxins containing Rhs repeat domains represent a recently identified and as-yet poorly characterized class of T6SS-dependent effectors. Previous work had revealed that the potent antibacterial T6SS of the opportunistic pathogen Serratia marcescens promotes intraspecies as well as interspecies competition (S. L. Murdoch, K. Trunk, G. English, M. J. Fritsch, E. Pourkarimi, and S. J. Coulthurst, J Bacteriol 193:6057–6069, 2011, http://dx.doi.org/10.1128/JB.05671-11). In this study, two new Rhs family antibacterial effectors delivered by this T6SS have been identified. One of these was shown to act as a DNase toxin, while the other contains a novel, cytoplasmic-acting toxin domain. Importantly, using S. marcescens, it has been demonstrated for the first time that Rhs proteins, rather than other T6SS-secreted effectors, can be the primary determinant of intraspecies competition. Furthermore, a new family of accessory proteins associated with T6SS effectors has been identified, exemplified by S. marcescens EagR1, which is specifically required for deployment of its associated Rhs effector. Together, these findings provide new insight into how bacteria can use the T6SS to deploy Rhs-family effectors and mediate different types of interbacterial interactions. IMPORTANCE Infectious diseases caused by bacterial pathogens represent a continuing threat to health and economic prosperity. To counter this threat, we must understand how such organisms survive and prosper. The type VI secretion system is a weapon that many pathogens deploy to compete against rival bacterial cells by injecting multiple antibacterial toxins into them. The ability to compete is vital considering that bacteria generally live in mixed communities. We aimed to identify new toxins and understand their deployment and role in interbacterial competition. We describe two new type VI secretion system-delivered toxins of the Rhs class, demonstrate that this class can play a primary role in competition between closely related bacteria, and identify a new accessory factor needed for their delivery. PMID:25939831

  20. Glioma associated cancer-initiating cells induce immune suppression

    PubMed Central

    Wei, Jun; Barr, Jason; Kong, Ling-Yuan; Wang, Yongtao; Wu, Adam; Sharma, Amit K.; Gumin, Joy; Henry, Verlene; Colman, Howard; Sawaya, Raymond; Lang, Frederick F.; Heimberger, Amy B.

    2010-01-01

    Purpose Glioblastoma multiforme (GBM) is a lethal cancer that responds poorly to therapy. GBM cancer-initiating cells have been shown to mediate resistance to both chemotherapy and radiation; however, it is unknown to what extent these cells contribute to the profound immune suppression in GBM patients and if strategies that alter their differentiation state can reduce this immune suppression. Experimental Design We isolated a subpopulation of cells from GBMs that possessed the capacity for self-renewal, formed neurospheres in vitro, were capable of pluripotent differentiation and could initiate tumors in vivo. These cells immune phenotype was characterized including the elaboration of immunosuppressive cytokines and chemokines by enzyme-linked immunosorbent assay. Functional immunosuppressive properties were characterized based on the inhibition of T cell proliferation and effector responses, triggering of T cell apoptosis and the induction FoxP3+ regulatory T cells. Upon altering their differentiation state, the immune suppressive phenotype and functional assays were reevaluated. Results We found that the cancer-initiating cells markedly inhibited T cell proliferation and activation, induced regulatory T cells and triggered T cell apoptosis that were mediated by B7-H1 and soluble Galectin-3. These immunosuppressive properties were diminished upon altering the differentiation of the cancer-initiating cells. Conclusion Cancer-initiating cells contribute to tumor evasion of the immune surveillance and approaches that alter the differentiation state may have immune therapeutic potential. PMID:20068105

  1. Effector-Mining in the Poplar Rust Fungus Melampsora larici-populina Secretome

    PubMed Central

    Lorrain, Cécile; Hecker, Arnaud; Duplessis, Sébastien

    2015-01-01

    The poplar leaf rust fungus, Melampsora larici-populina has been established as a tree-microbe interaction model. Understanding the molecular mechanisms controlling infection by pathogens appears essential for durable management of tree plantations. In biotrophic plant-parasites, effectors are known to condition host cell colonization. Thus, investigation of candidate secreted effector proteins (CSEPs) is a major goal in the poplar–poplar rust interaction. Unlike oomycetes, fungal effectors do not share conserved motifs and candidate prediction relies on a set of a priori criteria established from reported bona fide effectors. Secretome prediction, genome-wide analysis of gene families and transcriptomics of M. larici-populina have led to catalogs of more than a thousand secreted proteins. Automatized effector-mining pipelines hold great promise for rapid and systematic identification and prioritization of CSEPs for functional characterization. In this review, we report on and discuss the current status of the poplar rust fungus secretome and prediction of candidate effectors from this species. PMID:26697026

  2. A Salmonella type three secretion effector/chaperone complex adopts a hexameric ring-like structure.

    PubMed

    Roblin, Pierre; Dewitte, Frédérique; Villeret, Vincent; Biondi, Emanuele G; Bompard, Coralie

    2015-02-15

    Many bacterial pathogens use type three secretion systems (T3SS) to inject virulence factors, named effectors, directly into the cytoplasm of target eukaryotic cells. Most of the T3SS components are conserved among plant and animal pathogens, suggesting a common mechanism of recognition and secretion of effectors. However, no common motif has yet been identified for effectors allowing T3SS recognition. In this work, we performed a biochemical and structural characterization of the Salmonella SopB/SigE chaperone/effector complex by small-angle X-ray scattering (SAXS). Our results showed that the SopB/SigE complex is assembled in dynamic homohexameric-ring-shaped structures with an internal tunnel. In this ring, the chaperone maintains a disordered N-terminal end of SopB molecules, in a good position to be reached and processed by the T3SS. This ring dimensionally fits the ring-organized molecules of the injectisome, including ATPase hexameric rings; this organization suggests that this structural feature is important for ATPase recognition by T3SS. Our work constitutes the first evidence of the oligomerization of an effector, analogous to the organization of the secretion machinery, obtained in solution. As effectors share neither sequence nor structural identity, the quaternary oligomeric structure could constitute a strategy evolved to promote the specificity and efficiency of T3SS recognition. PMID:25404693

  3. Proteogenomic analysis of the Venturia pirina (Pear Scab Fungus) secretome reveals potential effectors.

    PubMed

    Cooke, Ira R; Jones, Dan; Bowen, Joanna K; Deng, Cecilia; Faou, Pierre; Hall, Nathan E; Jayachandran, Vignesh; Liem, Michael; Taranto, Adam P; Plummer, Kim M; Mathivanan, Suresh

    2014-08-01

    A proteogenomic analysis is presented for Venturia pirina, a fungus that causes scab disease on European pear (Pyrus communis). V. pirina is host-specific, and the infection is thought to be mediated by secreted effector proteins. Currently, only 36 V. pirina proteins are catalogued in GenBank, and the genome sequence is not publicly available. To identify putative effectors, V. pirina was grown in vitro on and in cellophane sheets mimicking its growth in infected leaves. Secreted extracts were analyzed by tandem mass spectrometry, and the data (ProteomeXchange identifier PXD000710) was queried against a protein database generated by combining in silico predicted transcripts with six frame translations of a whole genome sequence of V. pirina (GenBank Accession JEMP00000000 ). We identified 1088 distinct V. pirina protein groups (FDR 1%) including 1085 detected for the first time. Thirty novel (not in silico predicted) proteins were found, of which 14 were identified as potential effectors based on characteristic features of fungal effector protein sequences. We also used evidence from semitryptic peptides at the protein N-terminus to corroborate in silico signal peptide predictions for 22 proteins, including several potential effectors. The analysis highlights the utility of proteogenomics in the study of secreted effectors. PMID:24965097

  4. Spatiotemporal dynamics of effector CD8+ T cell responses within the liver.

    PubMed

    Inverso, Donato; Iannacone, Matteo

    2016-01-01

    CD8(+) T cells play a critical role in controlling hepatotropic viral infections, such as those caused by hepatitis B and hepatitis C viruses. The capacity of these cells to protect against such pathogens is mediated by antigen-experienced effector cells and relies on their ability to home to the liver, recognize pathogen-derived antigens, and deploy effector functions. Here, we review how dynamic imaging of hepatic effector CD8(+) T cell migration and function in mouse models of hepatitis B virus pathogenesis has recently revealed a unique and novel mode of adaptive immune surveillance. Circulating effector CD8(+) T cells initially arrest within liver sinusoids by docking onto adherent platelets and then actively crawl along the liver vasculature, probing hepatocytes for the presence of antigens by extending protrusions through the fenestrated sinusoidal endothelial cells. Hepatocellular antigen recognition and effector functions occur while CD8(+) T cells are still confined to the intravascular space and are inhibited by the pathologic processes that characterize liver fibrosis. A detailed understanding of the spatiotemporal dynamics of effector CD8(+) T cells within the liver is important for the rational design of targeted immunotherapeutic approaches for chronic liver infections. PMID:26188075

  5. Arabidopsis TAO1 is a TIR-NB-LRR protein that contributes to disease resistance induced by the Pseudomonas syringae effector AvrB.

    PubMed

    Eitas, Timothy K; Nimchuk, Zachary L; Dangl, Jeffery L

    2008-04-29

    The type III effector protein encoded by avirulence gene B (AvrB) is delivered into plant cells by pathogenic strains of Pseudomonas syringae. There, it localizes to the plasma membrane and triggers immunity mediated by the Arabidopsis coiled-coil (CC)-nucleotide binding (NB)-leucine-rich repeat (LRR) disease resistance protein RPM1. The sequence unrelated type III effector avirulence protein encoded by avirulence gene Rpm1 (AvrRpm1) also activates RPM1. AvrB contributes to virulence after delivery from P. syringae in leaves of susceptible soybean plants, and AvrRpm1 does the same in Arabidopsis rpm1 plants. Conditional overexpression of AvrB in rpm1 plants results in leaf chlorosis. In a genetic screen for mutants that lack AvrB-dependent chlorosis in an rpm1 background, we isolated TAO1 (target of AvrB operation), which encodes a Toll-IL-1 receptor (TIR)-NB-LRR disease resistance protein. In rpm1 plants, TAO1 function results in the expression of the pathogenesis-related protein 1 (PR-1) gene, suggestive of a defense response. In RPM1 plants, TAO1 contributes to disease resistance in response to Pto (P. syringae pathovars tomato) DC3000(avrB), but not against Pto DC3000(avrRpm1). The tao1-5 mutant allele, a stop mutation in the LRR domain of TAO1, posttranscriptionally suppresses RPM1 accumulation. These data provide evidence of genetically separable disease resistance responses to AvrB and AvrRpm1 in Arabidopsis. AvrB activates both RPM1, a CC-NB-LRR protein, and TAO1, a TIR-NB-LRR protein. These NB-LRR proteins then act additively to generate a full disease resistance response to P. syringae expressing this type III effector. PMID:18424557

  6. Pressure suppression containment system

    DOEpatents

    Gluntz, D.M.; Townsend, H.E.

    1994-03-15

    A pressure suppression containment system includes a containment vessel surrounding a reactor pressure vessel and defining a drywell therein containing a non-condensable gas. An enclosed wetwell pool is disposed inside the containment vessel, and a gravity driven cooling system (GDCS) pool is disposed above the wetwell pool in the containment vessel. The wetwell pool includes a plenum for receiving the non-condensable gas carried with steam from the drywell following a loss-of-coolant-accident (LOCA). The wetwell plenum is vented to a plenum above the GDCS pool following the LOCA for suppressing pressure rise within the containment vessel. A method of operation includes channeling steam released into the drywell following the LOCA into the wetwell pool for cooling along with the non-condensable gas carried therewith. The GDCS pool is then drained by gravity, and the wetwell plenum is vented into the GDCS plenum for channeling the non-condensable gas thereto. 6 figures.

  7. Pressure suppression containment system

    DOEpatents

    Gluntz, Douglas M. (San Jose, CA); Townsend, Harold E. (San Jose, CA)

    1994-03-15

    A pressure suppression containment system includes a containment vessel surrounding a reactor pressure vessel and defining a drywell therein containing a non-condensable gas. An enclosed wetwell pool is disposed inside the containment vessel, and a gravity driven cooling system (GDCS) pool is disposed above the wetwell pool in the containment vessel. The wetwell pool includes a plenum for receiving the non-condensable gas carried with steam from the drywell following a loss-of coolant-accident (LOCA). The wetwell plenum is vented to a plenum above the GDCS pool following the LOCA for suppressing pressure rise within the containment vessel. A method of operation includes channeling steam released into the drywell following the LOCA into the wetwell pool for cooling along with the non-condensable gas carried therewith. The GDCS pool is then drained by gravity, and the wetwell plenum is vented into the GDCS plenum for channeling the non-condensable gas thereto.

  8. Naturally Occurring Nonpathogenic Isolates of the Plant Pathogen Pseudomonas syringae Lack a Type III Secretion System and Effector Gene Orthologues? †

    PubMed Central

    Mohr, Toni J.; Liu, Haijie; Yan, Shuangchun; Morris, Cindy E.; Castillo, José A.; Jelenska, Joanna; Vinatzer, Boris A.

    2008-01-01

    Pseudomonas syringae causes plant diseases, and the main virulence mechanism is a type III secretion system (T3SS) that translocates dozens of effector proteins into plant cells. Here we report the existence of a subgroup of P. syringae isolates that do not cause disease on any plant species tested. This group is monophyletic and most likely evolved from a pathogenic P. syringae ancestor through loss of the T3SS. In the nonpathogenic isolate P. syringae 508 the genomic region that in pathogenic P. syringae strains contains the hrp-hrc cluster coding for the T3SS and flanking effector genes is absent. P. syringae 508 was also surveyed for the presence of effector orthologues from the closely related pathogenic strain P. syringae pv. syringae B728a, but none were detected. The absence of the hrp-hrc cluster and effector orthologues was confirmed for other nonpathogenic isolates. Using the AvrRpt2 effector as reporter revealed the inability of P. syringae 508 to translocate effectors into plant cells. Adding a plasmid-encoded T3SS and the P. syringae pv. syringae 61 effector gene hopA1 increased in planta growth almost 10-fold. This suggests that P. syringae 508 supplemented with a T3SS could be used to determine functions of individual effectors in the context of a plant infection, avoiding the confounding effect of other effectors with similar functions present in effector mutants of pathogenic isolates. PMID:18263729

  9. Nonsense suppression in archaea.

    PubMed

    Bhattacharya, Arpita; Köhrer, Caroline; Mandal, Debabrata; RajBhandary, Uttam L

    2015-05-12

    Bacterial strains carrying nonsense suppressor tRNA genes played a crucial role in early work on bacterial and bacterial viral genetics. In eukaryotes as well, suppressor tRNAs have played important roles in the genetic analysis of yeast and worms. Surprisingly, little is known about genetic suppression in archaea, and there has been no characterization of suppressor tRNAs or identification of nonsense mutations in any of the archaeal genes. Here, we show, using the ?-gal gene as a reporter, that amber, ochre, and opal suppressors derived from the serine and tyrosine tRNAs of the archaeon Haloferax volcanii are active in suppression of their corresponding stop codons. Using a promoter for tRNA expression regulated by tryptophan, we also show inducible and regulatable suppression of all three stop codons in H. volcanii. Additionally, transformation of a ?pyrE2 H. volcanii strain with plasmids carrying the genes for a pyrE2 amber mutant and the serine amber suppressor tRNA yielded transformants that grow on agar plates lacking uracil. Thus, an auxotrophic amber mutation in the pyrE2 gene can be complemented by expression of the amber suppressor tRNA. These results pave the way for generating archaeal strains carrying inducible suppressor tRNA genes on the chromosome and their use in archaeal and archaeviral genetics. We also provide possible explanations for why suppressor tRNAs have not been identified in archaea. PMID:25918386

  10. Nonsense suppression in archaea

    PubMed Central

    Bhattacharya, Arpita; Köhrer, Caroline; Mandal, Debabrata; RajBhandary, Uttam L.

    2015-01-01

    Bacterial strains carrying nonsense suppressor tRNA genes played a crucial role in early work on bacterial and bacterial viral genetics. In eukaryotes as well, suppressor tRNAs have played important roles in the genetic analysis of yeast and worms. Surprisingly, little is known about genetic suppression in archaea, and there has been no characterization of suppressor tRNAs or identification of nonsense mutations in any of the archaeal genes. Here, we show, using the ?-gal gene as a reporter, that amber, ochre, and opal suppressors derived from the serine and tyrosine tRNAs of the archaeon Haloferax volcanii are active in suppression of their corresponding stop codons. Using a promoter for tRNA expression regulated by tryptophan, we also show inducible and regulatable suppression of all three stop codons in H. volcanii. Additionally, transformation of a ?pyrE2 H. volcanii strain with plasmids carrying the genes for a pyrE2 amber mutant and the serine amber suppressor tRNA yielded transformants that grow on agar plates lacking uracil. Thus, an auxotrophic amber mutation in the pyrE2 gene can be complemented by expression of the amber suppressor tRNA. These results pave the way for generating archaeal strains carrying inducible suppressor tRNA genes on the chromosome and their use in archaeal and archaeviral genetics. We also provide possible explanations for why suppressor tRNAs have not been identified in archaea. PMID:25918386

  11. Structure of the Type VI Effector-Immunity Complex (Tae4-Tai4) Provides Novel Insights into the Inhibition Mechanism of the Effector by Its Immunity Protein*

    PubMed Central

    Zhang, Heng; Zhang, Heng; Gao, Zeng-Qiang; Wang, Wen-Jia; Liu, Guang-Feng; Xu, Jian-Hua; Su, Xiao-Dong; Dong, Yu-Hui

    2013-01-01

    The type VI secretion system (T6SS), a multisubunit needle-like apparatus, has recently been found to play a role in interspecies interactions. The Gram-negative bacteria harboring T6SS (donor) deliver the effectors into their neighboring cells (recipient) to kill them. Meanwhile, the cognate immunity proteins were employed to protect the donor cells against the toxic effectors. Tae4 (type VI amidase effector 4) and Tai4 (type VI amidase immunity 4) are newly identified T6SS effector-immunity pairs. Here, we report the crystal structures of Tae4 from Enterobacter cloacae and Tae4-Tai4 complexes from both E. cloacae and Salmonella typhimurium. Tae4 acts as a dl-endopeptidase and displays a typical N1pC/P60 domain. Unlike Tsi1 (type VI secretion immunity 1), Tai4 is an all-helical protein and forms a dimer in solution. The small angle x-ray scattering study combined with the analytical ultracentrifugation reveal that the Tae4-Tai4 complex is a compact heterotetramer that consists of a Tai4 dimer and two Tae4 molecules in solution. Structure-based mutational analysis of the Tae4-Tai4 interface shows that a helix (?3) of one subunit in dimeric Tai4 plays a major role in binding of Tae4, whereas a protruding loop (L4) in the other subunit is mainly responsible for inhibiting Tae4 activity. The inhibition process requires collaboration between the Tai4 dimer. These results reveal a novel and unique inhibition mechanism in effector-immunity pairs and suggest a new strategy to develop antipathogen drugs. PMID:23288853

  12. A bacterial type III secretion assay for delivery of fungal effector proteins into wheat.

    PubMed

    Upadhyaya, Narayana M; Mago, Rohit; Staskawicz, Brian J; Ayliffe, Michael A; Ellis, Jeffrey G; Dodds, Peter N

    2014-03-01

    Large numbers of candidate effectors from fungal pathogens are being identified through whole-genome sequencing and in planta expression studies. Although Agrobacterium-mediated transient expression has enabled high-throughput functional analysis of effectors in dicot plants, this assay is not effective in cereal leaves. Here, we show that a nonpathogenic Pseudomonas fluorescens engineered to express the type III secretion system (T3SS) of P. syringae and the wheat pathogen Xanthomonas translucens can deliver fusion proteins containing T3SS signals from P. syringae (AvrRpm1) and X. campestris (AvrBs2) avirulence (Avr) proteins, respectively, into wheat leaf cells. A calmodulin-dependent adenylate cyclase reporter protein was delivered effectively into wheat and barley by both bacteria. Absence of any disease symptoms with P. fluorescens makes it more suitable than X. translucens for detecting a hypersensitive response (HR) induced by an effector protein with avirulence activity. We further modified the delivery system by removal of the myristoylation site from the AvrRpm1 fusion to prevent its localization to the plasma membrane which could inhibit recognition of an Avr protein. Delivery of the flax rust AvrM protein by the modified delivery system into transgenic tobacco leaves expressing the corresponding M resistance protein induced a strong HR, indicating that the system is capable of delivering a functional rust Avr protein. In a preliminary screen of effectors from the stem rust fungus Puccinia graminis f. sp. tritici, we identified one effector that induced a host genotype-specific HR in wheat. Thus, the modified AvrRpm1:effector-Pseudomonas fluorescens system is an effective tool for large-scale screening of pathogen effectors for recognition in wheat. PMID:24156769

  13. A single plant resistance gene promoter engineered to recognize multiple TAL effectors from disparate pathogens.

    PubMed

    Römer, Patrick; Recht, Sabine; Lahaye, Thomas

    2009-12-01

    Plant pathogenic bacteria of the genus Xanthomonas inject transcription-activator like (TAL) effector proteins that manipulate the hosts' transcriptome to promote disease. However, in some cases plants take advantage of this mechanism to trigger defense responses. For example, transcription of the pepper Bs3 and rice Xa27 resistance (R) genes are specifically activated by the respective TAL effectors AvrBs3 from Xanthomonas campestris pv. vesicatoria (Xcv), and AvrXa27 from X. oryzae pv. oryzae (Xoo). Recognition of AvrBs3 was shown to be mediated by interaction with the corresponding UPT (UPregulated by TAL effectors) box UPT(AvrBs3) present in the promoter R gene Bs3 from the dicot pepper. In contrast, it was not known how the Xoo TAL effector AvrXa27 transcriptionally activates the matching R gene Xa27 from the monocot rice. Here we identified a 16-bp UPT(AvrXa27) box present in the rice Xa27 promoter that when transferred into the Bs3 promoter confers AvrXa27-dependent inducibility. We demonstrate that polymorphisms between the UPT(AvrXa27) box of the AvrXa27-inducible Xa27 promoter and the corresponding region of the noninducible xa27 promoter account for their distinct inducibility and affinity, with respect to AvrXa27. Moreover, we demonstrate that three functionally distinct UPT boxes targeted by separate TAL effectors retain their function and specificity when combined into one promoter. Given that many economically important xanthomonads deliver multiple TAL effectors, the engineering of R genes capable of recognizing multiple TAL effectors provides a potential approach for engineering broad spectrum and durable disease resistance. PMID:19910532

  14. Quantitative Proteomic Analysis of Burkholderia pseudomallei Bsa Type III Secretion System Effectors Using Hypersecreting Mutants

    PubMed Central

    Vander Broek, Charles W.; Chalmers, Kevin J.; Stevens, Mark P.; Stevens, Joanne M.

    2015-01-01

    Burkholderia pseudomallei is an intracellular pathogen and the causative agent of melioidosis, a severe disease of humans and animals. One of the virulence factors critical for early stages of infection is the Burkholderia secretion apparatus (Bsa) Type 3 Secretion System (T3SS), a molecular syringe that injects bacterial proteins, called effectors, into eukaryotic cells where they subvert cellular functions to the benefit of the bacteria. Although the Bsa T3SS itself is known to be important for invasion, intracellular replication, and virulence, only a few genuine effector proteins have been identified and the complete repertoire of proteins secreted by the system has not yet been fully characterized. We constructed a mutant lacking bsaP, a homolog of the T3SS “gatekeeper” family of proteins that exert control over the timing and magnitude of effector protein secretion. Mutants lacking BsaP, or the T3SS translocon protein BipD, were observed to hypersecrete the known Bsa effector protein BopE, providing evidence of their role in post-translational control of the Bsa T3SS and representing key reagents for the identification of its secreted substrates. Isobaric Tags for Relative and Absolute Quantification (iTRAQ), a gel-free quantitative proteomics technique, was used to compare the secreted protein profiles of the Bsa T3SS hypersecreting mutants of B. pseudomallei with the isogenic parent strain and a bsaZ mutant incapable of effector protein secretion. Our study provides one of the most comprehensive core secretomes of B. pseudomallei described to date and identified 26 putative Bsa-dependent secreted proteins that may be considered candidate effectors. Two of these proteins, BprD and BapA, were validated as novel effector proteins secreted by the Bsa T3SS of B. pseudomallei. PMID:25635268

  15. Role of Regulatory T Cells (Treg) and the Treg Effector Molecule Fibrinogen-like Protein 2 in Alloimmunity and Autoimmunity

    PubMed Central

    Chruscinski, Andrzej; Sadozai, Hassan; Rojas-Luengas, Vanessa; Bartczak, Agata; Khattar, Ramzi; Selzner, Nazia; Levy, Gary A.

    2015-01-01

    CD4+CD25+Foxp3+ regulatory T cells (Treg) are critical to the maintenance of immune tolerance. Treg are known to utilize a number of molecular pathways to control immune responses and maintain immune homeostasis. Fibrinogen-like protein 2 (FGL2) has been identified by a number of investigators as an important immunosuppressive effector of Treg, which exerts its immunoregulatory activity by binding to inhibitory Fc?RIIB receptors expressed on antigen-presenting cells including dendritic cells, endothelial cells, and B cells. More recently, it has been suggested that FGL2 accounts for the immunosuppressive activity of a highly suppressive subset of Treg that express T cell immunoreceptor with Ig and ITIM domains (TIGIT). Here we discuss the important role of Treg and FGL2 in preventing alloimmune and autoimmune disease. The FGL2–Fc?RIIB pathway is also known to be utilized by viruses and tumor cells to evade immune surveillance. Moving forward, therapies based on modulation of the FGL2–Fc?RIIB pathway hold promise for the treatment of a wide variety of conditions ranging from autoimmunity to cancer. PMID:26241231

  16. DEEP--a tool for differential expression effector prediction.

    PubMed

    Degenhardt, Jost; Haubrock, Martin; Dönitz, Jürgen; Wingender, Edgar; Crass, Torsten

    2007-07-01

    High-throughput methods for measuring transcript abundance, like SAGE or microarrays, are widely used for determining differences in gene expression between different tissue types, dignities (normal/malignant) or time points. Further analysis of such data frequently aims at the identification of gene interaction networks that form the causal basis for the observed properties of the systems under examination. To this end, it is usually not sufficient to rely on the measured gene expression levels alone; rather, additional biological knowledge has to be taken into account in order to generate useful hypotheses about the molecular mechanism leading to the realization of a certain phenotype. We present a method that combines gene expression data with biological expert knowledge on molecular interaction networks, as described by the TRANSPATH database on signal transduction, to predict additional--and not necessarily differentially expressed--genes or gene products which might participate in processes specific for either of the examined tissues or conditions. In a first step, significance values for over-expression in tissue/condition A or B are assigned to all genes in the expression data set. Genes with a significance value exceeding a certain threshold are used as starting points for the reconstruction of a graph with signaling components as nodes and signaling events as edges. In a subsequent graph traversal process, again starting from the previously identified differentially expressed genes, all encountered nodes 'inherit' all their starting nodes' significance values. In a final step, the graph is visualized, the nodes being colored according to a weighted average of their inherited significance values. Each node's, or sub-network's, predominant color, ranging from green (significant for tissue/condition A) over yellow (not significant for either tissue/condition) to red (significant for tissue/condition B), thus gives an immediate visual clue on which molecules--differentially expressed or not--may play pivotal roles in the tissues or conditions under examination. The described method has been implemented in Java as a client/server application and a web interface called DEEP (Differential Expression Effector Prediction). The client, which features an easy-to-use graphical interface, can freely be downloaded from the following URL: http://deep.bioinf.med.uni-goettingen.de. PMID:17584786

  17. Secreted Fungal Effector Lipase Releases Free Fatty Acids to Inhibit Innate Immunity-Related Callose Formation during Wheat Head Infection[W][OPEN

    PubMed Central

    Blümke, Antje; Falter, Christian; Herrfurth, Cornelia; Sode, Björn; Bode, Rainer; Schäfer, Wilhelm; Feussner, Ivo; Voigt, Christian A.

    2014-01-01

    The deposition of the (1,3)-?-glucan cell wall polymer callose at sites of attempted penetration is a common plant defense response to intruding pathogens and part of the plant’s innate immunity. Infection of the Fusarium graminearum disruption mutant ?fgl1, which lacks the effector lipase FGL1, is restricted to inoculated wheat (Triticum aestivum) spikelets, whereas the wild-type strain colonized the whole wheat spike. Our studies here were aimed at analyzing the role of FGL1 in establishing full F. graminearum virulence. Confocal laser-scanning microscopy revealed that the ?fgl1 mutant strongly induced the deposition of spot-like callose patches in vascular bundles of directly inoculated spikelets, while these callose deposits were not observed in infections by the wild type. Elevated concentrations of the polyunsaturated free fatty acids (FFAs) linoleic and ?-linolenic acid, which we detected in F. graminearum wild type-infected wheat spike tissue compared with ?fgl1-infected tissue, provided clear evidence for a suggested function of FGL1 in suppressing callose biosynthesis. These FFAs not only inhibited plant callose biosynthesis in vitro and in planta but also partially restored virulence to the ?fgl1 mutant when applied during infection of wheat spikelets. Additional FFA analysis confirmed that the purified effector lipase FGL1 was sufficient to release linoleic and ?-linolenic acids from wheat spike tissue. We concluded that these two FFAs have a major function in the suppression of the innate immunity-related callose biosynthesis and, hence, the progress of F. graminearum wheat infection. PMID:24686113

  18. Next generation fire suppressants

    NASA Technical Reports Server (NTRS)

    Brown, Jerry A.

    1995-01-01

    Spectrex, Inc., located in Cedar Grove, NJ is a manufacturer of fire detection and suppression equipment. Spectrex is one of the original pioneers in high speed fire detection and suppression systems for combat vehicles. Spectrex has installed fire suppressions systems in thousands of combat vehicles and ships throughout the world. Additionally, they manufacture flame explosion detectors, ship damage control systems, and optical gas and vapor detectors. The culmination of several years of research and development has recently produced an innovative electro-optical continuous monitoring systems called SharpEye 20/20I IR(sup 3) and SAFEYE that provide fast and reliable gas, vapor, aerosol, flame, and explosion detection. SharpEye 20/20I IR(sup 3) is a self-contained triple spectrum flame detector which scans for oscillating IR radiation (1 to 10 Hz) in the spectral bands ranging from 4.0 to 5.0 microns and uses programmed algorithms to check the ratio and correlation of data received by the three sensors to make the system highly immune to false alarms. It is extremely sensitive as it can detect a 1 x 1 square foot gasoline pan fire at 200 feet in less than 3 seconds. The sensitivity is user programmable, offering 4 ranges of detection. SAFEYE is comprised of a selected number of multispectral ban microprocessors controlled detectors which are in communication with one or more radiation sources that is projected along a 600 feet optical path. The signals from the selected narrow bands are processed and analyzed by highly sophisticated algorithms. It is ideal for high risk, remote, large areas such as petroleum and chemical manufacturing sites, waste dumps, aircraft cargo bays, and ship compartments. The SAFEYE will perform direct readings of the presence or rate of rise of concentrations of gases, vapors, or aerosols at the range of parts per million and provide alarms at various set points at different levels of concentrations.

  19. The effector repertoire of Fusarium oxysporum determines the tomato xylem proteome composition following infection

    PubMed Central

    Gawehns, Fleur; Ma, Lisong; Bruning, Oskar; Houterman, Petra M.; Boeren, Sjef; Cornelissen, Ben J. C.; Rep, Martijn; Takken, Frank L. W.

    2015-01-01

    Plant pathogens secrete small proteins, of which some are effectors that promote infection. During colonization of the tomato xylem vessels the fungus Fusarium oxysporum f.sp. lycopersici (Fol) secretes small proteins that are referred to as SIX (Secreted In Xylem) proteins. Of these, Six1 (Avr3), Six3 (Avr2), Six5, and Six6 are required for full virulence, denoting them as effectors. To investigate their activities in the plant, the xylem sap proteome of plants inoculated with Fol wild-type or either AVR2, AVR3, SIX2, SIX5, or SIX6 knockout strains was analyzed with nano-Liquid Chromatography-Mass Spectrometry (nLC-MSMS). Compared to mock-inoculated sap 12 additional plant proteins appeared while 45 proteins were no longer detectable in the xylem sap of Fol-infected plants. Of the 285 proteins found in both uninfected and infected plants the abundance of 258 proteins changed significantly following infection. The xylem sap proteome of plants infected with four Fol effector knockout strains differed significantly from plants infected with wild-type Fol, while that of the SIX2-knockout inoculated plants remained unchanged. Besides an altered abundance of a core set of 24 differentially accumulated proteins (DAPs), each of the four effector knockout strains affected specifically the abundance of a subset of DAPs. Hence, Fol effectors have both unique and shared effects on the composition of the tomato xylem sap proteome. PMID:26583031

  20. Wind Tunnel Test of an RPV with Shape-Change Control Effector and Sensor Arrays

    NASA Technical Reports Server (NTRS)

    Raney, David L.; Cabell, Randolph H.; Sloan, Adam R.; Barnwell, William G.; Lion, S. Todd; Hautamaki, Bret A.

    2004-01-01

    A variety of novel control effector concepts have recently emerged that may enable new approaches to flight control. In particular, the potential exists to shift the composition of the typical aircraft control effector suite from a small number of high authority, specialized devices (rudder, aileron, elevator, flaps), toward larger numbers of smaller, less specialized, distributed device arrays. The concept envisions effector and sensor networks composed of relatively small high-bandwidth devices able to simultaneously perform a variety of control functions using feedback from disparate data sources. To investigate this concept, a remotely piloted flight vehicle has been equipped with an array of 24 trailing edge shape-change effectors and associated pressure measurements. The vehicle, called the Multifunctional Effector and Sensor Array (MESA) testbed, was recently tested in NASA Langley's 12-ft Low Speed wind tunnel to characterize its stability properties, control authorities, and distributed pressure sensitivities for use in a dynamic simulation prior to flight testing. Another objective was to implement and evaluate a scheme for actively controlling the spanwise pressure distribution using the shape-change array. This report describes the MESA testbed, design of the pressure distribution controller, and results of the wind tunnel test.

  1. Perturbation of host ubiquitin systems by plant pathogen/pest effector proteins

    PubMed Central

    Banfield, Mark J

    2015-01-01

    Microbial pathogens and pests of animals and plants secrete effector proteins into host cells, altering cellular physiology to the benefit of the invading parasite. Research in the past decade has delivered significant new insights into the molecular mechanisms of how these effector proteins function, with a particular focus on modulation of host immunity-related pathways. One host system that has emerged as a common target of effectors is the ubiquitination system in which substrate proteins are post-translationally modified by covalent conjugation with the small protein ubiquitin. This modification, typically via isopeptide bond formation through a lysine side chain of ubiquitin, can result in target degradation, relocalization, altered activity or affect protein–protein interactions. In this review, I focus primarily on how effector proteins from bacterial and filamentous pathogens of plants and pests perturb host ubiquitination pathways that ultimately include the 26S proteasome. The activities of these effectors, in how they affect ubiquitin pathways in plants, reveal how pathogens have evolved to identify and exploit weaknesses in this system that deliver increased pathogen fitness. PMID:25339602

  2. Engraftment of human central memory-derived effector CD8+ T cells in immunodeficient mice.

    PubMed

    Wang, Xiuli; Berger, Carolina; Wong, ChingLam W; Forman, Stephen J; Riddell, Stanley R; Jensen, Michael C

    2011-02-10

    In clinical trials of adoptive T-cell therapy, the persistence of transferred cells correlates with therapeutic efficacy. However, properties of human T cells that enable their persistence in vivo are poorly understood, and model systems that enable investigation of the fate of human effector T cells (T(E)) have not been described. Here, we analyzed the engraftment of adoptively transferred human cytomegalovirus pp65-specific CD8(+) T(E) cells derived from purified CD45RO(+)CD62L(+) central memory (T(CM)) or CD45RO(+)CD62L(-) effector memory (T(EM)) precursors in an immunodeficient mouse model. The engraftment of T(CM)-derived effector cells (T(CM/E)) was dependent on human interleukin-15, and superior in magnitude and duration to T(EM)-derived effector cells (T(EM/E)). T-cell receptor V? analysis of persisting cells demonstrated that CD8(+) T(CM/E) engraftment was polyclonal, suggesting that the ability to engraft is a general feature of T(CM/E.) CD8(+) T(EM/E) proliferated extensively after transfer but underwent rapid apoptosis. In contrast, T(CM/E) were less prone to apoptosis and established a persistent reservoir of functional T cells in vivo characterized by higher CD28 expression. These studies predict that human CD8(+) effector T cells derived from T(CM) precursors may be preferred for adoptive therapy based on superior engraftment fitness. PMID:21123821

  3. An Unbiased Method for Clustering Bacterial Effectors Using Host Cellular Phenotypes

    PubMed Central

    Hodgson, David J.

    2014-01-01

    We present a novel method implementing unbiased high-content morphometric cell analysis to classify bacterial effector phenotypes. This clustering methodology represents a significant advance over more qualitative visual approaches and can also be used to classify, and therefore predict the likely function of, unknown effector genes from any microbial genome. As a proof of concept, we use this approach to investigate 23 genetic regions predicted to encode antimacrophage effectors located across the genome of the insect and human pathogen Photorhabdus asymbiotica. Statistical cluster analysis using multiple cellular measures categorized treated macrophage phenotypes into three major groups relating to their putative functionality: (i) adhesins, (ii) cytolethal toxins, and (iii) cytomodulating toxins. Further investigation into their effects on phagocytosis revealed that several effectors also modulate this function and that the nature of this modulation (increased or decreased phagocytosis) is linked to the phenotype cluster group. Categorizing potential functionalities in this way allows rapid functional follow-up of key candidates for more-directed cell biological or biochemical investigation. Such an unbiased approach to the classification of candidate effectors will be useful for describing virulence-related regions in a wide range of genomes and will be useful in assigning putative functions to the growing number of microbial genes whose function remains unclear from homology searching. PMID:24296505

  4. 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 effectors. PMID:26257749

  5. Bacterial effectors target the plant cell nucleus to subvert host transcription

    PubMed Central

    Canonne, Joanne; Rivas, Susana

    2012-01-01

    In order to promote virulence, Gram-negative bacteria have evolved the ability to inject so-called type III effector proteins into host cells. The plant cell nucleus appears to be a subcellular compartment repeatedly targeted by bacterial effectors. In agreement with this observation, mounting evidence suggests that manipulation of host transcription is a major strategy developed by bacteria to counteract plant defense responses. It has been suggested that bacterial effectors may adopt at least three alternative, although not mutually exclusive, strategies to subvert host transcription. T3Es may (1) act as transcription factors that directly activate transcription in host cells, (2) affect histone packing and chromatin configuration, and/or (3) directly target host transcription factor activity. Here, we provide an overview on how all these strategies may lead to host transcriptional re-programming and, as a result, to improved bacterial multiplication inside plant cells. PMID:22353865

  6. Toluene 4-Monooxygenase and its Complex with Effector Protein T4moD

    SciTech Connect

    Bailey, Lucas J.; Fox, Brian G.

    2012-10-16

    Toluene 4-monooxygenase (T4MO) is a multiprotein diiron enzyme complex that catalyzes the regiospecific oxidation of toluene to p-cresol. Catalytic function requires the presence of a small protein, called the effector protein. Effector protein exerts substantial control on the diiron hydroxylase catalytic cycle through protein-protein interactions. High-resolution crystal structures of the stoichometric hydroxylase and effector protein complex described here reveal how protein-protein interactions and reduction of the diiron center produce an active site configuration poised for reaction with O{sub 2}. Further information from crystal structures of mutated isoforms of the hydroxylase and a peroxo adduct is combined with catalytic results to give a fuller picture of the geometry of the enzyme-substrate complex used for the high fidelity oxidation of hydrocarbon substrates.

  7. Antibacterial effector/immunity systems: it's just the tip of the iceberg.

    PubMed

    Benz, Juliane; Meinhart, Anton

    2014-02-01

    Bacteria do not live anchoretic; rather they are constantly in touch with their eukaryotic hosts and with other bacteria sharing their habitat. Therefore, bacteria have evolved sophisticated proteinaceous weapons. To harm other bacteria, they produce antibacterial effector proteins, which they either release into the environment or export via direct intercellular contact. Contact-dependent killing is mediated by two specialized secretion systems, the type V and VI secretion system, whereas contact-independent processes hijack other transport mechanisms. Regardless of the transport system, cells co-express immunity proteins to protect themselves from suicide and fratricide. In general, effector protein activities and secretion mechanisms differ between Gram-positive and Gram-negative bacteria and evidence is emerging that different effector/immunity systems act synergistically and thus extend the bacterial armory. PMID:24581686

  8. Structural Basis for Sequence-Specific Recognition of DNA by TAL Effectors

    PubMed Central

    Deng, Dong; Yan, Chuangye; Pan, Xiaojing; Mahfouz, Magdy; Wang, Jiawei; Zhu, Jian-Kang; Shi, Yigong; Yan, Nieng

    2013-01-01

    TAL (transcription activator–like) effectors, secreted by phytopathogenic bacteria, recognize host DNA sequences through a central domain of tandem repeats. Each repeat comprises 33 to 35 conserved amino acids and targets a specific base pair by using two hypervariable residues [known as repeat variable diresidues (RVDs)] at positions 12 and 13. Here, we report the crystal structures of an 11.5-repeat TAL effector in both DNA-free and DNA-bound states. Each TAL repeat comprises two helices connected by a short RVD-containing loop. The 11.5 repeats form a right-handed, superhelical structure that tracks along the sense strand of DNA duplex, with RVDs contacting the major groove. The 12th residue stabilizes the RVD loop, whereas the 13th residue makes a base-specific contact. Understanding DNA recognition by TAL effectors may facilitate rational design of DNA-binding proteins with biotechnological applications. PMID:22223738

  9. Transcription Factor Networks Directing the Development, Function, and Evolution of Innate Lymphoid Effectors

    PubMed Central

    Kang, Joonsoo; Malhotra, Nidhi

    2015-01-01

    Mammalian lymphoid immunity is mediated by fast and slow responders to pathogens. Fast innate lymphocytes are active within hours after infections in mucosal tissues. Slow adaptive lymphocytes are conventional T and B cells with clonal antigen receptors that function days after pathogen exposure. A transcription factor (TF) regulatory network guiding early T cell development is at the core of effector function diversification in all innate lymphocytes, and the kinetics of immune responses is set by developmental programming. Operational units within the innate lymphoid system are not classified by the types of pathogen-sensing machineries but rather by discrete effector functions programmed by regulatory TF networks. Based on the evolutionary history of TFs of the regulatory networks, fast effectors likely arose earlier in the evolution of animals to fortify body barriers, and in mammals they often develop in fetal ontogeny prior to the establishment of fully competent adaptive immunity. PMID:25650177

  10. Volumetric reach comparison of possible end-effectors for the articulated transporter and manipulator system

    SciTech Connect

    Kress, R.L.; Babcock, S.M.; Hamel, W.R. ); Bills, K.C. )

    1990-01-01

    The goal of this research was to investigate the performance of the Articulated Transporter and Manipulator System (ATMS) during various tasks relative to the choice of wrist/end-effector configuration. The approach taken was to generate computer graphics-aided three-dimensional interactive application (CATIA) system-based models of four wrist/end-effector combinations and consider the volumetric reach of each of these configurations based on the capacity of the ATMS. The results indicate that a simple, lightweight end-effector provides a greater volumetric reach. The greatest variation presented herein is {approximately}40% when comparing a 7-degree-of-freedom (DOF) dexterous arm with a simple 3-DOF arm; however, the benefit of increasing volumetric reach by only 40% by using a simple arm may be outweighed by the loss of dexterity. 10 refs., 5 figs., 3 tabs.

  11. Uncoupling T-cell expansion from effector differentiation in cell-based immunotherapy

    PubMed Central

    2013-01-01

    Summary Adoptive cellular immunotherapy (ACT) is a potentially curative therapy for patients with advanced cancer. Eradication of tumor in mouse models and humans correlates with both a high dose of adoptively transferred cells and cells with a minimally differentiated phenotype that maintain replicative capacity and multipotency. We speculate that response to ACT not only requires transfer of cells with immediate cytolytic effector function to kill the bulk of fast-growing tumor, but also transfer of tumor-specific cells that maintain an ability for self-renewal and the capacity to produce a continual supply of cytolytic effector progeny until all malignant cells are eliminated. Current in vitro methods to expand cells to sufficient numbers and still maintain a minimally differentiated phenotype are hindered by the biological coupling of clonal expansion and effector differentiation. Therefore, a better understanding of the physiologic mechanism that couples cell expansion and differentiation in CD8+ T cells may improve the efficacy of ACT. PMID:24329803

  12. Transcriptional Regulation with CRISPR/Cas9 Effectors in Mammalian Cells.

    PubMed

    Pham, Hannah; Kearns, Nicola A; Maehr, René

    2016-01-01

    CRISPR/Cas9-based regulation of gene expression provides the scientific community with a new high-throughput tool to dissect the role of genes in molecular processes and cellular functions. Single-guide RNAs allow for recruitment of a nuclease-dead Cas9 protein and transcriptional Cas9-effector fusion proteins to specific genomic loci, thereby modulating gene expression. We describe the application of a CRISPR-Cas9 effector system from Streptococcus pyogenes for transcriptional regulation in mammalian cells resulting in activation or repression of transcription. We present methods for appropriate target site selection, sgRNA design, and delivery of dCas9 and dCas9-effector system components into cells through lentiviral transgenesis to modulate transcription. PMID:26463376

  13. Activation of CD4 T cells by Raf-independent effectors of Ras.

    PubMed

    Czyzyk, Jan; Brogdon, Jennifer L; Badou, Abdallah; Henegariu, Octavian; Preston Hurlburt, Paula; Flavell, Richard; Bottomly, Kim

    2003-05-13

    Small GTPase Ras is capable of mediating activation in T lymphocytes by using Raf kinase-dependent signaling pathway. Other effectors of Ras exist, however, suggesting that targets of Ras alternative to Raf may also contribute to T cell functions. Here we demonstrate that Ras(V12G37) mutant that fails to bind Raf, potently increases intracellular calcium concentration and cytokine production in primary antigen-stimulated T cells. From three known effectors which retain the ability to interact with Ras(V12G37), overexpression of phospholipase C epsilon but not that of RIN1 or Ral guanine nucleotide exchange factors enhanced cytokine and nuclear factor-activated T cell reporter T cell responses. Hence T cell activation can be critically regulated by the Ras effector pathway independent from Raf that can be mimicked by phospholipase C epsilon. PMID:12721365

  14. Activation of CD4 T cells by Raf-independent effectors of Ras

    PubMed Central

    Czyzyk, Jan; Brogdon, Jennifer L.; Badou, Abdallah; Henegariu, Octavian; Preston Hurlburt, Paula; Flavell, Richard; Bottomly, Kim

    2003-01-01

    Small GTPase Ras is capable of mediating activation in T lymphocytes by using Raf kinase-dependent signaling pathway. Other effectors of Ras exist, however, suggesting that targets of Ras alternative to Raf may also contribute to T cell functions. Here we demonstrate that RasV12G37 mutant that fails to bind Raf, potently increases intracellular calcium concentration and cytokine production in primary antigen-stimulated T cells. From three known effectors which retain the ability to interact with RasV12G37, overexpression of phospholipase C ? but not that of RIN1 or Ral guanine nucleotide exchange factors enhanced cytokine and nuclear factor-activated T cell reporter T cell responses. Hence T cell activation can be critically regulated by the Ras effector pathway independent from Raf that can be mimicked by phospholipase C ?. PMID:12721365

  15. 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

  16. CD152 (CTLA-4) regulates effector functions of CD8+ T lymphocytes by repressing Eomesodermin.

    PubMed

    Hegel, Johannes K; Knieke, Karin; Kolar, Paula; Reiner, Steven L; Brunner-Weinzierl, Monika C

    2009-03-01

    CD8(+) T lymphocytes are required for effective host defense against pathogens and also for mediating effector responses against uncontrolled proliferating self-tissues. In this study, we determine that individual CD8(+) T cells are tightly controlled in their effector functions by CD152 (CTLA-4). We demonstrate that signals induced by CD152 reduce the frequency of IFN-gamma and granzyme B expressing CD8(+) T cells independently of the transcription factors T-bet or cKrox by selectively inhibiting accumulation of Eomesodermin mRNA and protein. Ectopic expression of Eomesodermin reversed the CD152-mediated inhibition of effector molecule production. Additionally, enhanced cytotoxicity of individual CD8(+) T cells differentiated in the absence of CD152 signaling was determined in vivo. These novel insights extend our understanding of how immune responses of CD8(+) T cells are selectively modulated. PMID:19224637

  17. Factors influencing dust suppressant effectiveness

    SciTech Connect

    Copeland, C.R.; Eisele, T.C.; Chesney, D.J.; Kawatra, S.K.

    2008-11-15

    Water sprays are a common method used to reduce particulate matter (PM) emissions. Various factors such as wettability, surface area coverage, fine particle engulfment rates, interparticle adhesion forces, suppressant penetration and suppressant longevity have all been suggested as critical factors in achieving effective PM control. However, it has not been established which of these factors are the most important. Experimental work indicated that suppressant penetration is the most critical of these factors. The length of time after application that suppressants were effective was also improved by using hygroscopic reagents that retained moisture to prevent evaporation. Maximizing suppressant penetration and improving suppressant longevity led to an average 86% reduction in PM10 concentrations in laboratory dust tower tests.

  18. Coxiella burnetii Effector Proteins That Localize to the Parasitophorous Vacuole Membrane Promote Intracellular Replication

    PubMed Central

    Larson, Charles L.; Beare, Paul A.; Voth, Daniel E.; Howe, Dale; Cockrell, Diane C.; Bastidas, Robert J.; Valdivia, Raphael H.

    2014-01-01

    The intracellular bacterial pathogen Coxiella burnetii directs biogenesis of a parasitophorous vacuole (PV) that acquires host endolysosomal components. Formation of a PV that supports C. burnetii replication requires a Dot/Icm type 4B secretion system (T4BSS) that delivers bacterial effector proteins into the host cell cytosol. Thus, a subset of T4BSS effectors are presumed to direct PV biogenesis. Recently, the PV-localized effector protein CvpA was found to promote C. burnetii intracellular growth and PV expansion. We predict additional C. burnetii effectors localize to the PV membrane and regulate eukaryotic vesicle trafficking events that promote pathogen growth. To identify these vacuolar effector proteins, a list of predicted C. burnetii T4BSS substrates was compiled using bioinformatic criteria, such as the presence of eukaryote-like coiled-coil domains. Adenylate cyclase translocation assays revealed 13 proteins were secreted in a Dot/Icm-dependent fashion by C. burnetii during infection of human THP-1 macrophages. Four of the Dot/Icm substrates, termed Coxiella vacuolar protein B (CvpB), CvpC, CvpD, and CvpE, labeled the PV membrane and LAMP1-positive vesicles when ectopically expressed as fluorescently tagged fusion proteins. C. burnetii ?cvpB, ?cvpC, ?cvpD, and ?cvpE mutants exhibited significant defects in intracellular replication and PV formation. Genetic complementation of the ?cvpD and ?cvpE mutants rescued intracellular growth and PV generation, whereas the growth of C. burnetii ?cvpB and ?cvpC was rescued upon cohabitation with wild-type bacteria in a common PV. Collectively, these data indicate C. burnetii encodes multiple effector proteins that target the PV membrane and benefit pathogen replication in human macrophages. PMID:25422265

  19. Coxiella burnetii effector proteins that localize to the parasitophorous vacuole membrane promote intracellular replication.

    PubMed

    Larson, Charles L; Beare, Paul A; Voth, Daniel E; Howe, Dale; Cockrell, Diane C; Bastidas, Robert J; Valdivia, Raphael H; Heinzen, Robert A

    2015-02-01

    The intracellular bacterial pathogen Coxiella burnetii directs biogenesis of a parasitophorous vacuole (PV) that acquires host endolysosomal components. Formation of a PV that supports C. burnetii replication requires a Dot/Icm type 4B secretion system (T4BSS) that delivers bacterial effector proteins into the host cell cytosol. Thus, a subset of T4BSS effectors are presumed to direct PV biogenesis. Recently, the PV-localized effector protein CvpA was found to promote C. burnetii intracellular growth and PV expansion. We predict additional C. burnetii effectors localize to the PV membrane and regulate eukaryotic vesicle trafficking events that promote pathogen growth. To identify these vacuolar effector proteins, a list of predicted C. burnetii T4BSS substrates was compiled using bioinformatic criteria, such as the presence of eukaryote-like coiled-coil domains. Adenylate cyclase translocation assays revealed 13 proteins were secreted in a Dot/Icm-dependent fashion by C. burnetii during infection of human THP-1 macrophages. Four of the Dot/Icm substrates, termed Coxiella vacuolar protein B (CvpB), CvpC, CvpD, and CvpE, labeled the PV membrane and LAMP1-positive vesicles when ectopically expressed as fluorescently tagged fusion proteins. C. burnetii ?cvpB, ?cvpC, ?cvpD, and ?cvpE mutants exhibited significant defects in intracellular replication and PV formation. Genetic complementation of the ?cvpD and ?cvpE mutants rescued intracellular growth and PV generation, whereas the growth of C. burnetii ?cvpB and ?cvpC was rescued upon cohabitation with wild-type bacteria in a common PV. Collectively, these data indicate C. burnetii encodes multiple effector proteins that target the PV membrane and benefit pathogen replication in human macrophages. PMID:25422265

  20. Effector-Independent Motor Sequence Representations Exist in Extrinsic and Intrinsic Reference Frames

    PubMed Central

    Wiestler, Tobias; Waters-Metenier, Sheena

    2014-01-01

    Many daily activities rely on the ability to produce meaningful sequences of movements. Motor sequences can be learned in an effector-specific fashion (such that benefits of training are restricted to the trained hand) or an effector-independent manner (meaning that learning also facilitates performance with the untrained hand). Effector-independent knowledge can be represented in extrinsic/world-centered or in intrinsic/body-centered coordinates. Here, we used functional magnetic resonance imaging (fMRI) and multivoxel pattern analysis to determine the distribution of intrinsic and extrinsic finger sequence representations across the human neocortex. Participants practiced four sequences with one hand for 4 d, and then performed these sequences during fMRI with both left and right hand. Between hands, these sequences were equivalent in extrinsic or intrinsic space, or were unrelated. In dorsal premotor cortex (PMd), we found that sequence-specific activity patterns correlated higher for extrinsic than for unrelated pairs, providing evidence for an extrinsic sequence representation. In contrast, primary sensory and motor cortices showed effector-independent representations in intrinsic space, with considerable overlap of the two reference frames in caudal PMd. These results suggest that effector-independent representations exist not only in world-centered, but also in body-centered coordinates, and that PMd may be involved in transforming sequential knowledge between the two. Moreover, although effector-independent sequence representations were found bilaterally, they were stronger in the hemisphere contralateral to the trained hand. This indicates that intermanual transfer relies on motor memories that are laid down during training in both hemispheres, but preferentially draws upon sequential knowledge represented in the trained hemisphere. PMID:24695723

  1. Characteristics necessary for an interconvertible enzyme cascade to generate a highly sensitive response to an effector.

    PubMed

    Cárdenas, M L; Cornish-Bowden, A

    1989-01-15

    A monocyclic interconvertible enzyme cascade, in which active and inactive states of an enzyme are interconverted by two opposing enzyme-catalysed reactions, does not necessarily produce a greater degree of sensitivity to an effector than one could expect from direct interaction between effector and target reaction. On the contrary, a cascade in which an effector acts on one of the enzymes catalysing the interconversion reactions by altering the apparent value of its specificity constant will always generate a less sensitive response than direct interaction would give. Nonetheless, even if both interconversion reactions obey Michaelis-Menten kinetics with the ordinary types of inhibition and activation, one can easily generate an enormous sensitivity in which a 0.5% change in concentration can increase the proportion of target enzyme in the active state from 10% to 90%: this corresponds approximately to a Hill coefficient of 800. To maximize the sensitivity, the following conditions must be satisfied: (1) both modifier enzymes must act under conditions of near saturation; (2) the effector must act on both of them in opposite directions; (3) it must alter the apparent values of their catalytic constants; (4) the enzyme subject to inhibition by the effector must respond at much lower effector concentrations than the enzyme subject to activation. As the last of these conditions appears to be counter-intuitive, it suggests that feeble activation of modifier enzymes in real systems may have passed unnoticed, or been dismissed as physiologically insignificant, although in reality crucial to the effective response of the system. PMID:2930453

  2. A novel C-terminal region within the multicargo type III secretion chaperone CesT contributes to effector secretion.

    PubMed

    Ramu, Thangadurai; Prasad, Madhulika Esther; Connors, Erica; Mishra, Amit; Thomassin, Jenny-Lee; Leblanc, Jason; Rainey, Jan K; Thomas, Nikhil A

    2013-02-01

    The enteropathogenic Escherichia coli (EPEC) multicargo chaperone CesT interacts with at least 10 effector proteins and is central to pathogenesis. CesT has been implicated in coordinating effector hierarchy, although the mechanisms behind this regulation are poorly understood. To address this question, we set out to functionally characterize CesT with respect to roles in (i) effector binding, (ii) effector recruitment to the type III secretion system (T3SS), and (iii) effector translocation into host cells. A CesT variant expression library was screened in EPEC using a newly developed semi-high-throughput secretion assay. Among many deficient CesT variants, a predominant number were localized to a novel CesT C-terminal region. These CesT C-terminal variants exhibited normal effector binding yet reduced effector secretion levels. Structural correlation and thermal spectroscopy analyses of purified CesT variants implicated multiple surface-exposed residues, a terminal helix region, and a flexible C-terminal triple-serine stretch in effector secretion. Site-directed mutagenesis of the flexible CesT C-terminal triple-serine sequence produced differential effector secretion, implicating this region in secretion events. Infection assays further indicated that the C-terminal region of CesT was important for NleA translocation into host cells but was dispensable for Tir translocation. The findings implicate the CesT C terminus in effector secretion and contribute to a model for multiple-cargo chaperone function and effector translocation into host cells during infection. PMID:23222727

  3. Genetic analysis of the individual contribution to virulence of the type III effector inventory of Pseudomonas syringae pv. phaseolicola.

    PubMed

    Macho, Alberto P; Zumaquero, Adela; Gonzalez-Plaza, Juan J; Ortiz-Martín, Inmaculada; Rufián, José S; Beuzón, Carmen R

    2012-01-01

    Several reports have recently contributed to determine the effector inventory of the sequenced strain Pseudomonas syringae pv. phaseolicola (Pph) 1448a. However, the contribution to virulence of most of these effectors remains to be established. Genetic analysis of the contribution to virulence of individual P. syringae effectors has been traditionally hindered by the lack of phenotypes of the corresponding knockout mutants, largely attributed to a high degree of functional redundancy within their effector inventories. In support of this notion, effectors from Pseudomonas syringae pv. tomato (Pto) DC3000 have been classified into redundant effector groups (REGs), analysing virulence of polymutants in the model plant Nicotiana benthamiana. However, using competitive index (CI) as a virulence assay, we were able to establish the individual contribution of AvrPto1(Pto) (DC3000) to Pto DC3000 virulence in tomato, its natural host, even though typically, contribution to virulence of AvrPto1 is only shown in strains also lacking AvrPtoB (also called HopAB2), a member of its REG. This report raised the possibility that even effectors targeting the same defence signalling pathway may have an individual contribution to virulence, and pointed out to CI assays as the means to establish such a contribution for individual effectors. In this work, we have analysed the individual contribution to virulence of the majority of previously uncharacterised Pph 1448a effectors, by monitoring the development of disease symptoms and determining the CI of single knockout mutants at different stages of growth within bean, its natural host. Despite their potential functional redundancy, we have found individual contributions to virulence for six out of the fifteen effectors analysed. In addition, we have analysed the functional relationships between effectors displaying individual contribution to virulence, highlighting the diversity that these relationships may present, and the interest of analysing their functions within the context of the infection. PMID:22558247

  4. Pressure suppression system

    DOEpatents

    Gluntz, D.M.

    1994-10-04

    A pressure suppression system includes a containment vessel surrounding a reactor pressure vessel and defining a drywell therein containing a non-condensable gas. An enclosed wetwell pool is disposed inside the containment vessel, and an enclosed gravity driven cooling system (GDCS) pool is disposed above the wetwell pool in the containment vessel. The GDCS pool includes a plenum for receiving through an inlet the non-condensable gas carried with steam from the drywell following a loss-of-coolant accident (LOCA). A condenser is disposed in the GDCS plenum for condensing the steam channeled therein and to trap the non-condensable gas therein. A method of operation includes draining the GDCS pool following the LOCA and channeling steam released into the drywell following the LOCA into the GDCS plenum for cooling along with the non-condensable gas carried therewith for trapping the gas therein. 3 figs.

  5. Pressure suppression system

    DOEpatents

    Gluntz, Douglas M. (San Jose, CA)

    1994-01-01

    A pressure suppression system includes a containment vessel surrounding a reactor pressure vessel and defining a drywell therein containing a non-condensable gas. An enclosed wetwell pool is disposed inside the containment vessel, and an enclosed gravity driven cooling system (GDCS) pool is disposed above the wetwell pool in the containment vessel. The GDCS pool includes a plenum for receiving through an inlet the non-condensable gas carried with steam from the drywell following a loss-of-coolant accident (LOCA). A condenser is disposed in the GDCS plenum for condensing the steam channeled therein and to trap the non-condensable gas therein. A method of operation includes draining the GDCS pool following the LOCA and channeling steam released into the drywell following the LOCA into the GDCS plenum for cooling along with the non-condensable gas carried therewith for trapping the gas therein.

  6. Ankyrin repeat proteins comprise a diverse family of bacterial type IV effectors.

    PubMed

    Pan, Xiaoxiao; Lührmann, Anja; Satoh, Ayano; Laskowski-Arce, Michelle A; Roy, Craig R

    2008-06-20

    Specialized secretion systems are used by many bacteria to deliver effector proteins into host cells that can either mimic or disrupt the function of eukaryotic factors. We found that the intracellular pathogens Legionella pneumophila and Coxiella burnetii use a type IV secretion system to deliver into eukaryotic cells a large number of different bacterial proteins containing ankyrin repeat homology domains called Anks. The L. pneumophila AnkX protein prevented microtubule-dependent vesicular transport to interfere with fusion of the L. pneumophila-containing vacuole with late endosomes after infection of macrophages, which demonstrates that Ank proteins have effector functions important for bacterial infection of eukaryotic host cells. PMID:18566289

  7. Assembly of Customized TAL Effectors Through Advanced ULtiMATE System.

    PubMed

    Yang, Junjiao; Guo, Shengjie; Yuan, Pengfei; Wei, Wensheng

    2016-01-01

    Transcription activator-like effectors (TALEs) have been widely applied in gene targeting. Here we describe an advanced ULtiMATE (USER-based Ligation-Mediated Assembly of TAL Effector) system that utilizes USER fusion technique and archive of 512 tetramer templates to achieve highly efficient construction of TALEs, which takes only half a day to accomplish the assembly of any given TALE construct. This system is also suitable for large-scale assembly of TALENs and any other TALE-based constructions. PMID:26443213

  8. Design criteria for the light duty utility arm system end effectors

    SciTech Connect

    Pardini, A.F.

    1995-01-03

    This document provides the criteria for the design of end effectors that will be used as part of the Light Duty Utility Arm (LDUA) System. The LDUA System consists of a deployment vehicle, a vertical positioning mast, a light duty multi-axis robotic arm, a tank riser interface and confinement, a tool interface plate, a control system, and an operations control trailer. The criteria specified in this document will apply to all end effector systems being developed for use on or with the LDUA system at the Hanford site. The requirement stipulated in this document are mandatory.

  9. Platelets: versatile effector cells in hemostasis, inflammation, and the immune continuum

    PubMed Central

    Vieira-de-Abreu, Adriana; Campbell, Robert A.; Weyrich, Andrew S.

    2015-01-01

    Platelets are chief effector cells in hemostasis. In addition, however, their specializations include activities and intercellular interactions that make them key effectors in inflammation and in the continuum of innate and adaptive immunity. This review focuses on the immune features of human platelets and platelets from experimental animals and on interactions between inflammatory, immune, and hemostatic activities of these anucleate but complex and versatile cells. The experimental findings and evidence for physiologic immune functions include previously unrecognized biologic characteristics of platelets and are paralleled by new evidence for unique roles of platelets in inflammatory, immune, and thrombotic diseases. PMID:21818701

  10. Tomato immune receptor Ve1 recognizes effector of multiple fungal pathogens uncovered by genome and RNA sequencing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fungal plant pathogens secrete effector molecules to establish disease on their hosts, while plants in turn utilize immune receptors to intercept these effectors. The tomato immune receptor Ve1 governs resistance to race 1 strains of the soil-borne vascular wilt fungi Verticillium dahliae and V. alb...

  11. CXCR3 directs antigen-specific effector CD4+ T cell migration to the lung during parainfluenza virus infection1

    PubMed Central

    Kohlmeier, Jacob E.; Cookenham, Tres; Miller, Shannon C.; Roberts, Alan D.; Christensen, Jan P.; Thomsen, Allan R.; Woodland, David L.

    2009-01-01

    Effector T cells are a crucial component of the adaptive immune response to respiratory virus infections. Although it was previously reported that the chemokine receptors CCR5 and CXCR3 are expressed on respiratory virus-specific effector CD8+ T cells, it is unclear whether these receptors govern effector CD4+ T cell migration to the lungs. To assess the role of CCR5 and CXCR3 in vivo, we directly compared the migration of antigen-specific wild-type and chemokine receptor-deficient effector T cells in mixed bone marrow chimeric mice during a parainfluenza virus infection. CXCR3-deficient effector CD4+ T cells were 5-10 fold less efficient at migrating to the lung compared with wild-type cells, whereas CCR5-deficient effector T cells were not impaired in their migration to the lung. In contrast to its role in trafficking, CXCR3 had no impact on effector CD4+ T cell proliferation, phenotype, or function in any of the tissues examined. These findings demonstrate that CXCR3 controls virus-specific effector CD4+ T cell migration in vivo, and suggest that blocking CXCR3-mediated recruitment may limit T cell-induced immunopathology during respiratory virus infections. PMID:19734208

  12. 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

  13. MODULATION OF THE NF-KAPPA B SIGNALING PATHWAY BY THE BACTERIAL TYPE III SECRETION SYSTEM EFFECTORS

    E-print Network

    Gao, Xiaofei

    2012-08-31

    The type III secretion system (T3SS) is a bacterial injection system expressed by many Gram-negative bacteria. During the last two decades, the repertoire of T3SS effectors has been greatly explored, and several mechanisms of these effectors have...

  14. An Alternative to Thought Suppression?

    ERIC Educational Resources Information Center

    Boice, Robert

    2012-01-01

    Comments on the original article, "Setting free the bears: Escape from thought suppression," by D. M. Wegner (see record 2011-25622-008). While Wegner supposed that we might have to learn to live with bad thoughts, the present author discusses the use of imagination and guided imagery as an alternative to forced thought suppression.

  15. STRV Cryocooler Tip Motion Suppression

    NASA Technical Reports Server (NTRS)

    Glaser, R.; Ross, R. G., Jr.; Johnson, D. L.

    1994-01-01

    The Space Technology Research Vehicle (STRV-1b) scheduled to fly at the beginning of June 1994, has a cryocooler vibration suppression experiment aboard doing motion suppression of the tip of the coldfinger. STRV-1b is a bread box sized satellite to be launched on the next flight of the Ariane-4.

  16. NIK1-mediated translation suppression functions as a plant antiviral immunity mechanism.

    PubMed

    Zorzatto, Cristiane; Machado, João Paulo B; Lopes, Kênia V G; Nascimento, Kelly J T; Pereira, Welison A; Brustolini, Otávio J B; Reis, Pedro A B; Calil, Iara P; Deguchi, Michihito; Sachetto-Martins, Gilberto; Gouveia, Bianca C; Loriato, Virgílio A P; Silva, Marcos A C; Silva, Fabyano F; Santos, Anésia A; Chory, Joanne; Fontes, Elizabeth P B

    2015-04-30

    Plants and plant pathogens are subject to continuous co-evolutionary pressure for dominance, and the outcomes of these interactions can substantially impact agriculture and food security. In virus-plant interactions, one of the major mechanisms for plant antiviral immunity relies on RNA silencing, which is often suppressed by co-evolving virus suppressors, thus enhancing viral pathogenicity in susceptible hosts. In addition, plants use the nucleotide-binding and leucine-rich repeat (NB-LRR) domain-containing resistance proteins, which recognize viral effectors to activate effector-triggered immunity in a defence mechanism similar to that employed in non-viral infections. Unlike most eukaryotic organisms, plants are not known to activate mechanisms of host global translation suppression to fight viruses. Here we demonstrate in Arabidopsis that the constitutive activation of NIK1, a leucine-rich repeat receptor-like kinase (LRR-RLK) identified as a virulence target of the begomovirus nuclear shuttle protein (NSP), leads to global translation suppression and translocation of the downstream component RPL10 to the nucleus, where it interacts with a newly identified MYB-like protein, L10-INTERACTING MYB DOMAIN-CONTAINING PROTEIN (LIMYB), to downregulate translational machinery genes fully. LIMYB overexpression represses ribosomal protein genes at the transcriptional level, resulting in protein synthesis inhibition, decreased viral messenger RNA association with polysome fractions and enhanced tolerance to begomovirus. By contrast, the loss of LIMYB function releases the repression of translation-related genes and increases susceptibility to virus infection. Therefore, LIMYB links immune receptor LRR-RLK activation to global translation suppression as an antiviral immunity strategy in plants. PMID:25707794

  17. Effector diversification within compartments of the Leptosphaeria maculans genome affected by Repeat-Induced Point mutations

    PubMed Central

    Rouxel, Thierry; Grandaubert, Jonathan; Hane, James K.; Hoede, Claire; van de Wouw, Angela P.; Couloux, Arnaud; Dominguez, Victoria; Anthouard, Véronique; Bally, Pascal; Bourras, Salim; Cozijnsen, Anton J.; Ciuffetti, Lynda M.; Degrave, Alexandre; Dilmaghani, Azita; Duret, Laurent; Fudal, Isabelle; Goodwin, Stephen B.; Gout, Lilian; Glaser, Nicolas; Linglin, Juliette; Kema, Gert H. J.; Lapalu, Nicolas; Lawrence, Christopher B.; May, Kim; Meyer, Michel; Ollivier, Bénédicte; Poulain, Julie; Schoch, Conrad L.; Simon, Adeline; Spatafora, Joseph W.; Stachowiak, Anna; Turgeon, B. Gillian; Tyler, Brett M.; Vincent, Delphine; Weissenbach, Jean; Amselem, Joëlle; Quesneville, Hadi; Oliver, Richard P.; Wincker, Patrick; Balesdent, Marie-Hélène; Howlett, Barbara J.

    2011-01-01

    Fungi are of primary ecological, biotechnological and economic importance. Many fundamental biological processes that are shared by animals and fungi are studied in fungi due to their experimental tractability. Many fungi are pathogens or mutualists and are model systems to analyse effector genes and their mechanisms of diversification. In this study, we report the genome sequence of the phytopathogenic ascomycete Leptosphaeria maculans and characterize its repertoire of protein effectors. The L. maculans genome has an unusual bipartite structure with alternating distinct guanine and cytosine-equilibrated and adenine and thymine (AT)-rich blocks of homogenous nucleotide composition. The AT-rich blocks comprise one-third of the genome and contain effector genes and families of transposable elements, both of which are affected by repeat-induced point mutation, a fungal-specific genome defence mechanism. This genomic environment for effectors promotes rapid sequence diversification and underpins the evolutionary potential of the fungus to adapt rapidly to novel host-derived constraints. PMID:21326234

  18. Action observation supports effector-dependent learning of finger movement sequences.

    PubMed

    Osman, Magda; Bird, Geoffrey; Heyes, Cecilia

    2005-08-01

    Practising a motor skill can result in effector-dependent learning (learning that does not transfer from the set of muscles used in training to a new set of muscles). Proceeding from neurophysiological evidence of motor activation during action observation, this study asked whether observational learning, learning through observation of skilled performance, can also be effector-dependent. Adult human participants observed a model's right hand as the model responded to an eight-item sequence in a serial reaction time (SRT) task. Their sequence learning was then compared in two tests with that of controls who had observed the model's right hand responding to random targets during training. All participants performed the SRT task with their right hand in the first test and with their left hand in the second. Evidence of observational learning was obtained in the right hand test but not in the left hand test. This implies that sequence learning based on observation of right hand performance did not transfer to the left hand, and therefore that observational learning can support effector-dependent learning of finger movement sequences. A second experiment used the same procedure to assess learning by a group of participants who observed a sequence of response locations only. This group did not observe the model's responses. Results suggested that action observation was necessary for the effector-dependent observational learning demonstrated in Experiment 1. PMID:15883806

  19. Optimal effector functions in human natural killer cells rely upon autocrine bone morphogenetic protein signaling

    PubMed Central

    Mc Alpine, Tristan; Wei, Heng; Martínez, Víctor G.; Entrena, Ana; Melen, Gustavo J; MacDonald, Andrew S.; Phythian-Adams, Alexander; Sacedón, Rosa; Maraskovsky, Eugene; Cebon, Jonathan; Ramírez, Manuel

    2014-01-01

    Natural killer (NK) cells are critical for innate tumor immunity due to their specialized ability to recognize and kill neoplastically transformed cells. However, NK cells require a specific set of cytokine-mediated signals to achieve optimal effector function. Th1-associated cytokines promote effector functions which are inhibited by the prototypic Th-2 cytokine IL-4 and the TGF-? superfamily members TGF-?1 and activin-A. Interestingly, the largest subgroup of the TGF-? superfamily are the bone morphogenetic proteins (BMP), but the effects of BMP signaling to NK cell effector functions have not been evaluated. Here we demonstrate that blood-circulating NK cells express type I and II BMP receptors, BMP-2 and BMP-6 ligands, and phosphorylated isoforms of Smad-1/-5/-8 which mediate BMP family member signaling. In opposition to the inhibitory effects of TGF-?1 or activin-A, autocrine BMP signaling was supportive to NK cell function. Mechanistic investigations in cytokine and TLR-L activated NK cells revealed that BMP signaling optimized IFN-? and global cytokine and chemokine production; phenotypic activation and proliferation; autologous DC activation and target cytotoxicity. Collectively, our findings identify a novel auto-activatory pathway that is essential for optimal NK cell effector function, one which might be therapeutically manipulated to help eradicate tumors. PMID:25038228

  20. A type III effector ADP-ribosylates RNA-binding proteins and quells plant

    E-print Network

    , Dorothee Staiger5 & James R. Alfano1 The bacterial plant pathogen Pseudomonas syringae injects effector immunity by affecting RNA metabolism and the plant defence transcriptome. Many Gram-negative pathogens of plants and animals and other eukaryotic-associated bacteria use type III protein secretion sys- tems1

  1. High Efficiency Ex Vivo Cloning of Antigen-Specific Human Effector T Cells

    PubMed Central

    Neller, Michelle A.; Lai, Michael H.-L.; Lanagan, Catherine M.; O?Connor, Linda E.; Pritchard, Antonia L.; Martinez, Nathan R.; Schmidt, Christopher W.

    2014-01-01

    While cloned T cells are valuable tools for the exploration of immune responses against viruses and tumours, current cloning methods do not allow inferences to be made about the function and phenotype of a clone's in vivo precursor, nor can precise cloning efficiencies be calculated. Additionally, there is currently no general method for cloning antigen-specific effector T cells directly from peripheral blood mononuclear cells, without the need for prior expansion in vitro. Here we describe an efficient method for cloning effector T cells ex vivo. Functional T cells are detected using optimised interferon gamma capture following stimulation with viral or tumour cell-derived antigen. In combination with multiple phenotypic markers, single effector T cells are sorted using a flow cytometer directly into multi-well plates, and cloned using standard, non antigen-specific expansion methods. We provide examples of this novel technology to generate antigen-reactive clones from healthy donors using Epstein-Barr virus and cytomegalovirus as representative viral antigen sources, and from two melanoma patients using autologous melanoma cells. Cloning efficiency, clonality, and retention/loss of function are described. Ex vivo effector cell cloning provides a rapid and effective method of deriving antigen-specific T cells clones with traceable in vivo precursor function and phenotype. PMID:25368986

  2. The rust transferred proteins-a new family of effector proteins exhibiting protease inhibitor function.

    PubMed

    Pretsch, Klara; Kemen, Ariane; Kemen, Eric; Geiger, Matthias; Mendgen, Kurt; Voegele, Ralf

    2013-01-01

    Only few fungal effectors have been described to be delivered into the host cell during obligate biotrophic interactions. RTP1p, from the rust fungi Uromyces fabae and U.?striatus, was the first fungal protein for which localization within the host cytoplasm could be demonstrated directly. We investigated the occurrence of RTP1 homologues in rust fungi and examined the structural and biochemical characteristics of the corresponding gene products. The analysis of 28 homologues showed that members of the RTP family are most likely to occur ubiquitously in rust fungi and to be specific to the order Pucciniales. Sequence analyses indicated that the structure of the RTPp effectors is bipartite, consisting of a variable N-terminus and a conserved and structured C-terminus. The characterization of Uf-RTP1p mutants showed that four conserved cysteine residues sustain structural stability. Furthermore, the C-terminal domain exhibits similarities to that of cysteine protease inhibitors, and it was shown that Uf-RTP1p and Us-RTP1p are able to inhibit proteolytic activity in Pichia pastoris culture supernatants. We conclude that the RTP1p homologues constitute a rust fungi-specific family of modular effector proteins comprising an unstructured N-terminal domain and a structured C-terminal domain, which exhibit protease inhibitory activity possibly associated with effector function during biotrophic interactions. PMID:22998218

  3. Disturbance of Arabidopsis thaliana microRNA-regulated pathways by Xcc bacterial effector proteins.

    PubMed

    Kurubanjerdjit, Nilubon; Tsai, Jeffrey J P; Huang, Chien-Hung; Ng, Ka-Lok

    2014-04-01

    Plants are continuously subjected to infection by pathogens, including bacteria and viruses. Bacteria can inject a variety of effector proteins into the host to reprogram host defense mechanism. It is known that microRNAs participate in plant disease resistance to bacterial pathogens and previous studies have suggested that some bacterial effectors have evolved to disturb the host's microRNA-regulated pathways; and so enabling infection. In this study, the inter-species interaction between an Xanthomonas campestris pv campestris (Xcc) pathogen effector and Arabidopsis thaliana microRNA transcription promoter was investigated using three methods: (1) interolog, (2) alignment based on using transcription factor binding site profile matrix, and (3) the web-based binding site prediction tool, PATSER. Furthermore, we integrated another two data sets from our previous study into the present web-based system. These are (1) microRNA target genes and their downstream effects mediated by protein-protein interaction (PPI), and (2) the Xcc-Arabidopsis PPI information. This present work is probably the first comprehensive study of constructing pathways that comprises effector, microRNA, target genes and PPI for the study of pathogen-host interactions. It is expected that this study may help to elucidate the role of pathogen-host interplay in a plant's immune system. The database is freely accessible at: http://ppi.bioinfo.asia.edu.tw/EDMRP . PMID:24385242

  4. Repertoire Development and the Control of Cytotoxic/Effector Function in Human ?? T Cells

    PubMed Central

    Urban, Elizabeth M.; Chapoval, Andrei I.; Pauza, C. David

    2010-01-01

    T cells develop into two major populations distinguished by their T cell receptor (TCR) chains. Cells with the ?? TCR generally express CD4 or CD8 lineage markers and mostly fall into helper or cytotoxic/effector subsets. Cells expressing the alternate ?? TCR in humans generally do not express lineage markers, do not require MHC for antigen presentation, and recognize nonpeptidic antigens. We are interested in the dominant V?2V?2+ T cell subset in human peripheral blood and the control of effector function in this population. We review the literature on ?? T cell generation and repertoire selection, along with recent work on CD56 expression and defining a cytotoxic/effector lineage within the phosphoantigen-reactive V?2V?2 cells. A unique mechanism for MHC-independent repertoire selection is linked to the control of effector function that is vital to the role for ?? T cells in tumor surveillance. Better understanding of these mechanisms will improve our ability to exploit this population for tumor immunotherapy. PMID:20396597

  5. nAture methods | VOL.11 NO.4 | APRIL2014 | 429 Although transcription activatorlike effector nucleases

    E-print Network

    Liu, David R.

    ­like effector nucleases (tAlens) can be designed to cleave chosen dnA sequences, tAlens have activity against related off-target sequences. to better understand tAlen specificity, we profiled 30 unique tAlens accessible and modified by tAlens in human cells. the results suggest that (i) tAle repeats bind dn

  6. Soluble Epoxide Hydrolase Is a Main Effector of Angiotensin IIInduced Hypertension

    E-print Network

    Hammock, Bruce D.

    Soluble Epoxide Hydrolase Is a Main Effector of Angiotensin II­Induced Hypertension Oliver JungEH contributes to angiotensin II­induced hypertension and tested the effects of a water-soluble sEH inhibitor, 12­induced hypertension (1 mg/kg per day). The effect of AUDA was accompanied by an increase in urinary salt and water

  7. A Novel End-Effector Design for Robotics in Image Guided Needle Procedures

    PubMed Central

    Sun, David; Willingham, Chris; Durrani, Amir; King, Paul; Cleary, Kevin; Wood, Bradford

    2008-01-01

    Robotic end-effectors are being developed to facilitate image-guided minimally-invasive needle-based procedures such as tumor ablation, biopsy, thoracentesis, and blood sampling. A novel mechanical end-effector was designed to address the challenges associated with any major needle-based procedure, focusing on liver biopsy and ablation. In this end-effector embodiment, the distal end of a single articulating arm can grip needles and instruments and allow a fairly high number of degrees of freedom of movement during the complex motions associated with positioning and driving needles, as well as the periodic motions associated with breathing patterns. Tightening a cable that runs through the articulations fixes the arm in a rigid state, allowing insertion of the gripped needle. In its final form, we diagram a design that will require electro-mechanical stimulation and remote joystick control. Moreover, we discuss how cranial-caudal motion of soft tissue organs and the associated forces affect design constraints. A simulation protocol describes the use of tissue phantoms with mechanical properties in the range of hepatic tissue and the overlying abdominal wall. Finally, an in vivo protocol details the possible use of a robotic arm coupled with our end-effector in an image-guided interventional suite. Such a switchable and flexible mode for a robotic arm overcomes much of the current limitations for automated needle placements for mobile targets, subject to breathing or patient motion and the inherent risks thereof. PMID:17520618

  8. Recognition of the Hyaloperonospora parasitica effector ATR13 triggers resistance against oomycete, bacterial, and viral pathogens.

    PubMed

    Rentel, Maike C; Leonelli, Lauriebeth; Dahlbeck, Douglas; Zhao, Bingyu; Staskawicz, Brian J

    2008-01-22

    Phytopathogenic oomycetes cause some of the most devastating diseases affecting agricultural crops. Hyaloperonospora parasitica is a native oomycete pathogen of Arabidopsis and is related to other oomycete phytopathogens that include several species of Phytophthora, including the causal agent of potato late blight. Recently, four oomycete effector genes have been isolated, and several oomycete genomes have been sequenced. We have developed an efficient and genetically amenable system to test putative effector genes using the bacterial pathogen Pseudomonas syringae pv. tomato DC3000. The H. parasitica effector protein ATR13 was delivered via P. syringae by fusing the ATR13 gene with the avrRpm1 type three secretion signal peptide, a bacterial sequence that allows transfer of proteins into the host cell through the bacterial type III secretion system. We also inserted ATR13 into the genome of the turnip mosaic virus, a single-stranded RNA virus. Our results show that delivery of ATR13 via the bacterial or viral pathogen triggers defense responses in plants containing the cognate resistance protein RPP13(Nd), which restricts proliferation of both pathogens. Hence, recognition of ATR13 by RPP13 initiates defense responses that are effective against oomycete, bacterial and viral pathogens, pointing to a common defense mechanism. We have characterized regions of the RPP13(Nd) resistance protein that are essential for effector recognition and/or downstream signaling, using transient coexpression in Nicotiana benthamiana. PMID:18198274

  9. Representation of the Speech Effectors in the Human Motor Cortex: Somatotopy or Overlap?

    ERIC Educational Resources Information Center

    Takai, Osamu; Brown, Steven; Liotti, Mario

    2010-01-01

    Somatotopy within the orofacial region of the human motor cortex has been a central concept in interpreting the results of neuroimaging and transcranial magnetic stimulation studies of normal and disordered speech. Yet, somatotopy has been challenged by studies showing overlap among the effectors within the homunculus. In order to address this…

  10. A genetic screen to isolate type III effectors translocated into pepper cells during

    E-print Network

    ) The bacterial pathogen Xanthomonas campestris pv. vesicatoria (Xcv) uses a type III secretion system (TTSS outside plant cells (3). We are interested in elucidating how Xanthomonas campestris pv. vesicatoria (XcvA genetic screen to isolate type III effectors translocated into pepper cells during Xanthomonas

  11. Structural Analysis of Substrate and Effector Binding in Mycobacterium tuberculosis D-3-Phosphoglycerate Dehydrogenase,

    E-print Network

    Grant, Gregory

    , NADH, in any of the crystals examined led us to study binding by stopped flow kinetic analysis. The kinetic data suggest that productive NADH binding, that would support catalytic turnover, is dependentStructural Analysis of Substrate and Effector Binding in Mycobacterium tuberculosis D-3

  12. Differential homing mechanisms regulate regionalized effector CD8??+ T cell accumulation within the small intestine

    PubMed Central

    Stenstad, Hanna; Svensson, Marcus; Cucak, Helena; Kotarsky, Knut; Agace, William W.

    2007-01-01

    The CC chemokine receptor (CCR)9 is expressed on the majority of small intestinal, but few colonic, T cells, whereas its ligand CCL25 is constitutively expressed by small intestinal epithelial cells. As such, CCR9/CCL25 have been proposed to play a central role in regulating small intestinal but not colonic immune responses and thus to organize regionalized immunity within the intestinal mucosa. Here, we demonstrate that CCL25 is expressed at reduced levels by epithelial cells in the distal compared with proximal small intestine, which correlated with less efficient CCR9-dependent effector CD8??+ T cell entry into the ileal epithelium. In vitro-generated ?4?7+ effector CD8??+ T cell entry into the lamina propria was less dependent on CCR9 than entry into the epithelium along the entire length of the small intestine and in particular in the ileum. CCR9-independent ?4?7+ effector CD8??+ T cell entry was pertussis toxin-sensitive, suggesting a role for additional G?I-linked G protein-coupled receptors. Finally, in vivo-primed effector CD8??+ T cells displayed regionalized differences in their entry to the small intestinal epithelium with enhanced CCR9-independent entry to the ileum. These results highlight a hitherto underappreciated compartmentalization of immune responses within the small intestine and have direct implications for targeting strategies aimed at regulating T cell localization to the small intestinal mucosa. PMID:17551016

  13. Quantitative analysis of receptor tyrosine kinase-effector coupling at functionally relevant stimulus levels.

    PubMed

    Li, Simin; Bhave, Devayani; Chow, Jennifer M; Riera, Thomas V; Schlee, Sandra; Rauch, Simone; Atanasova, Mariya; Cate, Richard L; Whitty, Adrian

    2015-04-17

    A major goal of current signaling research is to develop a quantitative understanding of how receptor activation is coupled to downstream signaling events and to functional cellular responses. Here, we measure how activation of the RET receptor tyrosine kinase on mouse neuroblastoma cells by the neurotrophin artemin (ART) is quantitatively coupled to key downstream effectors. We show that the efficiency of RET coupling to ERK and Akt depends strongly on ART concentration, and it is highest at the low (?100 pM) ART levels required for neurite outgrowth. Quantitative discrimination between ERK and Akt pathway signaling similarly is highest at this low ART concentration. Stimulation of the cells with 100 pM ART activated RET at the rate of ?10 molecules/cell/min, leading at 5-10 min to a transient peak of ?150 phospho-ERK (pERK) molecules and ?50 pAkt molecules per pRET, after which time the levels of these two signaling effectors fell by 25-50% while the pRET levels continued to slowly rise. Kinetic experiments showed that signaling effectors in different pathways respond to RET activation with different lag times, such that the balance of signal flux among the different pathways evolves over time. Our results illustrate that measurements using high, super-physiological growth factor levels can be misleading about quantitative features of receptor signaling. We propose a quantitative model describing how receptor-effector coupling efficiency links signal amplification to signal sensitization between receptor and effector, thereby providing insight into design principles underlying how receptors and their associated signaling machinery decode an extracellular signal to trigger a functional cellular outcome. PMID:25635057

  14. Ehrlichia chaffeensis Exploits Host SUMOylation Pathways To Mediate Effector-Host Interactions and Promote Intracellular Survival

    PubMed Central

    Dunphy, Paige Selvy; Luo, Tian

    2014-01-01

    Ehrlichia chaffeensis is an obligately intracellular Gram-negative bacterium that selectively infects mononuclear phagocytes. We recently reported that E. chaffeensis utilizes a type 1 secretion (T1S) system to export tandem repeat protein (TRP) effectors and demonstrated that these effectors interact with a functionally diverse array of host proteins. By way of these interactions, TRP effectors modulate host cell functions; however, the molecular basis of these interactions and their roles in ehrlichial pathobiology are not well defined. In this study, we describe the first bacterial protein posttranslational modification (PTM) by the small ubiquitin-like modifier (SUMO). The E. chaffeensis T1S effector TRP120 is conjugated to SUMO at a carboxy-terminal canonical consensus SUMO conjugation motif in vitro and in human cells. In human cells, TRP120 was selectively conjugated with SUMO2/3 isoforms. Disruption of TRP120 SUMOylation perturbed interactions with known host proteins, through predicted SUMO interaction motif-dependent and -independent mechanisms. E. chaffeensis infection did not result in dramatic changes in the global host SUMOylated protein profile, but a robust colocalization of predominately SUMO1 with ehrlichial inclusions was observed. Inhibiting the SUMO pathway with a small-molecule inhibitor had a significant impact on E. chaffeensis replication and recruitment of the TRP120-interacting protein polycomb group ring finger protein 5 (PCGF5) to the inclusion, indicating that the SUMO pathway is critical for intracellular survival. This study reveals the novel exploitation of the SUMO pathway by Ehrlichia, which facilitates effector-eukaryote interactions necessary to usurp the host and create a permissive intracellular niche. PMID:25047847

  15. Brucella Modulates Secretory Trafficking via Multiple Type IV Secretion Effector Proteins

    PubMed Central

    Myeni, Sebenzile; Child, Robert; Ng, Tony W.; Kupko, John J.; Wehrly, Tara D.; Porcella, Stephen F.; Knodler, Leigh A.; Celli, Jean

    2013-01-01

    The intracellular pathogenic bacterium Brucella generates a replicative vacuole (rBCV) derived from the endoplasmic reticulum via subversion of the host cell secretory pathway. rBCV biogenesis requires the expression of the Type IV secretion system (T4SS) VirB, which is thought to translocate effector proteins that modulate membrane trafficking along the endocytic and secretory pathways. To date, only a few T4SS substrates have been identified, whose molecular functions remain unknown. Here, we used an in silico screen to identify putative T4SS effector candidate proteins using criteria such as limited homology in other bacterial genera, the presence of features similar to known VirB T4SS effectors, GC content and presence of eukaryotic-like motifs. Using ?-lactamase and CyaA adenylate cyclase reporter assays, we identified eleven proteins translocated into host cells by Brucella, five in a VirB T4SS-dependent manner, namely BAB1_0678 (BspA), BAB1_0712 (BspB), BAB1_0847 (BspC), BAB1_1671 (BspE) and BAB1_1948 (BspF). A subset of the translocated proteins targeted secretory pathway compartments when ectopically expressed in HeLa cells, and the VirB effectors BspA, BspB and BspF inhibited protein secretion. Brucella infection also impaired host protein secretion in a process requiring BspA, BspB and BspF. Single or combined deletions of bspA, bspB and bspF affected Brucella ability to replicate in macrophages and persist in the liver of infected mice. Taken together, these findings demonstrate that Brucella modulates secretory trafficking via multiple T4SS effector proteins that likely act coordinately to promote Brucella pathogenesis. PMID:23950720

  16. Field performance of the waste retrieval end effectors in the Oak Ridge gunite tanks

    SciTech Connect

    Mullen, O.D.

    1997-09-01

    Waterjet-based tank waste retrieval end effectors have been developed by Retrieval Process Development and Enhancements through several generations of test articles targeted at deployment in Hanford underground storage tanks with a large robotic arm. The basic technology has demonstrated effectiveness for retrieval of simulants bounding a wide range of waste properties and compatibility with foreseen deployment systems. The Oak Ridge National Laboratory (ORNL) selected the waterjet scarifying end effector, the jet pump conveyance system, and the Modified Light Duty Utility Arm and Houdini Remotely Operated Vehicle deployment and manipulator systems for evaluation in the Gunite and Associated Tanks Treatability Study (GAAT-TS). The Retrieval Process Development and Enhancements (RPD&E) team was tasked with developing a version of the retrieval end effector tailored to the Oak Ridge tanks, waste, and deployment platforms. The conceptual design was done by the University of Missouri-Rolla in FY 1995-96. The university researchers conducted separate effects tests of the component concepts, scaled the basic design features, and constructed a full-scale test article incorporating their findings in early FY 1996. The test article was extensively evaluated in the Hanford Hydraulic Testbed and the design features were further refined. Detail design of the prototype item was started at Waterjet Technology, Inc. before the development testing was finished, and two of the three main subassemblies were substantially complete before final design of the waterjet manifold was determined from the Hanford hydraulic testbed (HTB) testing. The manifold on the first prototype was optimized for sludge retrieval; assembled with that manifold, the end effector is termed the Sludge Retrieval End Effector (SREE).

  17. Resequencing and Comparative Genomics of Stagonospora nodorum: Sectional Gene Absence and Effector Discovery

    PubMed Central

    Syme, Robert Andrew; Hane, James K.; Friesen, Timothy L.; Oliver, Richard P.

    2013-01-01

    Stagonospora nodorum is an important wheat (Triticum aestivum) pathogen in many parts of the world, causing major yield losses. It was the first species in the large fungal Dothideomycete class to be genome sequenced. The reference genome sequence (SN15) has been instrumental in the discovery of genes encoding necrotrophic effectors that induce disease symptoms in specific host genotypes. Here we present the genome sequence of two further S. nodorum strains (Sn4 and Sn79) that differ in their effector repertoire from the reference. Sn79 is avirulent on wheat and produces no apparent effectors when infiltrated onto many cultivars and mapping population parents. Sn4 is pathogenic on wheat and has virulences not found in SN15. The new strains, sequenced with short-read Illumina chemistry, are compared with SN15 by a combination of mapping and de novo assembly approaches. Each of the genomes contains a large number of strain-specific genes, many of which have no meaningful similarity to any known gene. Large contiguous sections of the reference genome are absent in the two newly sequenced strains. We refer to these differences as “sectional gene absences.” The presence of genes in pathogenic strains and absence in Sn79 is added to computationally predicted properties of known proteins to produce a list of likely effector candidates. Transposon insertion was observed in the mitochondrial genomes of virulent strains where the avirulent strain retained the likely ancestral sequence. The study suggests that short-read enabled comparative genomics is an effective way to both identify new S. nodorum effector candidates and to illuminate evolutionary processes in this species. PMID:23589517

  18. 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. PMID:26130320

  19. Crystal structure of the effector protein HopA1 from Pseudomonas syringae.

    PubMed

    Park, Yangshin; Shin, Inchul; Rhee, Sangkee

    2015-03-01

    Plants have evolved to protect themselves against pathogen attack; in these competitions, many Gram-negative bacteria translocate pathogen-originated proteins known as effectors directly into plant cells to interfere with cellular processes. Effector-triggered immunity (ETI) is a plant defense mechanism in which plant resistance proteins recognize the presence of effectors and initiate immune responses. Enhanced disease susceptibility 1 (EDS1) in Arabidopsis thaliana serves as a central node protein for basal immune resistance and ETI by interacting dynamically with other immune regulatory or resistance proteins. Recently, the effector HopA1 from Pseudomonas syringae was shown to affect these EDS1 complexes by binding EDS1 directly and activating the immune response signaling pathway. Here, we report the crystal structure of the effector HopA1 from P. syringae pv. syringae strain 61 and tomato strain DC3000. HopA1, a sequence-unrelated protein to EDS1, has an ?+? fold in which the central antiparallel ?-sheet is flanked by helices. A similar structural domain, an ?/? fold, is one of the two domains in both EDS1 and the EDS1-interacting protein SAG101, and plays a crucial role in forming the EDS1 complex. Further analyses suggest structural similarity and differences between HopA1 and the ?/? fold of SAG101, as well as between two HopA1s from different pathovars. Our structural analysis provides a foundation for understanding the molecular basis of the effect of HopA1 on plant immunity. PMID:25681297

  20. Pro-inflammatory effector Th cells transmigrate through anti-inflammatory environments into the murine fetus.

    PubMed

    Wienecke, J; Hebel, K; Hegel, K J; Pierau, M; Brune, T; Reinhold, D; Pethe, A; Brunner-Weinzierl, M C

    2012-01-01

    The presence of maternal DNA or even maternal cells within the offspring (microchimerism) has been reported for many fetal tissues, including the liver, heart, and spleen. Microchimerism is believed to be involved in the pathogenesis of autoimmune diseases; however, the cellular origin of this phenomenon remains unknown. Here, we determined whether differentiated T lymphocytes could transmigrate through the immunosuppressive environment of the placenta to reach the fetus. In vitro-differentiated effector/memory Th1 and Th17 cells from OVA???????-specific TCR(tg) T cells of OT-II mice were adoptively transferred (i.v.) into the tail veins of pregnant Ly5.1 mice at d15 and d19 of gestation. Mice were then sacrificed 40 h after adoptive cell transfer. Using radioactive labeling of T cells with sodium chromate [Cr?¹] prior to adoptive transfer, we observed that homing of pro-inflammatory Th cells was equally efficient in both pregnant and non-pregnant mice. Transmigration of Th1- and Th17-like cells through the highly immunosuppressive environment of the placenta into the fetus was significantly enhanced in experimental mice compared to control mice (P < 0.0001). In addition, a substantial amount of effector Th cells accumulated in the placenta. Finally, we found that treatment with Pertussis Toxin resulted in a 3-fold increase in the transmigration of effector Th17 cells into the fetus (P < 0.0001). When pro-inflammatory Th1-or Th17-like cells were injected into syngeneic mothers, almost all of the fetuses analyzed exhibited radioactivity, suggesting that transmigration of effector T cells occurs frequently. Our results suggest the possibility of novel roles for these maternal effector cells in the pathogenesis or reduction of disease. PMID:22093381

  1. IL-35 is a novel cytokine with therapeutic effects against collagen-induced arthritis through the expansion of regulatory T cells and suppression of Th17 cells.

    PubMed

    Niedbala, Wanda; Wei, Xiao-Qing; Cai, Beilei; Hueber, Axel J; Leung, Bernard P; McInnes, Iain B; Liew, Foo Y

    2007-11-01

    Epstein-Barr virus-induced gene 3 (EBI3) and the p35 subunit of IL-12 have been reported to form a heterodimeric hematopoietin in human and mouse. We have constructed a heterodimeric protein covalently linking EBI3 and p35, to form a novel cytokine which we now call IL-35. The Fc fusion protein of IL-35 induced proliferation of murine CD4(+)CD25(+) and CD4(+)CD25(-) T cells when stimulated with immobilized anti-CD3 and anti-CD28 antibodies in vitro. The IL-35-expanded CD4(+)CD25(+) T cell population expressed Foxp3 and produced elevated levels of IL-10, whereas the IL-35-induced CD4(+)CD25(-) T cells produced IFN-gamma but not IL-4. The in vitro expanded CD4(+)CD25(+) T cells retained their suppressive functions against CD4(+)CD25(-) effector cells. Furthermore, when cultured with soluble anti-CD3 antibody and antigen-presenting cells, IL-35 suppressed the proliferation of CD4(+)CD25(-) effector cells. Moreover, IL-35 inhibited the differentiation of Th17 cells in vitro. In vivo, IL-35 effectively attenuated established collagen-induced arthritis in mice, with concomitant suppression of IL-17 production but enhanced IFN-gamma synthesis. Thus, IL-35 is a novel anti-inflammatory cytokine suppressing the immune response through the expansion of regulatory T cells and suppression of Th17 cell development. PMID:17874423

  2. Loss of NOX-Derived Superoxide Exacerbates Diabetogenic CD4 T-Cell Effector Responses in Type 1 Diabetes.

    PubMed

    Padgett, Lindsey E; Anderson, Brian; Liu, Chao; Ganini, Douglas; Mason, Ronald P; Piganelli, Jon D; Mathews, Clayton E; Tse, Hubert M

    2015-12-01

    Reactive oxygen species (ROS) play prominent roles in numerous biological systems. While classically expressed by neutrophils and macrophages, CD4 T cells also express NADPH oxidase (NOX), the superoxide-generating multisubunit enzyme. Our laboratory recently demonstrated that superoxide-deficient nonobese diabetic (NOD.Ncf1(m1J)) mice exhibited a delay in type 1 diabetes (T1D) partially due to blunted IFN-? synthesis by CD4 T cells. For further investigation of the roles of superoxide on CD4 T-cell diabetogenicity, the NOD.BDC-2.5.Ncf1(m1J) (BDC-2.5.Ncf1(m1J)) mouse strain was generated, possessing autoreactive CD4 T cells deficient in NOX-derived superoxide. Unlike NOD.Ncf1(m1J), stimulated BDC-2.5.Ncf1(m1J) CD4 T cells and splenocytes displayed elevated synthesis of Th1 cytokines and chemokines. Superoxide-deficient BDC-2.5 mice developed spontaneous T1D, and CD4 T cells were more diabetogenic upon adoptive transfer into NOD.Rag recipients due to a skewing toward impaired Treg suppression. Exogenous superoxide blunted exacerbated Th1 cytokines and proinflammatory chemokines to approximately wild-type levels, concomitant with reduced IL-12R?2 signaling and P-STAT4 (Y693) activation. These results highlight the importance of NOX-derived superoxide in curbing autoreactivity due, in part, to control of Treg function and as a redox-dependent checkpoint of effector T-cell responses. Ultimately, our studies reveal the complexities of free radicals in CD4 T-cell responses. PMID:26269022

  3. Phytoplasma effector SAP54 hijacks plant reproduction by degrading MADS-box proteins and promotes insect colonization in a RAD23-dependent manner.

    PubMed

    MacLean, Allyson M; Orlovskis, Zigmunds; Kowitwanich, Krissana; Zdziarska, Anna M; Angenent, Gerco C; Immink, Richard G H; Hogenhout, Saskia A

    2014-04-01

    Pathogens that rely upon multiple hosts to complete their life cycles often modify behavior and development of these hosts to coerce them into improving pathogen fitness. However, few studies describe mechanisms underlying host coercion. In this study, we elucidate the mechanism by which an insect-transmitted pathogen of plants alters floral development to convert flowers into vegetative tissues. We find that phytoplasma produce a novel effector protein (SAP54) that interacts with members of the MADS-domain transcription factor (MTF) family, including key regulators SEPALLATA3 and APETALA1, that occupy central positions in the regulation of floral development. SAP54 mediates degradation of MTFs by interacting with proteins of the RADIATION SENSITIVE23 (RAD23) family, eukaryotic proteins that shuttle substrates to the proteasome. Arabidopsis rad23 mutants do not show conversion of flowers into leaf-like tissues in the presence of SAP54 and during phytoplasma infection, emphasizing the importance of RAD23 to the activity of SAP54. Remarkably, plants with SAP54-induced leaf-like flowers are more attractive for colonization by phytoplasma leafhopper vectors and this colonization preference is dependent on RAD23. An effector that targets and suppresses flowering while simultaneously promoting insect herbivore colonization is unprecedented. Moreover, RAD23 proteins have, to our knowledge, no known roles in flower development, nor plant defence mechanisms against insects. Thus SAP54 generates a short circuit between two key pathways of the host to alter development, resulting in sterile plants, and promotes attractiveness of these plants to leafhopper vectors helping the obligate phytoplasmas reproduce and propagate (zombie plants). PMID:24714165

  4. HIV Controllers Maintain a Population of Highly Efficient Th1 Effector Cells in Contrast to Patients Treated in the Long Term

    PubMed Central

    Vingert, Benoît; Benati, Daniela; Lambotte, Olivier; de Truchis, Pierre; Slama, Laurence; Jeannin, Patricia; Galperin, Moran; Perez-Patrigeon, Santiago; Boufassa, Faroudy; Kwok, William W.; Lemaître, Fabrice; Delfraissy, Jean-François; Thèze, Jacques

    2012-01-01

    HIV controllers are rare individuals who spontaneously control HIV replication in the absence of antiretroviral therapy. To identify parameters of the CD4 response that may contribute to viral control rather than merely reflect a persistently low viremia, we compared the T helper profiles in two groups of patients with more than 10 years of viral suppression: HIV controllers from the Agence Nationale de Recherche sur le SIDA et les Hépatites Virales (ANRS) CO18 cohort (n = 26) and efficiently treated patients (n = 16). Cells specific for immunodominant Gag and cytomegalovirus (CMV) peptides were evaluated for the production of 10 cytokines and cytotoxicity markers and were also directly quantified ex vivo by major histocompatibility complex (MHC) class II tetramer staining. HIV controller CD4+ T cells were characterized by a higher frequency of gamma interferon (IFN-?) production, perforin+/CD107a+ expression, and polyfunctionality in response to Gag peptides. While interleukin 4 (IL-4), IL-17, and IL-21 production did not differ between groups, the cells of treated patients produced more IL-10 in response to Gag and CMV peptides, pointing to persistent negative immunoregulation after long-term antiretroviral therapy. Gag293 tetramer-positive cells were detected at a high frequency (0.12%) and correlated positively with IFN-?-producing CD4+ T cells in the controller group (R = 0.73; P = 0.003). Tetramer-positive cells were fewer in the highly active antiretroviral therapy (HAART) group (0.04%) and did not correlate with IFN-? production, supporting the notion of a persistent immune dysfunction in HIV-specific CD4+ T cells of treated patients. In conclusion, HIV controllers maintained a population of highly efficient Th1 effectors directed against Gag in spite of a persistently low antigenemia, while patients treated in the long term showed a loss of CD4 effector functions. PMID:22837194

  5. Components of the Pseudomonas syringae type III secretion system can suppress and may elicit plant innate immunity.

    PubMed

    Oh, Hye-Sook; Park, Duck Hwan; Collmer, Alan

    2010-06-01

    The type III secretion system (T3SS) of Pseudomonas syringae translocates into plant cells multiple effectors that suppress pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). P. syringae pv. tomato DC3000 no longer delivers the T3SS translocation reporter AvrPto-Cya in Nicotiana benthamiana leaf tissue in which PTI was induced by prior inoculation with P. fluorescens(pLN18). Cosmid pLN18 expresses the T3SS system of P. syringae pv. syringae 61 but lacks the hopA1(Psy61) effector gene. P. fluorescens(pLN18) expressing HrpH(PtoDC3000) or HopP1(PtoDC3000), two T3SS-associated putative lytic transglycosylases, suppresses PTI, based on multiple assays involving DC3000 challenge inoculum (AvrPto-Cya translocation, hypersensitive response elicitation, and colony development in planta) or on plant responses (vascular dye uptake or callose deposition). Analysis of additional mutations in pHIR11 derivatives revealed that the pLN18-encoded T3SS elicits a higher level of reactive oxygen species (ROS) than does P. fluorescens without a T3SS, that enhanced ROS production is dependent on the HrpK1 translocator, and that HopA1(Psy61) suppresses ROS elicitation attributable to both the P. fluorescens PAMPs and the presence of a functional T3SS. PMID:20459312

  6. A ligation-independent cloning technique for high-throughput assembly of transcription activator–like effector genes.

    PubMed

    Schmid-Burgk, Jonathan L; Schmidt, Tobias; Kaiser, Vera; Höning, Klara; Hornung, Veit

    2013-01-01

    Transcription activator–like (TAL) effector proteins derived from Xanthomonas species have emerged as versatile scaffolds for engineering DNA-binding proteins of user-defined specificity and functionality. Here we describe a rapid, simple, ligation-independent cloning (LIC) technique for synthesis of TAL effector genes. Our approach is based on a library of DNA constructs encoding individual TAL effector repeat unit combinations that can be processed to contain long, unique single-stranded DNA overhangs suitable for LIC. Assembly of TAL effector arrays requires only the combinatorial mixing of fluids and has exceptional fidelity. TAL effector nucleases (TALENs) produced by this method had high genome-editing activity at endogenous loci in HEK 293T cells (64% were active). To maximize throughput, we generated a comprehensive 5-mer TAL effector repeat unit fragment library that allows automated assembly of >600 TALEN genes in a single day. Given its simplicity, throughput and fidelity, LIC assembly will permit the generation of TAL effector gene libraries for large-scale functional genomics studies. PMID:23242165

  7. Diverse evolutionary mechanisms shape the type III effector virulence factor repertoire in the plant pathogen Pseudomonas syringae.

    PubMed Central

    Rohmer, Laurence; Guttman, David S; Dangl, Jeffery L

    2004-01-01

    Many gram-negative pathogenic bacteria directly translocate effector proteins into eukaryotic host cells via type III delivery systems. Type III effector proteins are determinants of virulence on susceptible plant hosts; they are also the proteins that trigger specific disease resistance in resistant plant hosts. Evolution of type III effectors is dominated by competing forces: the likely requirement for conservation of virulence function, the avoidance of host defenses, and possible adaptation to new hosts. To understand the evolutionary history of type III effectors in Pseudomonas syringae, we searched for homologs to 44 known or candidate P. syringae type III effectors and two effector chaperones. We examined 24 gene families for distribution among bacterial species, amino acid sequence diversity, and features indicative of horizontal transfer. We assessed the role of diversifying and purifying selection in the evolution of these gene families. While some P. syringae type III effectors were acquired recently, others have evolved predominantly by descent. The majority of codons in most of these genes were subjected to purifying selection, suggesting selective pressure to maintain presumed virulence function. However, members of 7 families had domains subject to diversifying selection. PMID:15280247

  8. Augmented Followed by Suppressed Levels of Natural Cell-Mediated Cytotoxicity in Mice Infected with Toxoplasma gondii

    PubMed Central

    Kamiyama, Tsuneo; Hagiwara, Toshikatsu

    1982-01-01

    The cytotoxic activity of effector cells from mice infected with Toxoplasma gondii was tested in a 4- to 5-hr 51Cr release assay, using RL 0001000 0011100 0101010 0001000 0001000 0011100 0100010 1000001 1000001 1000001 0100010 0011100 1 and YAC-1 target cells. They showed enhanced cytotoxicity with a peak on the 3rd day postinfection followed by suppression with a peak on the 12th day. The cytotoxicity seemed to be exhibited by natural killer (NK) cells because: (i) pretreatment of the effector cells with antiasialo GM1 or antiasialo GM2 plus complement abolished the cytotoxic activity; (ii) the altered cytotoxicity levels were also induced in nude mice; and (iii) the activity was elicited by nonadherent-nonphagocytic cells. The alteration occurred simultaneously in various lymphoid organs with a similar profile. Neither spleen nor bone marrow cells of 12-day-postinfected mice inhibited NK activity of uninfected mice. Culture fluids of the infected mouse spleen and bone marrow cells did not affect the normal mouse NK activity. The proportion of effector cells capable of binding to target cells was constant during the infection. There was no positive correlation between NK activity and serum interferon level; i.e., interferon was detected in the serum of 12-day-postinfected mice but not in that of 3-day-postinfected or uninfected mice. Passively administered interferon or polyinosinic-polycytidylic acid could not restore the suppressed NK activity of 12-day-postinfected mice. Moreover, in vitro treatment of spleen cells from 12-day-postinfected mice with interferon failed to restore the suppressed NK activity. These results suggested that after toxoplasma infection, defective sensitivity to interferon was induced in NK precursor cells, and differentiation to functionally active NK cells might be blocked. PMID:6177634

  9. A formula for charmonium suppression

    E-print Network

    C. Pena; D. Blaschke

    2011-08-21

    In this work a formula for charmonium suppression obtained by Matsui in 1989 is analytically generalized for the case of complex c-cbar potential described by a 3-dimensional and isotropic time-dependent harmonic oscillator (THO). It is suggested that under certain conditions the formula can be applied to describe J/\\psi suppression in heavy-ion collisions at CERN-SPS, RHIC, and LHC with the advantage of analytical tractability.

  10. Size of attentional suppressive surround.

    PubMed

    Yoo, Sang-Ah; Tsotsos, John; Fallah, Mazyar

    2015-09-01

    The Selective Tuning (ST) model (Tsotsos, 1995) proposed that the visual focus of attention is accompanied by a suppressive surround in spatial and feature dimensions. Follow-up studies have provided behavioural and neurophysiological evidence for this proposal (Carrasco, 2011; Tsotsos, 2011). ST also predicts that the size of the suppressive surround is determined by the level of processing within the visual hierarchy. We, thus, hypothesized that the size of the suppressive surround corresponds to the receptive field size of a neuron that best represents the attended stimulus. We conducted a free-viewing visual search task to test this hypothesis and used two different types of features processed at different levels - (early ventral) orientation and (late ventral) Greebles (Gauthier & Tarr, 1997). The sizes of search displays and stimuli were scaled depending on the feature levels to match the receptive field size of targeted neurons (V2 and LO). We tracked participants' eye movements during free-viewing visual search to find rapid return saccades from a distractor to a target. During search, if attention falls on a distractor (D1) such that a target lies within its suppressive surround, that target is invisible until attention is released. An eye movement then reveals the target and triggers a return saccade (Sheinberg & Logothetis, 2001). In other words, shifting gaze from D1 to another distractor (D2) releases the target from the suppression and a short latency (return) saccade to target occurs. The distribution of distances between the target and D1 when return saccades occur provides a measure for the size of the suppressive surround. Searching for Greebles produced much larger suppressive surrounds than orientation. This indicates that the size of the suppressive surround reflects the processing level of the attended stimulus supporting ST's original prediction. Meeting abstract presented at VSS 2015. PMID:26326949

  11. A Novel Function of Molecular Chaperone HSP70: SUPPRESSION OF ONCOGENIC FOXM1 AFTER PROTEOTOXIC STRESS.

    PubMed

    Halasi, Marianna; Váraljai, Renáta; Benevolenskaya, Elizaveta; Gartel, Andrei L

    2016-01-01

    The oncogenic transcription factor FOXM1 is overexpressed in the majority of human cancers, and it is a potential target for anticancer therapy. We identified proteasome inhibitors as the first type of drugs that target FOXM1 in cancer cells. Here we found that HSP90 inhibitor PF-4942847 and heat shock also suppress FOXM1. The common effector, which was induced after treatment with proteasome and HSP90 inhibitors or heat shock, was the molecular chaperone HSP70. We show that HSP70 binds to FOXM1 following proteotoxic stress and that HSP70 inhibits FOXM1 DNA-binding ability. Inhibition of FOXM1 transcriptional autoregulation by HSP70 leads to the suppression of FOXM1 protein expression. In addition, HSP70 suppression elevates FOXM1 expression, and simultaneous inhibition of FOXM1 and HSP70 increases the sensitivity of human cancer cells to anticancer drug-induced apoptosis. Overall, we determined the unique and novel mechanism of FOXM1 suppression by proteasome inhibitors. PMID:26559972

  12. Suppressed Charmed B Decay

    SciTech Connect

    Snoek, Hella Leonie; /Vrije U., Amsterdam

    2011-11-28

    This thesis describes the measurement of the branching fractions of the suppressed charmed B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decays and the non-resonant B{sup 0} {yields} D{sup (*)-} {eta}{pi}{sup +} decays in approximately 230 million {Upsilon}(4S) {yields} B{bar B} events. The data have been collected with the BABAR detector at the PEP-II B factory at the Stanford Linear Accelerator Center in California. Theoretical predictions of the branching fraction of the B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decays show large QCD model dependent uncertainties. Non-factorizing terms, in the naive factorization model, that can be calculated by QCD factorizing models have a large impact on the branching fraction of these decay modes. The predictions of the branching fractions are of the order of 10{sup -6}. The measurement of the branching fraction gives more insight into the theoretical models. In general a better understanding of QCD models will be necessary to conduct weak interaction physics at the next level. The presence of CP violation in electroweak interactions allows the differentiation between matter and antimatter in the laws of physics. In the Standard Model, CP violation is incorporated in the CKM matrix that describes the weak interaction between quarks. Relations amongst the CKM matrix elements are used to present the two relevant parameters as the apex of a triangle (Unitarity Triangle) in a complex plane. The over-constraining of the CKM triangle by experimental measurements is an important test of the Standard Model. At this moment no stringent direct measurements of the CKM angle {gamma}, one of the interior angles of the Unitarity Triangle, are available. The measurement of the angle {gamma} can be performed using the decays of neutral B mesons. The B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decay is sensitive to the angle {gamma} and, in comparison to the current decays that are being employed, could significantly enhance the measurement of this angle. However, the low expected branching fraction for the B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decay channels could severely impact the measurement. A prerequisite of the measurement of the CKM angle is the observation of the B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decay on which this thesis reports. The BABAR experiment consists of the BABAR detector and the PEP-II e{sup +}e{sup -} collider. The design of the experiment has been optimized for the study of CP violation in the decays of neutral B mesons but is also highly suitable for the search for rare B decays such as the B{sup 0} {yields} D{sup (*)-} a{sub 0}{sup +} decay. The PEP-II collider operates at the {Upsilon}(4S) resonance and is a clean source of B{bar B} meson pairs.

  13. Amotl2a interacts with the Hippo effector Yap1 and the Wnt/?-catenin effector Lef1 to control tissue size in zebrafish

    PubMed Central

    Agarwala, Sobhika; Duquesne, Sandra; Liu, Kun; Boehm, Anton; Grimm, Lin; Link, Sandra; König, Sabine; Eimer, Stefan; Ronneberger, Olaf; Lecaudey, Virginie

    2015-01-01

    During development, proliferation must be tightly controlled for organs to reach their appropriate size. While the Hippo signaling pathway plays a major role in organ growth control, how it senses and responds to increased cell density is still unclear. In this study, we use the zebrafish lateral line primordium (LLP), a group of migrating epithelial cells that form sensory organs, to understand how tissue growth is controlled during organ formation. Loss of the cell junction-associated Motin protein Amotl2a leads to overproliferation and bigger LLP, affecting the final pattern of sensory organs. Amotl2a function in the LLP is mediated together by the Hippo pathway effector Yap1 and the Wnt/?-catenin effector Lef1. Our results implicate for the first time the Hippo pathway in size regulation in the LL system. We further provide evidence that the Hippo/Motin interaction is essential to limit tissue size during development. DOI: http://dx.doi.org/10.7554/eLife.08201.001 PMID:26335201

  14. LFA-1 blockade induces effector and regulatory T-cell enrichment in lymph nodes and synergizes with CTLA-4Ig to inhibit effector function

    PubMed Central

    Reisman, Natalie M.; Floyd, Tamara L.; Wagener, Maylene E.; Kirk, Allan D.; Larsen, Christian P.

    2011-01-01

    Despite encouraging results using lymphocyte function antigen-1 (LFA-1) blockade to inhibit BM and solid organ transplantation rejection in nonhuman primates and humans, the precise mechanisms underlying its therapeutic potential are still poorly understood. Using a fully allogeneic murine transplantation model, we assessed the relative distribution of total lymphocyte subsets in untreated versus anti–LFA-1–treated animals. Our results demonstrated a striking loss of naive T cells from peripheral lymph nodes, a concomitant gain in blood after LFA-1 blockade, and a shift in phenotype of the cells remaining in the node to a CD62LloCD44hi profile. We determined that this change was due to a specific enrichment of activated, graft-specific effectors in the peripheral lymph nodes of anti–LFA-1–treated mice compared with untreated controls, and not to a direct effect of anti–LFA-1 on CD62L expression. LFA-1 blockade also resulted in a dramatic increase in the frequency of CD4+ FoxP3+ regulatory T cells in graft-draining nodes. Our results suggest that the differential impact of LFA-1 blockade on the distribution of naive versus effector and regulatory T cells may underlie its ability to inhibit alloreactive T-cell responses after transplantation. PMID:21972294

  15. A protein phosphatase 2C, responsive to the bacterial effector AvrRpm1 but not to the AvrB effector, regulates defense responses in Arabidopsis.

    PubMed

    Widjaja, Ivy; Lassowskat, Ines; Bethke, Gerit; Eschen-Lippold, Lennart; Long, Hoang-Hoa; Naumann, Kai; Dangl, Jeffery L; Scheel, Dierk; Lee, Justin

    2010-01-01

    Using a proteomics approach, a PP2C-type phosphatase (renamed PIA1, for PP2C induced by AvrRpm1) was identified that accumulates following infection by Pseudomonas syringae expressing the type III effector AvrRpm1, and subsequent activation of the corresponding plant NB-LRR disease resistance protein RPM1. No accumulation of PIA1 protein was seen following infection with P. syringae expressing AvrB, another type III effector that also activates RPM1, although PIA transcripts were observed. Accordingly, mutation of PIA1 resulted in enhanced RPM1 function in response to P. syringae pathover tomato (Pto) DC3000 (avrRpm1) but not to Pto DC3000 (avrB). Thus, PIA1 is a protein marker that distinguishes AvrRpm1- and AvrB-dependent activation of RPM1. AvrRpm1-induced expression of the pathogenesis-related genes PR1, PR2 and PR3, and salicylic acid accumulation were reduced in two pia1 mutants. By contrast, expression of other defense-related genes, including PR5 and PDF1.2 (plant defensin), was elevated in unchallenged pia1 mutants. Hence, PIA1 is required for AvrRpm1-induced responses, and confers dual (both positive and negative) regulation of defense gene expression. PMID:19843314

  16. Colletotrichum orbiculare Secretes Virulence Effectors to a Biotrophic Interface at the Primary Hyphal Neck via Exocytosis Coupled with SEC22-Mediated Traffic[W

    PubMed Central

    Irieda, Hiroki; Maeda, Hitomi; Akiyama, Kaoru; Hagiwara, Asuka; Saitoh, Hiromasa; Uemura, Aiko; Terauchi, Ryohei; Takano, Yoshitaka

    2014-01-01

    The hemibiotrophic pathogen Colletotrichum orbiculare develops biotrophic hyphae inside cucumber (Cucumis sativus) cells via appressorial penetration; later, the pathogen switches to necrotrophy. C. orbiculare also expresses specific effectors at different stages. Here, we found that virulence-related effectors of C. orbiculare accumulate in a pathogen–host biotrophic interface. Fluorescence-tagged effectors accumulated in a ring-like region around the neck of the biotrophic primary hyphae. Fluorescence imaging of cellular components and transmission electron microscopy showed that the ring-like signals of the effectors localized at the pathogen–plant interface. Effector accumulation at the interface required induction of its expression during the early biotrophic phase, suggesting that transcriptional regulation may link to effector localization. We also investigated the route of effector secretion to the interface. An exocytosis-related component, the Rab GTPase SEC4, localized to the necks of biotrophic primary hyphae adjacent to the interface, thereby suggesting focal effector secretion. Disruption of SEC4 in C. orbiculare reduced virulence and impaired effector delivery to the ring signal interface. Disruption of the v-SNARE SEC22 also reduced effector delivery. These findings suggest that biotrophy-expressed effectors are secreted, via the endoplasmic reticulum-to-Golgi route and subsequent exocytosis, toward the interface generated between C. orbiculare and the host cell. PMID:24850852

  17. The Crystal Structure of TAL Effector PthXo1 Bound to Its DNA Target

    SciTech Connect

    Mak, Amanda Nga-Sze; Bradley, Philip; Cernadas, Raul A.; Bogdanove, Adam J.; Stoddard, Barry L.

    2012-02-10

    DNA recognition by TAL effectors is mediated by tandem repeats, each 33 to 35 residues in length, that specify nucleotides via unique repeat-variable diresidues (RVDs). The crystal structure of PthXo1 bound to its DNA target was determined by high-throughput computational structure prediction and validated by heavy-atom derivatization. Each repeat forms a left-handed, two-helix bundle that presents an RVD-containing loop to the DNA. The repeats self-associate to form a right-handed superhelix wrapped around the DNA major groove. The first RVD residue forms a stabilizing contact with the protein backbone, while the second makes a base-specific contact to the DNA sense strand. Two degenerate amino-terminal repeats also interact with the DNA. Containing several RVDs and noncanonical associations, the structure illustrates the basis of TAL effector-DNA recognition.

  18. Tailored Immune Responses: Novel Effector Helper T Cell Subsets in Protective Immunity

    PubMed Central

    Kara, Ervin E.; Comerford, Iain; Fenix, Kevin A.; Bastow, Cameron R.; Gregor, Carly E.; McKenzie, Duncan R.; McColl, Shaun R.

    2014-01-01

    Differentiation of naïve CD4+ cells into functionally distinct effector helper T cell subsets, characterised by distinct “cytokine signatures,” is a cardinal strategy employed by the mammalian immune system to efficiently deal with the rapidly evolving array of pathogenic microorganisms encountered by the host. Since the TH1/TH2 paradigm was first described by Mosmann and Coffman, research in the field of helper T cell biology has grown exponentially with seven functionally unique subsets having now been described. In this review, recent insights into the molecular mechanisms that govern differentiation and function of effector helper T cell subsets will be discussed in the context of microbial infections, with a focus on how these different helper T cell subsets orchestrate immune responses tailored to combat the nature of the pathogenic threat encountered. PMID:24586147

  19. Task-level testing of the JPL-OMV smart end effector

    NASA Technical Reports Server (NTRS)

    Hannaford, B.

    1987-01-01

    An intelligent end effector previously developed at JPL has been tested in over 21 hours of experimental teleoperation. The end effector provides local control of gripper clamping force and a 6-degree-of-freedom, wrist mounted force torque sensor. Resolved forces and torques were displayed to the test subjects, and the effect of this information on their performance of simulated satellite servicing tasks was assessed. The experienced subjects accomplished the tasks with lower levels of Remote Manipulator System (RMS) forces than intermediate and naive subjects, but the force levels were apparently uncorrelated with the presence or absence of the display. This negative finding was attributed to the lack of a suitable control mode in the manipulator control system.

  20. Secreted effectors in Toxoplasma gondii and related species: determinants of host range and pathogenesis?

    PubMed Central

    English, E D; Adomako-Ankomah, Y; Boyle, J P

    2015-01-01

    Recent years have witnessed the discovery of a number of secreted proteins in Toxoplasma gondii that play important roles in host–pathogen interactions and parasite virulence, particularly in the mouse model. However, the role that these proteins play in driving the unique features of T. gondii compared to some of its nearest apicomplexan relatives (Hammondia hammondi and Neospora caninum) is unknown. These unique features include distinct dissemination characteristics in vivo and a vast host range. In this review we comprehensively survey what is known about disease outcome, the host response and host range for T. gondii, H. hammondi, and N. caninum. We then review what is presently known about recently identified secreted virulence effectors in these three genetically related, but phenotypically distinct, species. Finally we exploit the existence of genome sequences for these three organisms and discuss what is known about the presence, and functionality, of key T. gondii effectors in these three species. PMID:25655311

  1. Linear Scarifying End-Effector Developed For Wall Cleaning In Underground Storage Tanks

    SciTech Connect

    Fitzgerald, C.L.F.

    2001-02-04

    This paper describes the development and performance of a Linear Scarifying End-Effector (LSEE) designed and fabricated for deployment by a remotely operated vehicle. The end-effector was designed to blast or scarify in-grained residual contamination from gunite tank walls using high-pressure water jets after the bulk sludge had been removed from the tanks using an integrated suite of remotely operated tools. Two generations of the LSEE were fabricated, tested, and deployed in the gunite tanks at the Oak Ridge National Laboratory, with varying levels of success. Because the LSEE was designed near the end of a four-year project to clean up the gunite tanks at Oak Ridge, a number of design constraints existed. The end-effector had to utilize pneumatic, hydraulic and electrical interfaces already available at the site; and to be deployable through one of the containment structures already in place for the other remote systems. Another primary design consideration was that the tool had to effectively extend the reach of an existing remotely operated vehicle from six ft. to at least ten ft. to allow cleaning the tank walls from floor to ceiling. In addition, the combined weight and thrust of the LSEE had to be manageable by the manipulator mounted on the vehicle. Finally, the end-effector had to follow an autonomous scarifying path such that the vehicle was only required to reposition the unit at the end of each pass after the mist had cleared from the tank. The prototypes successfully met each of these challenges, but did encounter other difficulties during actual tank operations.

  2. Test plan for the remote conveyance and innovative end effector demonstration

    SciTech Connect

    Rice, P.; Smith, A.M.; Peterson, R.

    1994-08-01

    This test plan describes the demonstration of innovative equipment and processes specifically designed to be superior to currently employed technology for buried waste retrieval. The dumping of dry soil into a funnel/dumpster arrangement has been found to be the primary mechanism for dust generation during the retrieval of buried transuranic waste. The primary goal of the innovative end effector is to reduce dust generation and the potential spread of airborne contaminants during the dumping operation. In addition, regardless of the excavation technique, exhumed waste will have to be conveyed away from the retrieval area to a packaging area or directly to a treatment facility. The remote conveyance system is aimed at developing a remotely controlled vehicle to convey retrieved waste that will operate on variable terrain and remove workers from the hazardous zone. To demonstrate the remote conveyance system and the innovative end effector, the Buried Waste Integrated Demonstration (BWID) Program has subcontracted with RAHCO International to provide equipment and services to perform a demonstration of the technologies. The demonstration will be performed in two phases. In Phase I, the subcontractor will perform a full scale demonstration to assess the ability of the innovative end effector to control dust generation and the potential spread of contamination during dumping operations. Phase II includes performing a retrieval/conveyance demonstration. This demonstration will excavate, dump, and convey simulated waste to demonstrate the functionality of the system (e.g., maneuverability, retrieval rates, and system integration). Phase II of the demonstration will include all elements of the remote conveyance and end effector system. This test plan will describe the demonstration objectives, data quality objectives, equipment operation, and methods for collecting data during the demonstration.

  3. Complex structure of type VI peptidoglycan muramidase effector and a cognate immunity protein

    SciTech Connect

    Wang, Tianyu; Ding, Jinjing; Zhang, Ying; Wang, Da-Cheng; Liu, Wei

    2013-10-01

    The structure of the Tse3–Tsi3 complex associated with the bacterial type VI secretion system of P. aeruginosa has been solved and refined at 1.9 Å resolution. The structural basis of the recognition of the muramidase effector and its inactivation by its cognate immunity protein is revealed. The type VI secretion system (T6SS) is a bacterial protein-export machine that is capable of delivering virulence effectors between Gram-negative bacteria. The T6SS of Pseudomonas aeruginosa transports two lytic enzymes, Tse1 and Tse3, to degrade cell-wall peptidoglycan in the periplasm of rival bacteria that are competing for niches via amidase and muramidase activities, respectively. Two cognate immunity proteins, Tsi1 and Tsi3, are produced by the bacterium to inactivate the two antibacterial effectors, thereby protecting its siblings from self-intoxication. Recently, Tse1–Tsi1 has been structurally characterized. Here, the structure of the Tse3–Tsi3 complex is reported at 1.9 Å resolution. The results reveal that Tse3 contains a C-terminal catalytic domain that adopts a soluble lytic transglycosylase (SLT) fold in which three calcium-binding sites were surprisingly observed close to the catalytic Glu residue. The electrostatic properties of the substrate-binding groove are also distinctive from those of known structures with a similar fold. All of these features imply that a unique catalytic mechanism is utilized by Tse3 in cleaving glycosidic bonds. Tsi3 comprises a single domain showing a ?-sandwich architecture that is reminiscent of the immunoglobulin fold. Three loops of Tsi3 insert deeply into the groove of Tse3 and completely occlude its active site, which forms the structural basis of Tse3 inactivation. This work is the first crystallographic report describing the three-dimensional structure of the Tse3–Tsi3 effector–immunity pair.

  4. Global impact of Salmonella pathogenicity island 2-secreted effectors on the host phosphoproteome.

    PubMed

    Imami, Koshi; Bhavsar, Amit P; Yu, Hongbing; Brown, Nat F; Rogers, Lindsay D; Finlay, B Brett; Foster, Leonard J

    2013-06-01

    During the late stages of infection, Salmonella secretes numerous effectors through a type III secretion system that is encoded within Salmonella pathogenicity island 2 (SPI2). Despite the importance of SPI2 as a major virulence factor leading to the systemic spread of the bacteria and diseases, a global view of its effects on host responses is still lacking. Here, we measured global impacts of SPI2 effectors on the host phosphorylation and protein expression levels in RAW264.7 and in HeLa cells, as macrophage and nonphagocytic models of infection. We observe that SPI2 effectors differentially modulate the host phosphoproteome and cellular processes (e.g. protein trafficking, cytoskeletal regulation, and immune signaling) in a host cell-dependent manner. Our unbiased approach reveals the involvement of many previously unrecognized proteins, including E3 ligases (HERC4, RanBP2, and RAD18), kinases (CDK, SIK3, and WNK1), and histones (H2B1F, H4, and H15), in late stages of Salmonella infection. Furthermore, from this phosphoproteome analysis and other quantitative screens, we identified HSP27 as a direct in vitro and in vivo molecular target of the only type III secreted kinase, SteC. Using biochemical and cell biological assays, we demonstrate that SteC phosphorylates multiple sites in HSP27 and induces actin rearrangement through this protein. Together, these results provide a broader landscape of host players contributing to specific processes/pathways mediated by SPI2 effectors than was previously appreciated. PMID:23459991

  5. Migratory Properties of Naive, Effector, and Memory Cd8+ T Cells

    PubMed Central

    Weninger, Wolfgang; Crowley, Maura A.; Manjunath, N.; von Andrian, Ulrich H.

    2001-01-01

    It has been proposed that two different antigen-experienced T cell subsets may be distinguishable by their preferential ability to home to lymphoid organs (central memory cells) or nonlymphoid tissues (effector memory/effector cells). We have shown recently that murine antigen-primed CD8+ T cells cultured in interleukin (IL)-15 (CD8IL-15) resemble central memory cells in phenotype and function. In contrast, primed CD8+ T cells cultured in IL-2 (CD8IL-2) become cytotoxic effector cells. Here, the migratory behavior of these two subsets was investigated. Naive, CD8IL-15 cells and, to a lesser degree, CD8IL-2 cells localized to T cell areas in the spleen, but only naive and CD8IL-15 cells homed to lymph nodes (LNs) and Peyer's patches. Intravital microscopy of peripheral LNs revealed that CD8IL-15 cells, but not CD8IL-2 cells, rolled and arrested in high endothelial venules (HEVs). Migration of CD8IL-15 cells to LNs depended on L-selectin and required chemokines that bind CC chemokine receptor (CCR)7. Both antigen-experienced populations, but not naive T cells, responded to inflammatory chemokines and accumulated at sites of inflammation. However, CD8IL-2 cells were 12 times more efficient in migrating to inflamed peritoneum than CD8IL-15 cells. Furthermore, CD8IL-15 cells proliferated rapidly upon reencounter with antigen at sites of inflammation. Thus, central memory-like CD8IL-15 cells home avidly to lymphoid organs and moderately to sites of inflammation, where they mediate rapid recall responses, whereas CD8IL-2 effector T cells accumulate in inflamed tissues, but are excluded from most lymphoid organs. PMID:11581317

  6. The actin-binding protein UNC-115 is an effector of Rac signaling during axon pathfinding in C-elegans

    E-print Network

    Struckhoff, Eric Charles; Lundquist, Erik A.

    2003-02-01

    Rac GTPases control cell shape by regulating downstream effectors that influence the actin cytoskeleton. UNC-115, a putative actin-binding protein similar to human abLIM/limatin, has previously been implicated in axon pathfinding. We have discovered...

  7. Dynamics of G protein effector interactions and their impact on timing and sensitivity of G protein-mediated signal transduction.

    PubMed

    Bodmann, Eva-Lisa; Wolters, Valerie; Bünemann, Moritz

    2015-01-01

    G protein coupled receptors regulate numerous cellular functions primarily via coupling to heterotrimeric G proteins and subsequent regulation of effector proteins such as ion channels, enzymes or GTP exchange factors for small G proteins. The dynamics of interactions between G protein subunits and effectors have been difficult to study particularly in a cellular context. The introduction of Förster resonance energy transfer methods into the field of GPCR research led to interesting insights into the temporal patterns of interactions between G protein subunits and their effectors. In this review we specifically focus on the interaction of G?i subunits with adenylyl cyclases and of G?q subunits with p63RhoGEF or G protein coupled receptor kinases type 2. Comparing the dynamics of these interactions revealed remarkable differences between different G protein effectors regarding the ability to be modulated by members of the regulator of G protein signalling protein family as well as the sensitivity towards receptor activation. PMID:26074197

  8. Joint-space adaptive control of a 6 DOF end-effector with closed-kinematic chain mechanism

    NASA Technical Reports Server (NTRS)

    Nguyen, Charles C.; Zhou, Zhen-Lei

    1989-01-01

    The development is presented for a joint-space adaptive scheme that controls the joint position of a six-degree-of-freedom (DOF) robot end-effector performing fine and precise motion within a very limited workspace. The end-effector was built to study autonomous assembly of NASA hardware in space. The design of the adaptive controller is based on the concept of model reference adaptive control (MRAC) and Lyapunov direct method. In the development, it is assumed that the end-effector performs slowly varying motion. Computer simulation is performed to investigate the performance of the developed control scheme on position control of the end-effector. Simulation results manifest that the adaptive control scheme provides excellent tracking of several test paths.

  9. Host-Induced gene silencing in barley powdery mildew reveals a class of ribonuclease-like effectors

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Obligate biotrophic pathogens of plants require the ability to circumvent host defenses to enable colonization. To establish compatibility, pathogens secrete a variety of effectors, which regulate host immunity, and thus, facilitate the establishment of haustorial feeding structures. These structur...

  10. Genome-wide assessment of differential effector gene use in embryogenesis.

    PubMed

    Barsi, Julius C; Tu, Qiang; Calestani, Cristina; Davidson, Eric H

    2015-11-15

    Six different populations of cells were isolated by fluorescence-activated cell sorting from disaggregated late blastula- and gastrula-stage sea urchin embryos according to the regulatory states expressed in these cells, as reported by recombineered bacterial artificial chromosomes producing fluorochromes. Transcriptomes recovered from these embryonic cell populations revealed striking, early differential expression of large cohorts of effector genes. The six cell populations were presumptive pigment cells, presumptive neurogenic cells, presumptive skeletogenic cells, cells from the stomodeal region of the oral ectoderm, ciliated band cells and cells from the endoderm/ectoderm boundary that will give rise both to hindgut and to border ectoderm. Transcriptome analysis revealed that each of these domains specifically expressed several hundred effector genes at significant levels. Annotation indicated the qualitative individuality of the functional nature of each cell population, even though they were isolated from embryos only 1-2?days old. In no case was more than a tiny fraction of the transcripts enriched in one population also enriched in any other of the six populations studied. As was particularly clear in the cases of the presumptive pigment, neurogenic and skeletogenic cells, all three of which represent precociously differentiating cell types of this embryo, most specifically expressed genes of given cell types are not significantly expressed at all in the other cell types. Thus, at the effector gene level, a dramatic, cell type-specific pattern of differential gene regulation is established well before any significant embryonic morphogenesis has occurred. PMID:26417044

  11. The haustorial transcriptomes of Uromyces appendiculatus and Phakopsora pachyrhizi and their candidate effector families.

    PubMed

    Link, Tobias I; Lang, Patrick; Scheffler, Brian E; Duke, Mary V; Graham, Michelle A; Cooper, Bret; Tucker, Mark L; van de Mortel, Martijn; Voegele, Ralf T; Mendgen, Kurt; Baum, Thomas J; Whitham, Steven A

    2014-05-01

    Haustoria of biotrophic rust fungi are responsible for the uptake of nutrients from their hosts and for the production of secreted proteins, known as effectors, which modulate the host immune system. The identification of the transcriptome of haustoria and an understanding of the functions of expressed genes therefore hold essential keys for the elucidation of fungus-plant interactions and the development of novel fungal control strategies. Here, we purified haustoria from infected leaves and used 454 sequencing to examine the haustorial transcriptomes of Phakopsora pachyrhizi and Uromyces appendiculatus, the causal agents of soybean rust and common bean rust, respectively. These pathogens cause extensive yield losses in their respective legume crop hosts. A series of analyses were used to annotate expressed sequences, including transposable elements and viruses, to predict secreted proteins from the assembled sequences and to identify families of candidate effectors. This work provides a foundation for the comparative analysis of haustorial gene expression with further insights into physiology and effector evolution. PMID:24341524

  12. An Effector Peptide Family Required for Drosophila Toll-Mediated Immunity

    PubMed Central

    Wasserman, Steven A.

    2015-01-01

    In Drosophila melanogaster, recognition of an invading pathogen activates the Toll or Imd signaling pathway, triggering robust upregulation of innate immune effectors. Although the mechanisms of pathogen recognition and signaling are now well understood, the functions of the immune-induced transcriptome and proteome remain much less well characterized. Through bioinformatic analysis of effector gene sequences, we have defined a family of twelve genes – the Bomanins (Boms) – that are specifically induced by Toll and that encode small, secreted peptides of unknown biochemical activity. Using targeted genome engineering, we have deleted ten of the twelve Bom genes. Remarkably, inactivating these ten genes decreases survival upon microbial infection to the same extent, and with the same specificity, as does eliminating Toll pathway function. Toll signaling, however, appears unaffected. Assaying bacterial load post-infection in wild-type and mutant flies, we provide evidence that the Boms are required for resistance to, rather than tolerance of, infection. In addition, by generating and assaying a deletion of a smaller subset of the Bom genes, we find that there is overlap in Bom activity toward particular pathogens. Together, these studies deepen our understanding of Toll-mediated immunity and provide a new in vivo model for exploration of the innate immune effector repertoire. PMID:25915418

  13. Entamoeba histolytica RacC selectively engages p21-activated kinase effectors.

    PubMed

    Bosch, Dustin E; Siderovski, David P

    2015-01-20

    Rho family GTPases modulate actin cytoskeleton dynamics by signaling through multiple effectors, including the p21-activated kinases (PAKs). The intestinal parasite Entamoeba histolytica expresses ?20 Rho family GTPases and seven isoforms of PAK, two of which have been implicated in pathogenesis-related processes such as amoebic motility and invasion and host cell phagocytosis. Here, we describe two previously unstudied PAK isoforms, EhPAK4 and EhPAK5, as highly specific effectors of EhRacC. A structural model based on 2.35 Å X-ray crystallographic data of a complex between EhRacC(Q65L)·GTP and the EhPAK4 p21 binding domain (PBD) reveals a fairly well-conserved Rho/effector interface despite deviation of the PBD ?-helix. A structural comparison with EhRho1 in complex with EhFormin1 suggests likely determinants of Rho family GTPase signaling specificity in E. histolytica. These findings suggest a high degree of Rho family GTPase diversity and specificity in the single-cell parasite E. histolytica. Because PAKs regulate pathogenesis-related processes in E. histolytica, they may be valid pharmacologic targets for anti-amoebiasis drugs. PMID:25529118

  14. Effector granules in human T lymphocytes: the luminal proteome of secretory lysosomes from human T cells

    PubMed Central

    2011-01-01

    Background Cytotoxic cells of the immune system have evolved a lysosomal compartment to store and mobilize effector molecules. In T lymphocytes and NK cells, the death factor FasL is one of the characteristic marker proteins of these so-called secretory lysosomes, which combine properties of conventional lysosomes and exocytotic vesicles. Although these vesicles are crucial for immune effector function, their protein content in T cells has so far not been investigated in detail. Results In the present study, intact membranous vesicles were enriched from homogenates of polyclonally activated T cells and initially characterized by Western blotting and electron microscopic inspection. The vesicular fraction that contained the marker proteins of secretory lysosomes was subsequently analyzed by 2D electrophoresis and mass spectrometry. The proteome analysis and data evaluation revealed that 70% of the 397 annotated proteins had been associated with different lysosome-related organelles in previous proteome studies. Conclusion We provide the first comprehensive proteome map of T cell-derived secretory lysosomes with only minor contaminations by cytosolic, nuclear or other proteins. This information will be useful to more precisely address the activation-dependent maturation and the specific distribution of effector organelles and proteins in individual T or NK cell populations in future studies. PMID:21255389

  15. miR-150 Regulates Differentiation and Cytolytic Effector Function in CD8+ T cells

    PubMed Central

    Smith, Norah L.; Wissink, Erin M.; Grimson, Andrew; Rudd, Brian D.

    2015-01-01

    MicroRNAs regulate most mammalian genes, and they control numerous aspects of immune system development and function. Their precise roles in the CD8+ T cell response, however, remain unclear. In this report, we show that in the absence of the microRNA miR-150, CD8+ T cells fail to undergo robust expansion and differentiation into short-lived terminal effector cells in response to primary infection with Listeria monocytogenes or Vaccinia virus. Notably, even after transitioning into the memory pool, miR-150?/? cells still mount a weaker recall response to secondary infection, and remain less differentiated than their wild-type counterparts. Transcriptome analysis shows miR-150 gene targets are globally upregulated in cells lacking miR-150, and amongst these targets, we found misregulation of genes associated with proliferation and effector cell function. These transcriptome data suggest that miR-150 deficient CD8+ T cells are less efficient in killing infected cells, which we validate experimentally. Together, these results reveal a cell-intrinsic role for miR-150 in the regulation of effector CD8+ T cell fate and function. PMID:26549197

  16. Subversion of Cell-Autonomous Immunity and Cell Migration by Legionella pneumophila Effectors

    PubMed Central

    Simon, Sylvia; Hilbi, Hubert

    2015-01-01

    Bacteria trigger host defense and inflammatory processes, such as cytokine production, pyroptosis, and the chemotactic migration of immune cells toward the source of infection. However, a number of pathogens interfere with these immune functions by producing specific so-called “effector” proteins, which are delivered to host cells via dedicated secretion systems. Air-borne Legionella pneumophila bacteria trigger an acute and potential fatal inflammation in the lung termed Legionnaires’ disease. The opportunistic pathogen L. pneumophila is a natural parasite of free-living amoebae, but also replicates in alveolar macrophages and accidentally infects humans. The bacteria employ the intracellular multiplication/defective for organelle trafficking (Icm/Dot) type IV secretion system and as many as 300 different effector proteins to govern host–cell interactions and establish in phagocytes an intracellular replication niche, the Legionella-containing vacuole. Some Icm/Dot-translocated effector proteins target cell-autonomous immunity or cell migration, i.e., they interfere with (i) endocytic, secretory, or retrograde vesicle trafficking pathways, (ii) organelle or cell motility, (iii) the inflammasome and programed cell death, or (iv) the transcription factor NF-?B. Here, we review recent mechanistic insights into the subversion of cellular immune functions by L. pneumophila. PMID:26441958

  17. Structure of the Legionella Effector, lpg1496, Suggests a Role in Nucleotide Metabolism.

    PubMed

    Wong, Kathy; Kozlov, Guennadi; Zhang, Yinglu; Gehring, Kalle

    2015-10-01

    Pathogenic Gram-negative bacteria use specialized secretion systems that translocate bacterial proteins, termed effectors, directly into host cells where they interact with host proteins and biochemical processes for the benefit of the pathogen. lpg1496 is a previously uncharacterized effector of Legionella pneumophila, the causative agent of Legionnaires disease. Here, we crystallized three nucleotide binding domains from lpg1496. The C-terminal domain, which is conserved among the SidE family of effectors, is formed of two largely ?-helical lobes with a nucleotide binding cleft. A structural homology search has shown similarity to phosphodiesterases involved in cleavage of cyclic nucleotides. We have also crystallized a novel domain that occurs twice in the N-terminal half of the protein that we term the KLAMP domain due to the presence of homologous domains in bacterial histidine kinase-like ATP binding region-containing proteins and S-adenosylmethionine-dependent methyltransferase proteins. Both KLAMP structures are very similar but selectively bind 3',5'-cAMP and ADP. A co-crystal of the KLAMP1 domain with 3',5'-cAMP reveals the contribution of Tyr-61 and Tyr-69 that produces ?-stacking interactions with the adenine ring of the nucleotide. Our study provides the first structural insights into two novel nucleotide binding domains associated with bacterial virulence. PMID:26294765

  18. Complement Effectors of Inflammation in Cystic Fibrosis Lung Fluid Correlate with Clinical Measures of Disease

    PubMed Central

    Sass, Laura A.; Hair, Pamela S.; Perkins, Amy M.; Shah, Tushar A.; Krishna, Neel K.; Cunnion, Kenji M.

    2015-01-01

    In cystic fibrosis (CF), lung damage is mediated by a cycle of obstruction, infection, and inflammation. Here we explored complement inflammatory effectors in CF lung fluid. In this study soluble fractions (sols) from sputum samples of 15 CF patients were assayed for complement effectors and analyzed with clinical measurements. The pro-inflammatory peptide C5a was increased 4.8-fold (P = 0.04) in CF sols compared with controls. Incubation of CF sols with P. aeruginosa or S. aureus increased C5a concentration 2.3-fold (P = 0.02). A peptide inhibitor of complement C1 (PIC1) completely blocked the increase in C5a concentration from P. aeruginosa in CF sol in vitro (P = 0.001). C5a concentration in CF sol correlated inversely with body mass index (BMI) percentile in children (r = -0.77, P = 0.04). C3a, which has anti-inflammatory effects, correlated positively with FEV1% predicted (rs = 0.63, P = 0.02). These results suggest that complement effectors may significantly impact inflammation in CF lung fluid. PMID:26642048

  19. The Salmonella effector protein SifA plays a dual role in virulence

    PubMed Central

    Zhao, Weidong; Moest, Thomas; Zhao, Yaya; Guilhon, Aude-Agnès; Buffat, Christophe; Gorvel, Jean-Pierre; Méresse, Stéphane

    2015-01-01

    The virulence of Salmonella relies on the expression of effector proteins that the bacterium injects inside infected cells. Salmonella enters eukaryotic cells and resides in a vacuolar compartment on which a number of effector proteins such as SifA are found. SifA plays an essential role in Salmonella virulence. It is made of two distinct domains. The N-terminal domain of SifA interacts with the host protein SKIP. This interaction regulates vacuolar membrane dynamics. The C-terminal has a fold similar to other bacterial effector domains having a guanine nucleotide exchange factor activity. Although SifA interacts with RhoA, it does not stimulate the dissociation of GDP and the activation of this GTPase. Hence it remains unknown whether the C-terminal domain contributes to the function of SifA in virulence. We used a model of SKIP knockout mice to show that this protein mediates the host susceptibility to salmonellosis and to establish that SifA also contributes to Salmonella virulence independently of its interaction with SKIP. We establish that the C-terminal domain of SifA mediates this SKIP-independent contribution. Moreover, we show that the two domains of SifA are functionally linked and participate to the same signalling cascade that supports Salmonella virulence. PMID:26268777

  20. miR-150 Regulates Differentiation and Cytolytic Effector Function in CD8+ T cells.

    PubMed

    Smith, Norah L; Wissink, Erin M; Grimson, Andrew; Rudd, Brian D

    2015-01-01

    MicroRNAs regulate most mammalian genes, and they control numerous aspects of immune system development and function. Their precise roles in the CD8+ T cell response, however, remain unclear. In this report, we show that in the absence of the microRNA miR-150, CD8+ T cells fail to undergo robust expansion and differentiation into short-lived terminal effector cells in response to primary infection with Listeria monocytogenes or Vaccinia virus. Notably, even after transitioning into the memory pool, miR-150(-/-) cells still mount a weaker recall response to secondary infection, and remain less differentiated than their wild-type counterparts. Transcriptome analysis shows miR-150 gene targets are globally upregulated in cells lacking miR-150, and amongst these targets, we found misregulation of genes associated with proliferation and effector cell function. These transcriptome data suggest that miR-150 deficient CD8+ T cells are less efficient in killing infected cells, which we validate experimentally. Together, these results reveal a cell-intrinsic role for miR-150 in the regulation of effector CD8+ T cell fate and function. PMID:26549197

  1. Activation of effector immune cells promotes tumor stochastic extinction: A homotopy analysis approach

    E-print Network

    Josep Sardanyés; Carla Rodrigues; Cristina Januário; Nuno Martins; Gabriel Gil-Gómez; Jorge Duarte

    2014-11-28

    In this article we provide homotopy solutions of a cancer nonlinear model describing the dynamics of tumor cells in interaction with healthy and effector immune cells. We apply a semi-analytic technique for solving strongly nonlinear systems - the Step Homotopy Analysis Method (SHAM). This algorithm, based on a modification of the standard homotopy analysis method (HAM), allows to obtain a one-parameter family of explicit series solutions. By using the homotopy solutions, we first investigate the dynamical effect of the activation of the effector immune cells in the deterministic dynamics, showing that an increased activation makes the system to enter into chaotic dynamics via a period-doubling bifurcation scenario. Then, by adding demographic stochasticity into the homotopy solutions, we show, as a difference from the deterministic dynamics, that an increased activation of the immune cells facilitates cancer clearance involving tumor cells extinction and healthy cells persistence. Our results highlight the importance of therapies activating the effector immune cells at early stages of cancer progression.

  2. Bacterial Internalization, Localization, and Effectors Shape the Epithelial Immune Response during Shigella flexneri Infection.

    PubMed

    Lippmann, Juliane; Gwinner, Frederik; Rey, Camille; Tamir, Uyanga; Law, Helen K W; Schwikowski, Benno; Enninga, Jost

    2015-09-01

    Intracellular pathogens are differentially sensed by the compartmentalized host immune system. Nevertheless, gene expression studies of infected cells commonly average the immune responses, neglecting the precise pathogen localization. To overcome this limitation, we dissected the transcriptional immune response to Shigella flexneri across different infection stages in bulk and single cells. This identified six distinct transcriptional profiles characterizing the dynamic, multilayered host response in both bystander and infected cells. These profiles were regulated by external and internal danger signals, as well as whether bacteria were membrane bound or cytosolic. We found that bacterial internalization triggers a complex, effector-independent response in bystander cells, possibly to compensate for the undermined host gene expression in infected cells caused by bacterial effectors, particularly OspF. Single-cell analysis revealed an important bacterial strategy to subvert host responses in infected cells, demonstrating that OspF disrupts concomitant gene expression of proinflammatory, apoptosis, and stress pathways within cells. This study points to novel mechanisms through which bacterial internalization, localization, and injected effectors orchestrate immune response transcriptional signatures. PMID:26123804

  3. Coevolution between a Family of Parasite Virulence Effectors and a Class of LINE-1 Retrotransposons

    PubMed Central

    Pedersen, Carsten; Skamnioti, Pari; Thordal-Christensen, Hans; Micali, Cristina; Brown, James K. M.; Ridout, Christopher J.

    2009-01-01

    Parasites are able to evolve rapidly and overcome host defense mechanisms, but the molecular basis of this adaptation is poorly understood. Powdery mildew fungi (Erysiphales, Ascomycota) are obligate biotrophic parasites infecting nearly 10,000 plant genera. They obtain their nutrients from host plants through specialized feeding structures known as haustoria. We previously identified the AVRk1 powdery mildew-specific gene family encoding effectors that contribute to the successful establishment of haustoria. Here, we report the extensive proliferation of the AVRk1 gene family throughout the genome of B. graminis, with sequences diverging in formae speciales adapted to infect different hosts. Also, importantly, we have discovered that the effectors have coevolved with a particular family of LINE-1 retrotransposons, named TE1a. The coevolution of these two entities indicates a mutual benefit to the association, which could ultimately contribute to parasite adaptation and success. We propose that the association would benefit 1) the powdery mildew fungus, by providing a mechanism for amplifying and diversifying effectors and 2) the associated retrotransposons, by providing a basis for their maintenance through selection in the fungal genome. PMID:19829700

  4. Engineering development of waste retrieval end effectors for the Oak Ridge gunite waste tanks

    SciTech Connect

    Mullen, O.D.

    1997-05-01

    The Gunite and Associated Tanks Treatability Study at Oak Ridge National Laboratory selected the waterjet scarifying end effector, the jet pump conveyance system, and the Modified Light Duty Utility Arm and Houdini Remotely Operated Vehicle deployment and manipulator systems for evaluation. The waterjet-based retrieval end effector had been developed through several generations of test articles targeted at deployment in Hanford underground storage tanks with a large robotic arm. The basic technology had demonstrated effectiveness at retrieval of simulants bounding the foreseen range of waste properties and indicated compatibility with the planned deployment systems. The Retrieval Process Development and Enhancements team was tasked with developing a version of the retrieval end effector tailored to the Oak Ridge tanks, waste and deployment platforms. The finished prototype was delivered to PNNL and subjected to a brief round of characterization and performance testing at the Hydraulic Testbed prior to shipment to Oak Ridge. It has undergone extensive operational testing in the Oak Ridge National Laboratory Tanks Technology Cold Test Facility and performed well, as expected. A second unit has been delivered outfitted with the high pressure manifold.

  5. Differential expression of candidate salivary effector proteins in field collections of Hessian fly, Mayetiola destructor

    PubMed Central

    Johnson, A J; Shukle, R H; Chen, M-S; Srivastava, S; Subramanyam, S; Schemerhorn, B J; Weintraub, P G; Abdel Moniem, H E M; Flanders, K L; Buntin, G D; Williams, C E

    2015-01-01

    Evidence is emerging that some proteins secreted by gall-forming parasites of plants act as effectors responsible for systemic changes in the host plant, such as galling and nutrient tissue formation. A large number of secreted salivary gland proteins (SSGPs) that are the putative effectors responsible for the physiological changes elicited in susceptible seedling wheat by Hessian fly, Mayetiola destructor (Say), larvae have been documented. However, how the genes encoding these candidate effectors might respond under field conditions is unknown. The goal of this study was to use microarray analysis to investigate variation in SSGP transcript abundance amongst field collections from different geographical regions (southeastern USA, central USA, and the Middle East). Results revealed significant variation in SSGP transcript abundance amongst the field collections studied. The field collections separated into three distinct groups that corresponded to the wheat classes grown in the different geographical regions as well as to recently described Hessian fly populations. These data support previous reports correlating Hessian fly population structure with micropopulation differences owing to agro-ecosystem parameters such as cultivation of regionally adapted wheat varieties, deployment of resistance genes and variation in climatic conditions. PMID:25528896

  6. Targeting an antimicrobial effector function in insect immunity as a pest control strategy

    PubMed Central

    Bulmer, Mark S.; Bachelet, Ido; Raman, Rahul; Rosengaus, Rebeca B.; Sasisekharan, Ram

    2009-01-01

    Insect pests such as termites cause damages to crops and man-made structures estimated at over $30 billion per year, imposing a global challenge for the human economy. Here, we report a strategy for compromising insect immunity that might lead to the development of nontoxic, sustainable pest control methods. Gram-negative bacteria binding proteins (GNBPs) are critical for sensing pathogenic infection and triggering effector responses. We report that termite GNBP-2 (tGNBP-2) shows ?(1,3)-glucanase effector activity previously unknown in animal immunity and is a pleiotropic pattern recognition receptor and an antimicrobial effector protein. Termites incorporate this protein into the nest building material, where it functions as a nest-embedded sensor that cleaves and releases pathogenic components, priming termites for improved antimicrobial defense. By means of rational design, we present an inexpensive, nontoxic small molecule glycomimetic that blocks tGNBP-2, thus exposing termites in vivo to accelerated infection and death from specific and opportunistic pathogens. Such a molecule, introduced into building materials and agricultural methods, could protect valuable assets from insect pests. PMID:19506247

  7. The Machinery at Endoplasmic Reticulum-Plasma Membrane Contact Sites Contributes to Spatial Regulation of Multiple Legionella Effector Proteins

    PubMed Central

    Hubber, Andree; Arasaki, Kohei; Nakatsu, Fubito; Hardiman, Camille; Lambright, David; De Camilli, Pietro; Nagai, Hiroki; Roy, Craig R.

    2014-01-01

    The Dot/Icm system of the intracellular pathogen Legionella pneumophila has the capacity to deliver over 270 effector proteins into host cells during infection. Important questions remain as to spatial and temporal mechanisms used to regulate such a large array of virulence determinants after they have been delivered into host cells. Here we investigated several L. pneumophila effector proteins that contain a conserved phosphatidylinositol-4-phosphate (PI4P)-binding domain first described in the effector DrrA (SidM). This PI4P binding domain was essential for the localization of effectors to the early L. pneumophila-containing vacuole (LCV), and DrrA-mediated recruitment of Rab1 to the LCV required PI4P-binding activity. It was found that the host cell machinery that regulates sites of contact between the plasma membrane (PM) and the endoplasmic reticulum (ER) modulates PI4P dynamics on the LCV to control localization of these effectors. Specifically, phosphatidylinositol-4-kinase III? (PI4KIII?) was important for generating a PI4P signature that enabled L. pneumophila effectors to localize to the PM-derived vacuole, and the ER-associated phosphatase Sac1 was involved in metabolizing the PI4P on the vacuole to promote the dissociation of effectors. A defect in L. pneumophila replication in macrophages deficient in PI4KIII? was observed, highlighting that a PM-derived PI4P signature is critical for biogenesis of a vacuole that supports intracellular multiplication of L. pneumophila. These data indicate that PI4P metabolism by enzymes controlling PM-ER contact sites regulate the association of L. pneumophila effectors to coordinate early stages of vacuole biogenesis. PMID:24992562

  8. Available online at www.sciencedirect.com How are TH1 and TH2 effector cells made?

    E-print Network

    Spilianakis, Charalampos G. "Babis"

    to parasites [1]. Cytokine induced TH1/TH2 induction Activation of nai¨ve CD4+ T cells in vitro in the presenceAvailable online at www.sciencedirect.com How are TH1 and TH2 effector cells made? Derk Amsen1 , Charalampos G Spilianakis2 and Richard A Flavell3,4 Differentiation of TH1 and TH2 effector cells proceeds

  9. Comparative Analysis of Type III Secreted Effector Genes Reflects Divergence of Acidovorax citrulli Strains into Three Distinct Lineages.

    PubMed

    Eckshtain-Levi, Noam; Munitz, Tamar; Zivanovi?, Marija; Traore, Sy M; Spröer, Cathrin; Zhao, Bingyu; Welbaum, Gregory; Walcott, Ron; Sikorski, Johannes; Burdman, Saul

    2014-11-01

    ABSTRACT Acidovorax citrulli causes bacterial fruit blotch of cucurbits, a serious economic threat to watermelon (Citrullus lanatus) and melon (Cucumis melo) production worldwide. Based on genetic and biochemical traits, A. citrulli strains have been divided into two distinct groups: group I strains have been mainly isolated from various non-watermelon hosts, while group II strains have been generally isolated from and are highly virulent on watermelon. The pathogen depends on a functional type III secretion system for pathogenicity. Annotation of the genome of the group II strain AAC00-1 revealed 11 genes encoding putative type III secreted (T3S) effectors. Due to the crucial role of type III secretion for A. citrulli pathogenicity, we hypothesized that group I and II strains differ in their T3S effector repertoire. Comparative analysis of the 11 effector genes from a collection of 22 A. citrulli strains confirmed this hypothesis. Moreover, this analysis led to the identification of a third A. citrulli group, which was supported by DNA:DNA hybridization, DNA fingerprinting, multilocus sequence analysis of conserved genes, and virulence assays. The effector genes assessed in this study are homologous to effectors from other plant-pathogenic bacteria, mainly belonging to Xanthomonas spp. and Ralstonia solanacearum. Analyses of the effective number of codons and gas chromatography content of effector genes relative to a representative set of housekeeping genes support the idea that these effector genes were acquired by lateral gene transfer. Further investigation is required to identify new T3S effectors of A. citrulli and to determine their contribution to virulence and host preferential association. PMID:24848275

  10. Suppression of T cell responses in the tumor microenvironment.

    PubMed

    Frey, Alan B

    2015-12-16

    The immune system recognizes protein antigens expressed in transformed cells evidenced by accumulation of antigen-specific T cells in tumor and tumor draining lymph nodes. However, despite demonstrable immune response, cancers grow progressively suggesting that priming of antitumor immunity is insufficiently vigorous or that antitumor immunity is suppressed, or both. Compared to virus infection, antitumor T cells are low abundance that likely contributes to tumor escape and enhancement of priming is a long-sought goal of experimental vaccination therapy. Furthermore, patient treatment with antigen-specific T cells can in some cases overcome deficient priming and cause tumor regression supporting the notion that low numbers of T cells permits tumor outgrowth. However, tumor-induced suppression of antitumor immune response is now recognized as a significant factor contributing to cancer growth and reversal of the inhibitory influences within the tumor microenvironment is a major research objective. Multiple cell types and factors can inhibit T cell functions in tumors and may be grouped in two general classes: T cell intrinsic and T cell extrinsic. T cell intrinsic factors are exemplified by T cell expression of cell surface inhibitory signaling receptors that, after contact with cells expressing a cognate ligand, inactivate proximal T Cell Receptor-mediated signal transduction therein rendering T cells dysfunctional. T cell extrinsic factors are more diverse in nature and are produced by tumors and various non-tumor cells in the tumor microenvironment. These include proteins secreted by tumor or stromal cells, highly reactive soluble oxygen and nitrogen species, cytokines, chemokines, gangliosides, and toxic metabolites. These factors may restrict T cell entrance into the tumor parenchyma, cause inactivation of effector phase T cell functions, or induce T cell apoptosis ultimately causing diminished cancer elimination. Here, we review the contributions of inhibitory factors to tumor T cell dysfunction leading to tumor escape. PMID:26403368

  11. Creating a customized intracellular niche: subversion of host cell signaling by Legionella type IV secretion system effectors.

    PubMed

    So, Ernest C; Mattheis, Corinna; Tate, Edward W; Frankel, Gad; Schroeder, Gunnar N

    2015-09-01

    The Gram-negative facultative intracellular pathogen Legionella pneumophila infects a wide range of different protozoa in the environment and also human alveolar macrophages upon inhalation of contaminated aerosols. Inside its hosts, it creates a defined and unique compartment, termed the Legionella-containing vacuole (LCV), for survival and replication. To establish the LCV, L. pneumophila uses its Dot/Icm type IV secretion system (T4SS) to translocate more than 300 effector proteins into the host cell. Although it has become apparent in the past years that these effectors subvert a multitude of cellular processes and allow Legionella to take control of host cell vesicle trafficking, transcription, and translation, the exact function of the vast majority of effectors still remains unknown. This is partly due to high functional redundancy among the effectors, which renders conventional genetic approaches to elucidate their role ineffective. Here, we review the current knowledge about Legionella T4SS effectors, highlight open questions, and discuss new methods that promise to facilitate the characterization of T4SS effector functions in the future. PMID:26059316

  12. Convergent targeting of a common host protein-network by pathogen effectors from three kingdoms of life

    PubMed Central

    Weßling, Ralf; Epple, Petra; Altmann, Stefan; He, Yijian; Yang, Li; Henz, Stefan R.; McDonald, Nathan; Wiley, Kristin; Bader, Kai Christian; Gläßer, Christine; Mukhtar, M. Shahid; Haigis, Sabine; Ghamsari, Lila; Stephens, Amber E.; Ecker, Joseph R.; Vidal, Marc; Jones, Jonathan D. G.; Mayer, Klaus F. X.; van Themaat, Emiel Ver Loren; Weigel, Detlef; Schulze-Lefert, Paul; Dangl, Jeffery L.; Panstruga, Ralph; Braun, Pascal

    2014-01-01

    SUMMARY While conceptual principles governing plant immunity are becoming clear, its systems-level organization and the evolutionary dynamic of the host-pathogen interface are still obscure. We generated a systematic protein-protein interaction network of virulence effectors from the ascomycete pathogen Golovinomyces orontii and Arabidopsis thaliana host proteins. We combined this dataset with corresponding data for the eubacterial pathogen Pseudomonas syringae and the oomycete pathogen Hyaloperonospora arabidopsidis. The resulting network identifies host proteins onto which intraspecies and interspecies pathogen effectors converge. Phenotyping of 124 Arabidopsis effector-interactor mutants revealed a correlation between intra- and interspecies convergence and several altered immune response phenotypes. The effectors and most heavily targeted host protein co-localized in sub-nuclear foci. Products of adaptively selected Arabidopsis genes are enriched for interactions with effector targets. Our data suggest the existence of a molecular host-pathogen interface that is conserved across Arabidopsis accessions, while evolutionary adaptation occurs in the immediate network neighborhood of effector targets. PMID:25211078

  13. Phytoplasma Effector SAP54 Induces Indeterminate Leaf-Like Flower Development in Arabidopsis Plants1[C][W][OA

    PubMed Central

    MacLean, Allyson M.; Sugio, Akiko; Makarova, Olga V.; Findlay, Kim C.; Grieve, Victoria M.; Tóth, Réka; Nicolaisen, Mogens; Hogenhout, Saskia A.

    2011-01-01

    Phytoplasmas are insect-transmitted bacterial plant pathogens that cause considerable damage to a diverse range of agricultural crops globally. Symptoms induced in infected plants suggest that these phytopathogens may modulate developmental processes within the plant host. We report herein that Aster Yellows phytoplasma strain Witches’ Broom (AY-WB) readily infects the model plant Arabidopsis (Arabidopsis thaliana) ecotype Columbia, inducing symptoms that are characteristic of phytoplasma infection, such as the production of green leaf-like flowers (virescence and phyllody) and increased formation of stems and branches (witches’ broom). We found that the majority of genes encoding secreted AY-WB proteins (SAPs), which are candidate effector proteins, are expressed in Arabidopsis and the AY-WB insect vector Macrosteles quadrilineatus (Hemiptera; Cicadellidae). To identify which of these effector proteins induce symptoms of phyllody and virescence, we individually expressed the effector genes in Arabidopsis. From this screen, we have identified a novel AY-WB effector protein, SAP54, that alters floral development, resulting in the production of leaf-like flowers that are similar to those produced by plants infected with this phytoplasma. This study offers novel insight into the effector profile of an insect-transmitted plant pathogen and reports to our knowledge the first example of a microbial pathogen effector protein that targets flower development in a host. PMID:21849514

  14. Bio-effectors from waste materials as growth promoters, an agronomic and metabolomic study

    NASA Astrophysics Data System (ADS)

    Alwanney, Deaa; Chami, Ziad Al; Angelica De Pascali, Sandra; Cavoski, Ivana; Fanizzi, Francesco Paolo

    2014-05-01

    Nowadays, improving plant performance by providing growth promoters is a main concern of the organic agriculture. As a consequence of increased food demands, more efficient and alternatives of the current plant nutrition strategies are becoming urgent. Recently, a novel concept "bio-effectors" raised on to describe a group of products that are able to improve plant performance and do not belong to fertilizers or pesticides. Agro-Food processing residues are promising materials as bio-effector. Three plant-derived materials: brewers' spent grain (BSG), fennel processing residues (FPR) and lemon processing residues (LPR) were chosen as bio-effector candidates. Plant-derived materials were characterized in term of total macro and micronutrients content. Green extraction methodology and solvent choice (aqueous; ethanol; and aqueous: ethanol mixture 1:1) was based on the extraction yield as main factor. Optimum extracts, to be used on the tomato test plant, were determined using phytotoxicity test (seed germination test) as main constraint. Thereafter, selected extracts were characterized and secondary metabolites profiling were detected by NMR technique. Selected extracts were applied on tomato in a growth chamber at different doses in comparison to humic-like substances as positive control (Ctrl+) and to a Hoagland solution as negative control (Ctrl-). At the end of the experiment, agronomical parameters were determined and NMR-metabolomic profiling were conducted on tomato seedlings. Results are summarized as follow: (i) raw showed an interesting content, either at nutritional or biological level; (ii) aqueous extraction resulted higher yield than other used solvent; (iii) at high extraction ratio (1:25 for BSG; 1:100 for FPR; and 1:200 for LPR) aqueous extracts were not phytotoxic on the tomato test plant; (iv) all aqueous extract are differently rich in nutrients, aminoacids, sugars and low molecular weight molecules; (v) all extract exhibited a growth promotion at low application doses; (vi) regarding plant metabolomics study, all treatments showed a different metabolites in respect to Ctrl- treatment. BSG, LPR and Ctrl+ treatments had similar metabolic profile. Finally, Metabolomic study provided an efficient tool and a key reporter about bio-effectors impact on plants. The visible effect and measured agronomical parameters was emphasized and demonstrated by metabolic profiling which offer insights into the affected plant metabolic pathways. As conclusion, our results supported the prediction that plant derived materials may interfere again in plant production regardless their nutritional content. Keywords: Bio-effectors; Metabolomics; Nuclear Magnetic Resonance (NMR); Barley; Fennel; Lemon; Tomato.

  15. Soft substrates suppress droplet splashing

    E-print Network

    Howland, Christopher J; Style, Robert W; Castrejón-Pita, A A

    2015-01-01

    Droplets splash when they impact dry, flat substrates above a critical velocity that depends on parameters such as droplet size, viscosity and air pressure. We show that substrate stiffness also impacts the splashing threshold by imaging ethanol drops impacting silicone gels of different stiffnesses. Splashing is significantly suppressed: droplets on the softest substrates need over 70% more kinetic energy to splash than they do on rigid substrates. We show that splash suppression is likely to be due to energy losses caused by deformations of soft substrates during the first few microseconds of impact. We find that solids with Youngs modulus $\\lesssim O(10^5)$Pa suppress splashing, in agreement with simple scaling arguments. Thus materials like soft gels and elastomers can be used as simple coatings for effective splash prevention.

  16. Vibration suppression using smart structures

    NASA Technical Reports Server (NTRS)

    Garcia, Ephrahim; Inman, Daniel J.; Dosch, Jeffrey

    1991-01-01

    The control of structures for vibration suppression is discussed in the context of using smart materials and structures. Here the use of smart structures refers to using embedded piezoelectric devices as both control actuators and sensors. Using embedded sensors and actuators allows great improvements in performance over traditional structures (both passive and active) for vibration suppression. The application of smart structures to three experimental flexible structures is presented. The first is a flexible beam, the second is a flexible beam undergoing slewing motion, the third is a ribbed antenna. A simple model of a piezoelectric actuator/sensor is presented. The equations of motion for each structure is presented. The control issues considered as those associated with multi-input, multi-output control, PID control and LQR control implementation. A modern control analysis illustrates the usefulness of smart structures for vibration suppression.

  17. A translocated effector required for Bartonella dissemination from derma to blood safeguards migratory host cells from damage by co-translocated effectors.

    PubMed

    Okujava, Rusudan; Guye, Patrick; Lu, Yun-Yueh; Mistl, Claudia; Polus, Florine; Vayssier-Taussat, Muriel; Halin, Cornelia; Rolink, Antonius G; Dehio, Christoph

    2014-06-01

    Numerous bacterial pathogens secrete multiple effectors to modulate host cellular functions. These effectors may interfere with each other to efficiently control the infection process. Bartonellae are Gram-negative, facultative intracellular bacteria using a VirB type IV secretion system to translocate a cocktail of Bartonella effector proteins (Beps) into host cells. Based on in vitro infection models we demonstrate here that BepE protects infected migratory cells from injurious effects triggered by BepC and is required for in vivo dissemination of bacteria from the dermal site of inoculation to blood. Human endothelial cells (HUVECs) infected with a ?bepE mutant of B. henselae (Bhe) displayed a cell fragmentation phenotype resulting from Bep-dependent disturbance of rear edge detachment during migration. A ?bepCE mutant did not show cell fragmentation, indicating that BepC is critical for triggering this deleterious phenotype. Complementation of ?bepE with BepEBhe or its homologues from other Bartonella species abolished cell fragmentation. This cyto-protective activity is confined to the C-terminal Bartonella intracellular delivery (BID) domain of BepEBhe (BID2.EBhe). Ectopic expression of BID2.EBhe impeded the disruption of actin stress fibers by Rho Inhibitor 1, indicating that BepE restores normal cell migration via the RhoA signaling pathway, a major regulator of rear edge retraction. An intradermal (i.d.) model for B. tribocorum (Btr) infection in the rat reservoir host mimicking the natural route of infection by blood sucking arthropods allowed demonstrating a vital role for BepE in bacterial dissemination from derma to blood. While the Btr mutant ?bepDE was abacteremic following i.d. inoculation, complementation with BepEBtr, BepEBhe or BIDs.EBhe restored bacteremia. Given that we observed a similar protective effect of BepEBhe on infected bone marrow-derived dendritic cells migrating through a monolayer of lymphatic endothelial cells we propose that infected dermal dendritic cells may be involved in disseminating Bartonella towards the blood stream in a BepE-dependent manner. PMID:24945914

  18. A Translocated Effector Required for Bartonella Dissemination from Derma to Blood Safeguards Migratory Host Cells from Damage by Co-translocated Effectors

    PubMed Central

    Okujava, Rusudan; Guye, Patrick; Lu, Yun-Yueh; Mistl, Claudia; Polus, Florine; Vayssier-Taussat, Muriel; Halin, Cornelia; Rolink, Antonius G.; Dehio, Christoph

    2014-01-01

    Numerous bacterial pathogens secrete multiple effectors to modulate host cellular functions. These effectors may interfere with each other to efficiently control the infection process. Bartonellae are Gram-negative, facultative intracellular bacteria using a VirB type IV secretion system to translocate a cocktail of Bartonella effector proteins (Beps) into host cells. Based on in vitro infection models we demonstrate here that BepE protects infected migratory cells from injurious effects triggered by BepC and is required for in vivo dissemination of bacteria from the dermal site of inoculation to blood. Human endothelial cells (HUVECs) infected with a ?bepE mutant of B. henselae (Bhe) displayed a cell fragmentation phenotype resulting from Bep-dependent disturbance of rear edge detachment during migration. A ?bepCE mutant did not show cell fragmentation, indicating that BepC is critical for triggering this deleterious phenotype. Complementation of ?bepE with BepEBhe or its homologues from other Bartonella species abolished cell fragmentation. This cyto-protective activity is confined to the C-terminal Bartonella intracellular delivery (BID) domain of BepEBhe (BID2.EBhe). Ectopic expression of BID2.EBhe impeded the disruption of actin stress fibers by Rho Inhibitor 1, indicating that BepE restores normal cell migration via the RhoA signaling pathway, a major regulator of rear edge retraction. An intradermal (i.d.) model for B. tribocorum (Btr) infection in the rat reservoir host mimicking the natural route of infection by blood sucking arthropods allowed demonstrating a vital role for BepE in bacterial dissemination from derma to blood. While the Btr mutant ?bepDE was abacteremic following i.d. inoculation, complementation with BepEBtr, BepEBhe or BIDs.EBhe restored bacteremia. Given that we observed a similar protective effect of BepEBhe on infected bone marrow-derived dendritic cells migrating through a monolayer of lymphatic endothelial cells we propose that infected dermal dendritic cells may be involved in disseminating Bartonella towards the blood stream in a BepE-dependent manner. PMID:24945914

  19. Nonredundant roles of IL-10 and TGF-? in suppression of immune responses to hepatic AAV-factor IX gene transfer.

    PubMed

    Hoffman, Brad E; Martino, Ashley T; Sack, Brandon K; Cao, Ou; Liao, Gongxian; Terhorst, Cox; Herzog, Roland W

    2011-07-01

    Hepatic gene transfer using adeno-associated viral (AAV) vectors has been shown to efficiently induce immunological tolerance to a variety of proteins. Regulatory T-cells (Treg) induced by this route suppress humoral and cellular immune responses against the transgene product. In this study, we examined the roles of immune suppressive cytokines interleukin-10 (IL-10) and transforming growth factor-? (TGF-?) in the development of tolerance to human coagulation factor IX (hF.IX). Interestingly, IL-10 deficient C57BL/6 mice receiving gene transfer remained tolerant to hF.IX and generated Treg that suppressed anti-hF.IX formation. Effects of TGF-? blockade were also minor in this strain. In contrast, in C3H/HeJ mice, a strain known to have stronger T-cell responses against hF.IX, IL-10 was specifically required for the suppression of CD8(+) T-cell infiltration of the liver. Furthermore, TGF-? was critical for tipping the balance toward an regulatory immune response. TGF-? was required for CD4(+)CD25(+)FoxP3(+) Treg induction, which was necessary for suppression of effector CD4(+) and CD8(+) T-cell responses as well as antibody formation. These results demonstrate the crucial, nonredundant roles of IL-10 and TGF-? in prevention of immune responses against AAV-F.IX-transduced hepatocytes. PMID:21386826

  20. Conditioned suppression, punishment, and aversion

    NASA Technical Reports Server (NTRS)

    Orme-Johnson, D. W.; Yarczower, M.

    1974-01-01

    The aversive action of visual stimuli was studied in two groups of pigeons which received response-contingent or noncontingent electric shocks in cages with translucent response keys. Presentation of grain for 3 sec, contingent on key pecking, was the visual stimulus associated with conditioned punishment or suppression. The responses of the pigeons in three different experiments are compared.

  1. Plasma suppression of beamstrahlung: Revision

    SciTech Connect

    Whittum, D.H.; Sessler, A.M.; Stewart, J.J.; Yu, S.S.

    1988-06-01

    We investigate the use of a plasma at the interaction point of two colliding beams to suppress beamstrahlung and related phenomena. We derive conditions for good current cancellation via plasma return currents and report on numerical simulations conducted to confirm our analytic results. 17 refs., 5 figs., 5 tabs.

  2. -uncertainty Anonymization by Partial Suppression

    E-print Network

    Zhu, Kenny Q.

    Results (a) Original Dataset TID Transaction 1 bread, milk, condom 2 bread, milk 3 milk, condom 4 flour, fruits 5 flour, condom 6 bread, fruits 7 fruits, condom (b) Global Suppression TID Transaction 1 bread, milk, condom 2 bread, milk 3 milk, condom 4 flour, fruits 5 flour, condom 6 bread, fruits 7 fruits

  3. DENDRITIC POLYMERS AS FIRE SUPPRESSANTS

    EPA Science Inventory

    This report describes an evaluation of the applicability of one of the latest advances in polymer technology (dendritic polymers) to suppressing fires, one of the greatest survivability threats to military personnel and vehicles. Certain types of alkali and transition metal compl...

  4. Fermionic suppression of dipolar relaxation.

    PubMed

    Burdick, Nathaniel Q; Baumann, Kristian; Tang, Yijun; Lu, Mingwu; Lev, Benjamin L

    2015-01-16

    We observe the suppression of inelastic dipolar scattering in ultracold Fermi gases of the highly magnetic atom dysprosium: the more energy that is released, the less frequently these exothermic reactions take place, and only quantum spin statistics can explain this counterintuitive effect. Inelastic dipolar scattering in nonzero magnetic fields leads to heating or to loss of the trapped population, both detrimental to experiments intended to study quantum many-body physics with strongly dipolar gases. Fermi statistics, however, is predicted to lead to a kinematic suppression of these harmful reactions. Indeed, we observe a 120-fold suppression of dipolar relaxation in fermionic versus bosonic Dy, as expected from theory describing universal inelastic dipolar scattering, though never before experimentally confirmed. Similarly, low inelastic cross sections are observed in spin mixtures, also with striking correspondence to predictions. The suppression of relaxation opens the possibility of employing fermionic dipolar species in studies of quantum many-body physics involving, e.g., synthetic gauge fields and pairing. PMID:25635544

  5. Structure and Function of a Fungal Adhesin that Binds Heparin and Mimics Thrombospondin-1 by Blocking T Cell Activation and Effector Function

    PubMed Central

    Brandhorst, T. Tristan; Roy, René; Wüthrich, Marcel; Nanjappa, Som; Filutowicz, Hanna; Galles, Kevin; Tonelli, Marco; McCaslin, Darrell R.; Satyshur, Kenneth; Klein, Bruce

    2013-01-01

    Blastomyces adhesin-1 (BAD-1) is a 120-kD surface protein on B. dermatitidis yeast. We show here that BAD-1 contains 41 tandem repeats and that deleting even half of them impairs fungal pathogenicity. According to NMR, the repeats form tightly folded 17-amino acid loops constrained by a disulfide bond linking conserved cysteines. Each loop contains a highly conserved WxxWxxW motif found in thrombospondin-1 (TSP-1) type 1 heparin-binding repeats. BAD-1 binds heparin specifically and saturably, and is competitively inhibited by soluble heparin, but not related glycosaminoglycans. According to SPR analysis, the affinity of BAD-1 for heparin is 33 nM±14 nM. Putative heparin-binding motifs are found both at the N-terminus and within each tandem repeat loop. Like TSP-1, BAD-1 blocks activation of T cells in a manner requiring the heparan sulfate-modified surface molecule CD47, and impairs effector functions. The tandem repeats of BAD-1 thus confer pathogenicity, harbor motifs that bind heparin, and suppress T-cell activation via a CD47-dependent mechanism, mimicking mammalian TSP-1. PMID:23853587

  6. Three New Pierce's Disease Pathogenicity Effectors Identified Using Xylella fastidiosa Biocontrol Strain EB92-1

    PubMed Central

    Zhang, Shujian; Chakrabarty, Pranjib K.; Fleites, Laura A.; Rayside, Patricia A.; Hopkins, Donald L.; Gabriel, Dean W.

    2015-01-01

    Xylella fastidiosa (X. fastidiosa) infects a wide range of plant hosts and causes economically serious diseases, including Pierce's Disease (PD) of grapevines. X. fastidiosa biocontrol strain EB92-1 was isolated from elderberry and is infectious and persistent in grapevines but causes only very slight symptoms under ideal conditions. The draft genome of EB92-1 revealed that it appeared to be missing genes encoding 10 potential PD pathogenicity effectors found in Temecula1. Subsequent PCR and sequencing analyses confirmed that EB92-1 was missing the following predicted effectors found in Temecula1: two type II secreted enzymes, including a lipase (LipA; PD1703) and a serine protease (PD0956); two identical genes encoding proteins similar to Zonula occludens toxins (Zot; PD0915 and PD0928), and at least one relatively short, hemagglutinin-like protein (PD0986). Leaves of tobacco and citrus inoculated with cell-free, crude protein extracts of E. coli BL21(DE3) overexpressing PD1703 exhibited a hypersensitive response (HR) in less than 24 hours. When cloned into shuttle vector pBBR1MCS-5, PD1703 conferred strong secreted lipase activity to Xanthomonas citri, E. coli and X. fastidiosa EB92-1 in plate assays. EB92-1/PD1703 transformants also showed significantly increased disease symptoms on grapevines, characteristic of PD. Genes predicted to encode PD0928 (Zot) and a PD0986 (hemagglutinin) were also cloned into pBBR1MCS-5 and moved into EB92-1; both transformants also showed significantly increased symptoms on V. vinifera vines, characteristic of PD. Together, these results reveal that PD effectors include at least a lipase, two Zot-like toxins and a possibly redundant hemagglutinin, none of which are necessary for parasitic survival of X. fastidiosa populations in grapevines or elderberry. PMID:26218423

  7. Three New Pierce's Disease Pathogenicity Effectors Identified Using Xylella fastidiosa Biocontrol Strain EB92-1.

    PubMed

    Zhang, Shujian; Chakrabarty, Pranjib K; Fleites, Laura A; Rayside, Patricia A; Hopkins, Donald L; Gabriel, Dean W

    2015-01-01

    Xylella fastidiosa (X. fastidiosa) infects a wide range of plant hosts and causes economically serious diseases, including Pierce's Disease (PD) of grapevines. X. fastidiosa biocontrol strain EB92-1 was isolated from elderberry and is infectious and persistent in grapevines but causes only very slight symptoms under ideal conditions. The draft genome of EB92-1 revealed that it appeared to be missing genes encoding 10 potential PD pathogenicity effectors found in Temecula1. Subsequent PCR and sequencing analyses confirmed that EB92-1 was missing the following predicted effectors found in Temecula1: two type II secreted enzymes, including a lipase (LipA; PD1703) and a serine protease (PD0956); two identical genes encoding proteins similar to Zonula occludens toxins (Zot; PD0915 and PD0928), and at least one relatively short, hemagglutinin-like protein (PD0986). Leaves of tobacco and citrus inoculated with cell-free, crude protein extracts of E. coli BL21(DE3) overexpressing PD1703 exhibited a hypersensitive response (HR) in less than 24 hours. When cloned into shuttle vector pBBR1MCS-5, PD1703 conferred strong secreted lipase activity to Xanthomonas citri, E. coli and X. fastidiosa EB92-1 in plate assays. EB92-1/PD1703 transformants also showed significantly increased disease symptoms on grapevines, characteristic of PD. Genes predicted to encode PD0928 (Zot) and a PD0986 (hemagglutinin) were also cloned into pBBR1MCS-5 and moved into EB92-1; both transformants also showed significantly increased symptoms on V. vinifera vines, characteristic of PD. Together, these results reveal that PD effectors include at least a lipase, two Zot-like toxins and a possibly redundant hemagglutinin, none of which are necessary for parasitic survival of X. fastidiosa populations in grapevines or elderberry. PMID:26218423

  8. Influence of glutathione chemical effectors in the response of maize to arsenic exposure.

    PubMed

    Requejo, Raquel; Tena, Manuel

    2012-05-01

    To support the key role of glutathione (GSH) in the mechanisms of tolerance and accumulation of arsenic in plants, this work examines the impact of several effectors of GSH synthesis or action in the response of maize (Zea mays L.) to arsenic. Maize was exposed in hydroponics to iso-toxic rates of 150 ?M arsenate or 75 ?M arsenite for 9 days and GSH effectors, flurazole (an herbicide safener), l-buthionine-sulfoximine (BSO, a known inhibitor of GSH biosynthesis), and dimercaptosuccinate (DMS) and dimercaptopropanesulfonate (DMPS) (two thiols able to displace GSH from arsenite-GSH complexes) were assayed. The main responses of plants to arsenic exposure consisted of a biomass reduction (fresh weight basis) of about 50%, an increase of non-protein thiol (NPTs) levels (especially in the GSH precursor ?-glutamylcysteine and the phytochelatins PC? and PC?) in roots, with little effect in shoots, and an accumulation of between 600 and 1000 ppm of As (dry weight basis) in roots with very little translocation to shoots. Growth inhibition caused by arsenic was partially or completely reversed in plants co-treated with flurazole and arsenate or arsenite, respectively, highly exacerbated in plants co-treated with BSO, and not modified in plants co-treated with DMS or DMPS. These responses correlated well with an increase of both NPTs levels in roots and glutathione transferase activity in roots and shoots due to flurazole treatment, the decrease of NPTs levels in roots caused by BSO and the lack of effect on NPT levels caused by both DMS and DMPS. Regarding to arsenic accumulation in roots, it was not modified by flurazole, highly reduced by BSO, and increased between 2.5- and 4.0-fold by DMS and DMPS. Therefore, tolerance and accumulation of arsenic by maize could be manipulated pharmacologically by chemical effectors of GSH. PMID:22418430

  9. Global impact of Salmonella type III secretion effector SteA on host cells

    SciTech Connect

    Cardenal-Muñoz, Elena Gutiérrez, Gabriel Ramos-Morales, Francisco

    2014-07-11

    Highlights: • We analyzed HeLa cells transcriptome in response to Salmonella SteA. • Significant differential expression was detected for 58 human genes. • They are involved in ECM organization and regulation of some signaling pathways. • Cell death, cell adhesion and cell migration were decreased in SteA-expressing cells. • These results contribute to understand the role of SteA during infections. - Abstract: Salmonella enterica is a Gram-negative bacterium that causes gastroenteritis, bacteremia and typhoid fever in several animal species including humans. Its virulence is greatly dependent on two type III secretion systems, encoded in pathogenicity islands 1 and 2. These systems translocate proteins called effectors into eukaryotic host cell. Effectors interfere with host signal transduction pathways to allow the internalization of pathogens and their survival and proliferation inside vacuoles. SteA is one of the few Salmonella effectors that are substrates of both type III secretion systems. Here, we used gene arrays and bioinformatics analysis to study the genetic response of human epithelial cells to SteA. We found that constitutive synthesis of SteA in HeLa cells leads to induction of genes related to extracellular matrix organization and regulation of cell proliferation and serine/threonine kinase signaling pathways. SteA also causes repression of genes related to immune processes and regulation of purine nucleotide synthesis and pathway-restricted SMAD protein phosphorylation. In addition, a cell biology approach revealed that epithelial cells expressing steA show altered cell morphology, and decreased cytotoxicity, cell–cell adhesion and migration.

  10. Multiple Myeloma Impairs Bone Marrow Localization of Effector Natural Killer Cells by Altering the Chemokine Microenvironment.

    PubMed

    Ponzetta, Andrea; Benigni, Giorgia; Antonangeli, Fabrizio; Sciumè, Giuseppe; Sanseviero, Emilio; Zingoni, Alessandra; Ricciardi, Maria Rosaria; Petrucci, Maria Teresa; Santoni, Angela; Bernardini, Giovanni

    2015-11-15

    Natural killer (NK) cells are key innate immune effectors against multiple myeloma, their activity declining in multiple myeloma patients with disease progression. To identify the mechanisms underlying NK cell functional impairment, we characterized the distribution of functionally distinct NK cell subsets in the bone marrow of multiple myeloma-bearing mice. Herein we report that the number of KLRG1(-) NK cells endowed with potent effector function rapidly and selectively decreases in bone marrow during multiple myeloma growth, this correlating with decreased bone marrow NK cell degranulation in vivo. Altered NK cell subset distribution was dependent on skewed chemokine/chemokine receptor axes in the multiple myeloma microenvironment, with rapid downmodulation of the chemokine receptor CXCR3 on NK cells, increased CXCL9 and CXCL10, and decreased CXCL12 expression in bone marrow. Similar alterations in chemokine receptor/chemokine axes were observed in patients with multiple myeloma. Adoptive transfer experiments demonstrated that KLRG1(-) NK cell migration to the bone marrow was more efficient in healthy than multiple myeloma-bearing mice. Furthermore, bone marrow localization of transferred CXCR3-deficient NK cells with respect to wild type was enhanced in healthy and multiple myeloma-bearing mice, suggesting that CXCR3 restrains bone marrow NK cell trafficking. Our results indicate that multiple myeloma-promoted CXCR3 ligand upregulation together with CXCL12 downmodulation act as exit signals driving effector NK cells outside the bone marrow, thus weakening the antitumor immune response at the primary site of tumor growth. Cancer Res; 75(22); 4766-77. ©2015 AACR. PMID:26438594

  11. Legionella pneumophila Effector LpdA Is a Palmitoylated Phospholipase D Virulence Factor.

    PubMed

    Schroeder, Gunnar N; Aurass, Philipp; Oates, Clare V; Tate, Edward W; Hartland, Elizabeth L; Flieger, Antje; Frankel, Gad

    2015-10-01

    Legionella pneumophila is a bacterial pathogen that thrives in alveolar macrophages, causing a severe pneumonia. The virulence of L. pneumophila depends on its Dot/Icm type IV secretion system (T4SS), which delivers more than 300 effector proteins into the host, where they rewire cellular signaling to establish a replication-permissive niche, the Legionella-containing vacuole (LCV). Biogenesis of the LCV requires substantial redirection of vesicle trafficking and remodeling of intracellular membranes. In order to achieve this, several T4SS effectors target regulators of membrane trafficking, while others resemble lipases. Here, we characterized LpdA, a phospholipase D effector, which was previously proposed to modulate the lipid composition of the LCV. We found that ectopically expressed LpdA was targeted to the plasma membrane and Rab4- and Rab14-containing vesicles. Subcellular targeting of LpdA required a C-terminal motif, which is posttranslationally modified by S-palmitoylation. Substrate specificity assays showed that LpdA hydrolyzed phosphatidylinositol, -inositol-3- and -4-phosphate, and phosphatidylglycerol to phosphatidic acid (PA) in vitro. In HeLa cells, LpdA generated PA at vesicles and the plasma membrane. Imaging of different phosphatidylinositol phosphate (PIP) and organelle markers revealed that while LpdA did not impact on membrane association of various PIP probes, it triggered fragmentation of the Golgi apparatus. Importantly, although LpdA is translocated inefficiently into cultured cells, an L. pneumophila ?lpdA mutant displayed reduced replication in murine lungs, suggesting that it is a virulence factor contributing to L. pneumophila infection in vivo. PMID:26216420

  12. Fingolimod modulates peripheral effector and regulatory T cells in MS patients.

    PubMed

    Serpero, Laura D; Filaci, Gilberto; Parodi, Alessia; Battaglia, Florinda; Kalli, Francesca; Brogi, Davide; Mancardi, Giovanni Luigi; Uccelli, Antonio; Fenoglio, Daniela

    2013-12-01

    Multiple sclerosis (MS) is a complex neurological disease where, in genetically predisposed individuals, the unbalanced interplay between pathogenic and regulatory T cells will result in the progression of the autoimmune assault to neural antigens. Fingolimod (FTY720), an oral sphingosine 1-phosphate modulator recently approved for the treatment of MS, inhibits the egress of T cells from lymph nodes acting specifically on naïve and memory T cells and sparing effector T cells. Here we characterized IL-17 and IFN? producing effector CD4 and CD8 positive T cells as well as CD4 positive CD25(high)CD127(low) regulatory T cells in MS patients before and 1 month after treatment was started. We observed that fingolimod did not significantly affect the percentage of CCR6 and CD161 positive T cells in both CD4 and CD8 compartments. In contrast, it significantly reduced the levels of both CD4+ CCR6+ CD161+ and CD8+ CCR6+ CD161+ producing IFN? alone or in combination with IL-17. The percentage of IL-17 secreting cells in both subsets was affected by the treatment to a lesser extent. Finally, we observed that CD4+ CD25(high)CD127(low) regulatory T cells were decreased in MS patients compared to healthy controls and fingolimod significantly increased their frequencies. All together these findings demonstrate that fingolimod functionally modulates the ability of potentially pathogenic effector cells to produce relevant pro-inflammatory cytokines and increases the number of circulating regulatory T cells possibly contributing in restoring a balance between these populations. PMID:23649711

  13. Novel cyclic di-GMP effectors of the YajQ protein family control bacterial virulence.

    PubMed

    An, Shi-qi; Caly, Delphine L; McCarthy, Yvonne; Murdoch, Sarah L; Ward, Joseph; Febrer, Melanie; Dow, J Maxwell; Ryan, Robert P

    2014-10-01

    Bis-(3',5') cyclic di-guanylate (cyclic di-GMP) is a key bacterial second messenger that is implicated in the regulation of many critical processes that include motility, biofilm formation and virulence. Cyclic di-GMP influences diverse functions through interaction with a range of effectors. Our knowledge of these effectors and their different regulatory actions is far from complete, however. Here we have used an affinity pull-down assay using cyclic di-GMP-coupled magnetic beads to identify cyclic di-GMP binding proteins in the plant pathogen Xanthomonas campestris pv. campestris (Xcc). This analysis identified XC_3703, a protein of the YajQ family, as a potential cyclic di-GMP receptor. Isothermal titration calorimetry showed that the purified XC_3703 protein bound cyclic di-GMP with a high affinity (K(d)?2 µM). Mutation of XC_3703 led to reduced virulence of Xcc to plants and alteration in biofilm formation. Yeast two-hybrid and far-western analyses showed that XC_3703 was able to interact with XC_2801, a transcription factor of the LysR family. Mutation of XC_2801 and XC_3703 had partially overlapping effects on the transcriptome of Xcc, and both affected virulence. Electromobility shift assays showed that XC_3703 positively affected the binding of XC_2801 to the promoters of target virulence genes, an effect that was reversed by cyclic di-GMP. Genetic and functional analysis of YajQ family members from the human pathogens Pseudomonas aeruginosa and Stenotrophomonas maltophilia showed that they also specifically bound cyclic di-GMP and contributed to virulence in model systems. The findings thus identify a new class of cyclic di-GMP effector that regulates bacterial virulence. PMID:25329577

  14. GCN2 sustains mTORC1 suppression upon amino acid deprivation by inducing Sestrin2.

    PubMed

    Ye, Jiangbin; Palm, Wilhelm; Peng, Min; King, Bryan; Lindsten, Tullia; Li, Ming O; Koumenis, Constantinos; Thompson, Craig B

    2015-11-15

    Mammalian cells possess two amino acid-sensing kinases: general control nonderepressible 2 (GCN2) and mechanistic target of rapamycin complex 1 (mTORC1). Their combined effects orchestrate cellular adaptation to amino acid levels, but how their activities are coordinated remains poorly understood. Here, we demonstrate an important link between GCN2 and mTORC1 signaling. Upon deprivation of various amino acids, activated GCN2 up-regulates ATF4 to induce expression of the stress response protein Sestrin2, which is required to sustain repression of mTORC1 by blocking its lysosomal localization. Moreover, Sestrin2 induction is necessary for cell survival during glutamine deprivation, indicating that Sestrin2 is a critical effector of GCN2 signaling that regulates amino acid homeostasis through mTORC1 suppression. PMID:26543160

  15. The role of effectors and host immunity in plant–necrotrophic fungal interactions

    PubMed Central

    Wang, Xuli; Jiang, Nan; Liu, Jinling; Liu, Wende; Wang, Guo-Liang

    2014-01-01

    Fungal diseases pose constant threats to the global economy and food safety. As the largest group of plant fungal pathogens, necrotrophic fungi cause heavy crop losses worldwide. The molecular mechanisms of the interaction between necrotrophic fungi and plants are complex and involve sophisticated recognition and signaling networks. Here, we review recent findings on the roles of phytotoxin and proteinaceous effectors, pathogen-associated molecular patterns (PAMPs), and small RNAs from necrotrophic fungi. We also consider the functions of damage-associated molecular patterns (DAMPs), the receptor-like protein kinase BIK1, and epigenetic regulation in plant immunity to necrotrophic fungi. PMID:25513773

  16. 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.

  17. Apoptosis and expression of cytotoxic T lymphocyte effector molecules in renal allografts.

    PubMed

    Olive, C; Cheung, C; Falk, M C

    1999-03-01

    Cytotoxic T lymphocyte (CTL) mediated apoptosis is thought to play a major role in the rejection of renal allografts following transplantation, however, the CTL effector mechanism that is primarily responsible for immunological rejection is unknown. The two major effector pathways of CTL killing which lead to apoptosis involve the Fas/Fas ligand (Fas L) lytic pathway, and the perforin/granzyme degranulation pathway. The expression of CTL effector molecules which influence these pathways include Fas, Fas L and TiA-1 (cytotoxic granule protein). This study has investigated apoptosis by in situ terminal deoxytransferase-catalysed DNA nick end labelling (TUNEL), and the expression of CTL effector molecules by immunohistochemistry, in renal allograft biopsies obtained from patients following kidney transplantation. Renal biopsies were classified into three histological groups; acute cellular rejection, chronic rejection, or no rejection. The extent of T-cell infiltration of renal tissues was assessed by immunohistochemical staining with an anti-CD3 monoclonal antibody. Numerous TUNEL positive cells were detected in all transplant biopsies examined; these consisted mainly of renal tubular cells and infiltrating cells, with some TUNEL positive cells also detected in the glomeruli. In the case of normal kidney tissue, renal cells also stained positive for TUNEL but there was no lymphocytic infiltration. There was significantly more T-cell infiltration observed in acute rejection biopsies compared to the no rejection biopsies. In the case of Fas L expression, there was little expression in all three biopsy groups, apart from one case of chronic rejection. Conversely, although there were no significant differences in TiA-1 expression between the three biopsy groups, TiA-1 expression was more prominent in acute rejection biopsies. Furthermore, Fas expression was significantly decreased in acute rejection biopsies when compared to those of chronic and no rejection in which Fas was predominantly localized in the renal tubular cells. These results indicate that the mechanism of CTL killing leading to the rejection of renal allografts may be different in acute and chronic rejection. Moreover, our data indicate the potential for cytotoxic granule-based CTL killing in acute renal allograft rejection but not in chronic rejection. PMID:10375075

  18. Physical and neural entrainment to rhythm: human sensorimotor coordination across tasks and effector systems

    PubMed Central

    Ross, Jessica Marie; Balasubramaniam, Ramesh

    2014-01-01

    The human sensorimotor system can be readily entrained to environmental rhythms, through multiple sensory modalities. In this review, we provide an overview of theories of timekeeping that make this neuroentrainment possible. First, we present recent evidence that contests the assumptions made in classic timekeeper models. The role of state estimation, sensory feedback and movement parameters on the organization of sensorimotor timing are discussed in the context of recent experiments that examined simultaneous timing and force control. This discussion is extended to the study of coordinated multi-effector movements and how they may be entrained. PMID:25136306

  19. Chemically Synthesized Molecules with the Targeting and Effector Functions of Antibodies

    PubMed Central

    2015-01-01

    This article reports the design, synthesis, and evaluation of a novel class of molecules of intermediate size (approximately 7000 Da), which possess both the targeting and effector functions of antibodies. These compounds—called synthetic antibody mimics targeting prostate cancer (SyAM-Ps)—bind simultaneously to prostate-specific membrane antigen and Fc gamma receptor I, thus eliciting highly selective cancer cell phagocytosis. SyAMs have the potential to combine the advantages of both small-molecule and biologic therapies, and may address many drawbacks associated with available treatments for cancer and other diseases. PMID:25514603

  20. A Discussion of Aerodynamic Control Effectors (ACEs) for Unmanned Air Vehicles (UAVs)

    NASA Technical Reports Server (NTRS)

    Wood, Richard M.

    2002-01-01

    A Reynolds number based, unmanned air vehicle classification structure has been developed which identifies four classes of unmanned air vehicle concepts. The four unmanned air vehicle (UAV) classes are; Micro UAV, Meso UAV, Macro UAV, and Mega UAV. In a similar fashion a labeling scheme for aerodynamic control effectors (ACE) was developed and eleven types of ACE concepts were identified. These eleven types of ACEs were laid out in a five (5) layer scheme. The final section of the paper correlated the various ACE concepts to the four UAV classes and ACE recommendations are offered for future design activities.

  1. Multi-Angle Effector Function Analysis of Human Monoclonal IgG Glycovariants

    PubMed Central

    Dashivets, Tetyana; Thomann, Marco; Rueger, Petra; Knaupp, Alexander; Buchner, Johannes; Schlothauer, Tilman

    2015-01-01

    Therapeutic performance of recombinant antibodies relies on two independent mechanisms: antigen recognition and Fc-mediated antibody effector functions. Interaction of Fc-fragment with different FcR triggers antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity and determines longevity of the antibody in serum. In context of therapeutic antibodies Fc?Rs play the most important role. It has been demonstrated that the Fc-attached sugar moiety is essential for IgG effector functionality, dictates its affinity to individual Fc?Rs and determines binding to different receptor classes: activating or inhibitory. In this study, we systematically analyze effector functions of monoclonal IgG1 and its eight enzymatically engineered glycosylation variants. The analysis of interaction of glycovariants with FcRs was performed for single, as well as for antigen-bound antibodies and IgGs in a form of immune complex. In addition to functional properties we addressed impact of glycosylation on the structural properties of the tested glycovariants. We demonstrate a clear impact of glycosylation pattern on antibody stability and interaction with different Fc?Rs. Consistent with previous reports, deglycosylated antibodies failed to bind all Fc?-receptors, with the exception of high affinity Fc?RI. The Fc?RII and Fc?RIIIa binding activity of IgG1 was observed to depend on the galactosylation level, and hypergalactosylated antibodies demonstrated increased receptor interaction. Sialylation did not decrease the Fc?R binding of the tested IgGs; in contrast, sialylation of antibodies improved binding to Fc?RIIa and IIb. We demonstrate that glycosylation influences to some extent IgG1 interaction with FcRn. However, independent of glycosylation pattern the interaction of IgG1 with a soluble monomeric target surprisingly resulted in an impaired receptor binding. Here, we demonstrate, that immune complexes (IC), induced by multimeric ligand, compensated for the decreased affinity of target bound antibody towards FcRs, showing the importance of the IC-formation for the FcR- mediated effector functions. PMID:26657484

  2. Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response

    PubMed Central

    2014-01-01

    Background Transcriptional activator-like (TAL) effectors, formerly known as the AvrBs3/PthA protein family, are DNA-binding effectors broadly found in Xanthomonas spp. that transactivate host genes upon injection via the bacterial type three-secretion system. Biologically relevant targets of TAL effectors, i.e. host genes whose induction is vital to establish a compatible interaction, have been reported for xanthomonads that colonize rice and pepper; however, citrus genes modulated by the TAL effectors PthA“s” and PthC“s” of the citrus canker bacteria Xanthomonas citri (Xc) and Xanthomonas aurantifolii pathotype C (XaC), respectively, are poorly characterized. Of particular interest, XaC causes canker disease in its host lemon (Citrus aurantifolia), but triggers a defense response in sweet orange. Results Based on, 1) the TAL effector-DNA binding code, 2) gene expression data of Xc and XaC-infiltrated sweet orange leaves, and 3) citrus hypocotyls transformed with PthA2, PthA4 or PthC1, we have identified a collection of Citrus sinensis genes potentially targeted by Xc and XaC TAL effectors. Our results suggest that similar with other strains of Xanthomonas TAL effectors, PthA2 and PthA4, and PthC1 to some extent, functionally converge. In particular, towards induction of genes involved in the auxin and gibberellin synthesis and response, cell division, and defense response. We also present evidence indicating that the TAL effectors act as transcriptional repressors and that the best scoring predicted DNA targets of PthA“s” and PthC“s” in citrus promoters predominantly overlap with or localize near to TATA boxes of core promoters, supporting the idea that TAL effectors interact with the host basal transcriptional machinery to recruit the RNA pol II and start transcription. Conclusions The identification of PthA“s” and PthC“s” targets, such as the LOB (LATERAL ORGAN BOUNDARY) and CCNBS genes that we report here, is key for the understanding of the canker symptoms development during host susceptibility, or the defenses of sweet orange against the canker bacteria. We have narrowed down candidate targets to a few, which pointed out the host metabolic pathways explored by the pathogens. PMID:24564253

  3. Phylogenomics of Xanthomonas field strains infecting pepper and tomato reveals diversity in effector repertoires and identifies determinants of host specificity

    PubMed Central

    Schwartz, Allison R.; Potnis, Neha; Timilsina, Sujan; Wilson, Mark; Patané, José; Martins, Joaquim; Minsavage, Gerald V.; Dahlbeck, Douglas; Akhunova, Alina; Almeida, Nalvo; Vallad, Gary E.; Barak, Jeri D.; White, Frank F.; Miller, Sally A.; Ritchie, David; Goss, Erica; Bart, Rebecca S.; Setubal, João C.; Jones, Jeffrey B.; Staskawicz, Brian J.

    2015-01-01

    Bacterial spot disease of pepper and tomato is caused by four distinct Xanthomonas species and is a severely limiting factor on fruit yield in these crops. The genetic diversity and the type III effector repertoires of a large sampling of field strains for this disease have yet to be explored on a genomic scale, limiting our understanding of pathogen evolution in an agricultural setting. Genomes of 67 Xanthomonas euvesicatoria (Xe), Xanthomonas perforans (Xp), and Xanthomonas gardneri (Xg) strains isolated from diseased pepper and tomato fields in the southeastern and midwestern United States were sequenced in order to determine the genetic diversity in field strains. Type III effector repertoires were computationally predicted for each strain, and multiple methods of constructing phylogenies were employed to understand better the genetic relationship of strains in the collection. A division in the Xp population was detected based on core genome phylogeny, supporting a model whereby the host-range expansion of Xp field strains on pepper is due, in part, to a loss of the effector AvrBsT. Xp-host compatibility was further studied with the observation that a double deletion of AvrBsT and XopQ allows a host range expansion for Nicotiana benthamiana. Extensive sampling of field strains and an improved understanding of effector content will aid in efforts to design disease resistance strategies targeted against highly conserved core effectors. PMID:26089818

  4. Structure of the effector-binding domain of the arabinose repressor AraR from Bacillus subtilis

    SciTech Connect

    Procházková, Kate?ina; ?ermáková, Kate?ina; Pachl, Petr; Sieglová, Irena; Fábry, Milan; Otwinowski, Zbyszek; ?ezá?ová, Pavlína

    2012-02-01

    The crystal structure of the effector-binding domain of the transcriptional repressor AraR from B. subtilis in complex with the effector molecule (l-arabinose) was determined at 2.2 Å resolution. A detailed analysis of the crystal identified a dimer organization that is distinctive from that of other members of the GalR/LacI family. In Bacillus subtilis, the arabinose repressor AraR negatively controls the expression of genes in the metabolic pathway of arabinose-containing polysaccharides. The protein is composed of two domains of different phylogenetic origin and function: an N-terminal DNA-binding domain belonging to the GntR family and a C-terminal effector-binding domain that shows similarity to members of the GalR/LacI family. The crystal structure of the C-terminal effector-binding domain of AraR in complex with the effector l-arabinose has been determined at 2.2 Å resolution. The l-arabinose binding affinity was characterized by isothermal titration calorimetry and differential scanning fluorimetry; the K{sub d} value was 8.4 ± 0.4 µM. The effect of l-arabinose on the protein oligomeric state was investigated in solution and detailed analysis of the crystal identified a dimer organization which is distinctive from that of other members of the GalR/LacI family.

  5. Translocation of Magnaporthe oryzae Effectors into Rice Cells and Their Subsequent Cell-to-Cell Movement[W][OA

    PubMed Central

    Khang, Chang Hyun; Berruyer, Romain; Giraldo, Martha C.; Kankanala, Prasanna; Park, Sook-Young; Czymmek, Kirk; Kang, Seogchan; Valent, Barbara

    2010-01-01

    Knowledge remains limited about how fungal pathogens that colonize living plant cells translocate effector proteins inside host cells to regulate cellular processes and neutralize defense responses. To cause the globally important rice blast disease, specialized invasive hyphae (IH) invade successive living rice (Oryza sativa) cells while enclosed in host-derived extrainvasive hyphal membrane. Using live-cell imaging, we identified a highly localized structure, the biotrophic interfacial complex (BIC), which accumulates fluorescently labeled effectors secreted by IH. In each newly entered rice cell, effectors were first secreted into BICs at the tips of the initially filamentous hyphae in the cell. These tip BICs were left behind beside the first-differentiated bulbous IH cells as the fungus continued to colonize the host cell. Fluorescence recovery after photobleaching experiments showed that the effector protein PWL2 (for prevents pathogenicity toward weeping lovegrass [Eragrostis curvula]) continued to accumulate in BICs after IH were growing elsewhere. PWL2 and BAS1 (for biotrophy-associated secreted protein 1), BIC-localized secreted proteins, were translocated into the rice cytoplasm. By contrast, BAS4, which uniformly outlines the IH, was not translocated into the host cytoplasm. Fluorescent PWL2 and BAS1 proteins that reached the rice cytoplasm moved into uninvaded neighbors, presumably preparing host cells before invasion. We report robust assays for elucidating the molecular mechanisms that underpin effector secretion into BICs, translocation to the rice cytoplasm, and cell-to-cell movement in rice. PMID:20435900

  6. Bio-effectors from waste materials as growth promoters for tomato plants, an agronomic and metabolomic study

    NASA Astrophysics Data System (ADS)

    Abou Chehade, Lara; Chami, Ziad Al; De Pascali, Sandra; Cavoski, Ivana; Fanizzi, Francesco Paolo

    2015-04-01

    In organic farming, where nutrient management is constrained and sustainability is claimed, bio-effectors pave their way. Considering selected bio-effectors, this study integrates metabolomics to agronomy in depicting induced relevant phenomena. Extracts of three agro-industrial wastes (Lemon processing residues, Fennel processing residues and Brewer's spent grain) are being investigated as sources of bio-effectors for the third trial consequently. Corresponding individual and mixture aqueous extracts are assessed for their synergistic and/or single agronomic and qualitative performances on soil-grown tomato, compared to both a control and humic acid treatments. A metabolomic profiling of tomato fruits via the Proton Nuclear Magnetic Resonance (NMR) spectroscopy, as holistic indicator of fruit quality and extract-induced responses, complements crop productivity and organoleptic/nutritional qualitative analyses. Results are expected to show mainly an enhancement of the fruit qualitative traits, and to confirm partly the previous results of better crop productivity and metabolism enhancement. Waste-derived bio-effectors could be, accordingly, demonstrated as potential candidates of plant-enhancing substances. Keywords: bio-effectors, organic farming, agro-industrial wastes, nuclear magnetic resonance (NMR), tomato.

  7. Noise suppressing capillary separation system

    DOEpatents

    Yeung, E.S.; Xue, Y.

    1996-07-30

    A noise-suppressing capillary separation system for detecting the real-time presence or concentration of an analyte in a sample is provided. The system contains a capillary separation means through which the analyte is moved, a coherent light source that generates a beam which is split into a reference beam and a sample beam that irradiate the capillary, and a detector for detecting the reference beam and the sample beam light that transmits through the capillary. The laser beam is of a wavelength effective to be absorbed by a chromophore in the capillary. The system includes a noise suppressing system to improve performance and accuracy without signal averaging or multiple scans. 13 figs.

  8. S-maltoheptaose targets syndecan-bound effectors to reduce smoking-related neutrophilic inflammation

    PubMed Central

    Lam, David CL; Chan, Stanley CH; Mak, Judith CW; Freeman, Craig; Ip, Mary SM; Shum, Daisy KY

    2015-01-01

    Cigarette smoke induces injury and neutrophilic inflammation in the airways of smokers. The stability and activity of inflammatory effectors, IL8 and neutrophil elastase (NE), can be prolonged by binding to airway heparan sulfate (HS)/syndecan-1, posing risk for developing chronic obstructive pulmonary disease(COPD). We hypothesize that antagonizing HS/syndecan-1 binding of the inflammatory effectors could reduce smoking-related neutrophil-mediated airway inflammation. Analysis of bronchoalveolar lavage fluid(BALF) of COPD patients found both total and unopposed NE levels to be significantly higher among smokers with COPD than non-COPD subjects. Similar NE burden was observed in smoke-exposed rats compared to sham air controls. We chose sulfated-maltoheptaose(SM), a heparin-mimetic, to antagonize HS/sydecan-1 binding of the inflammatory mediators in airway fluids and lung tissues of the smoke-exposed rat model. Airway treatment with SM resulted in displacement of CINC-1 and NE from complexation with bronchio-epithelial HS/syndecan-1, dissipating the chemokine gradient for neutrophil flux across to the bronchial lumen. Following SM displacement of NE from shed HS/syndecan-1 in bronchial fluids, NE became accessible to inhibition by ?1-antitrypsin endogenous in test samples. The antagonistic actions of SM against syndecan-1 binding of NE and CINC-1 in smoke-exposed airways suggest new therapeutic opportunities for modulating airway inflammation in smokers with SM delivery. PMID:26256047

  9. Modularly assembled designer TAL effector nucleases for targeted gene knockout and gene replacement in eukaryotes

    SciTech Connect

    Li, T; Huang, S; Zhao, XF; Wright, DA; Carpenter, S; Spalding, MH; Weeks, DP; Yang, B

    2011-08-08

    Recent studies indicate that the DNA recognition domain of transcription activator-like (TAL) effectors can be combined with the nuclease domain of FokI restriction enzyme to produce TAL effector nucleases (TALENs) that, in pairs, bind adjacent DNA target sites and produce double-strand breaks between the target sequences, stimulating non-homologous end-joining and homologous recombination. Here, we exploit the four prevalent TAL repeats and their DNA recognition cipher to develop a 'modular assembly' method for rapid production of designer TALENs (dTALENs) that recognize unique DNA sequence up to 23 bases in any gene. We have used this approach to engineer 10 dTALENs to target specific loci in native yeast chromosomal genes. All dTALENs produced high rates of site-specific gene disruptions and created strains with expected mutant phenotypes. Moreover, dTALENs stimulated high rates (up to 34%) of gene replacement by homologous recombination. Finally, dTALENs caused no detectable cytotoxicity and minimal levels of undesired genetic mutations in the treated yeast strains. These studies expand the realm of verified TALEN activity from cultured human cells to an intact eukaryotic organism and suggest that low-cost, highly dependable dTALENs can assume a significant role for gene modifications of value in human and animal health, agriculture and industry.

  10. Assessment of the Geographic Origins of Pinewood Nematode Isolates via Single Nucleotide Polymorphism in Effector Genes

    PubMed Central

    Figueiredo, Joana; Simões, Maria José; Gomes, Paula; Barroso, Cristina; Pinho, Diogo; Conceição, Luci; Fonseca, Luís; Abrantes, Isabel; Pinheiro, Miguel; Egas, Conceição

    2013-01-01

    The pinewood nematode, Bursaphelenchus xylophilus, is native to North America but it only causes damaging pine wilt disease in those regions of the world where it has been introduced. The accurate detection of the species and its dispersal routes are thus essential to define effective control measures. The main goals of this study were to analyse the genetic diversity among B. xylophilus isolates from different geographic locations and identify single nucleotide polymorphism (SNPs) markers for geographic origin, through a comparative transcriptomic approach. The transcriptomes of seven B. xylophilus isolates, from Continental Portugal (4), China (1), Japan (1) and USA (1), were sequenced in the next generation platform Roche 454. Analysis of effector gene transcripts revealed inter-isolate nucleotide diversity that was validated by Sanger sequencing in the genomic DNA of the seven isolates and eight additional isolates from different geographic locations: Madeira Island (2), China (1), USA (1), Japan (2) and South Korea (2). The analysis identified 136 polymorphic positions in 10 effector transcripts. Pairwise comparison of the 136 SNPs through Neighbor-Joining and the Maximum Likelihood methods and 5-mer frequency analysis with the alignment-independent bilinear multivariate modelling approach correlated the SNPs with the isolates geographic origin. Furthermore, the SNP analysis indicated a closer proximity of the Portuguese isolates to the Korean and Chinese isolates than to the Japanese or American isolates. Each geographic cluster carried exclusive alleles that can be used as SNP markers for B. xylophilus isolate identification. PMID:24391785

  11. Angiogenin mutants as novel effector molecules for the generation of fusion proteins with increased cytotoxic potential.

    PubMed

    Cremer, Christian; Vierbuchen, Tim; Hein, Lea; Fischer, Rainer; Barth, Stefan; Nachreiner, Thomas

    2015-04-01

    Human cytolytic fusion proteins (hCFPs) are therapeutically efficacious recombinant polypeptides comprising a target cell-specific binding component and a human effector domain that induces apoptosis. Compared with former generations of immunotoxins, which contain immunogenic cytotoxic domains derived from bacteria or plants, hCFPs contain solely human proteins that do not induce an immune response, thus avoiding the development of neutralizing antibodies. Here, we investigated the suitability of human angiogenin (Ang) mutants as effector domains. We engineered 3 different Ang variants that outperformed the wild-type enzyme by replacing amino acid residues with key roles in the protein's catalytic activity and its interaction with the ribonuclease inhibitor RNH1. The cytotoxic potential of these mutants was compared with wild-type Ang by fusing each to the CD64-specific single-chain variable fragment H22. All hCFPs were successfully expressed in HEK293T cells and purified from the cell culture supernatant by immobilized metal ion affinity chromatography. The Ang mutant-based hCFPs showed normal binding activity towards human interferon-?-stimulated CD64 HL-60 cells and activated human macrophages isolated from peripheral blood mononuclear cells, but increased cytotoxicity based on reduced affinity towards RNH1 and higher ribonucleolytic activity. PMID:25710248

  12. The Legionella pneumophila Effector Protein, LegC7, Alters Yeast Endosomal Trafficking

    PubMed Central

    O’Brien, Kevin M.; Lindsay, Elizabeth L.; Starai, Vincent J.

    2015-01-01

    The intracellular pathogen, Legionella pneumophila, relies on numerous secreted effector proteins to manipulate host endomembrane trafficking events during pathogenesis, thereby preventing fusion of the bacteria-laden phagosome with host endolysosomal compartments, and thus escaping degradation. Upon expression in the surrogate eukaryotic model Saccharomyces cerevisiae, we find that the L. pneumophila LegC7/YlfA effector protein disrupts the delivery of both biosynthetic and endocytic cargo to the yeast vacuole. We demonstrate that the effects of LegC7 are specific to the endosome:vacuole delivery pathways; LegC7 expression does not disrupt other known vacuole-directed pathways. Deletions of the ESCRT-0 complex member, VPS27, provide resistance to the LegC7 toxicity, providing a possible target for LegC7 function in vivo. Furthermore, a single amino acid substitution in LegC7 abrogates both its toxicity and ability to alter endosomal traffic in vivo, thereby identifying a critical functional domain. LegC7 likely inhibits endosomal trafficking during L. pneumophila pathogenesis to prevent entry of the phagosome into the endosomal maturation pathway and eventual fusion with the lysosome. PMID:25643265

  13. Multipart Chaperone-Effector Recognition in the Type III Secretion System of Chlamydia trachomatis.

    PubMed

    Shen, Li; Macnaughtan, Megan A; Frohlich, Kyla M; Cong, Yanguang; Goodwin, Octavia Y; Chou, Chau-Wen; LeCour, Louis; Krup, Kristen; Luo, Miao; Worthylake, David K

    2015-11-20

    Secretion of effector proteins into the eukaryotic host cell is required for Chlamydia trachomatis virulence. In the infection process, Scc1 and Scc4, two chaperones of the type III secretion (T3S) system, facilitate secretion of the important effector and plug protein, CopN, but little is known about the details of this event. Here we use biochemistry, mass spectrometry, nuclear magnetic resonance spectroscopy, and genetic analyses to characterize this trimolecular event. We find that Scc4 complexes with Scc1 and CopN in situ at the late developmental cycle of C. trachomatis. We show that Scc4 and Scc1 undergo dynamic interactions as part of the unique bacterial developmental cycle. Using alanine substitutions, we identify several amino acid residues in Scc4 that are critical for the Scc4-Scc1 interaction, which is required for forming the Scc4·Scc1·CopN ternary complex. These results, combined with our previous findings that Scc4 plays a role in transcription (Rao, X., Deighan, P., Hua, Z., Hu, X., Wang, J., Luo, M., Wang, J., Liang, Y., Zhong, G., Hochschild, A., and Shen, L. (2009) Genes Dev. 23, 1818-1829), reveal that the T3S process is linked to bacterial transcriptional events, all of which are mediated by Scc4 and its interacting proteins. A model describing how the T3S process may affect gene expression is proposed. PMID:26438824

  14. 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.

  15. A tetrapod-like repertoire of innate immune receptors and effectors for coelacanths

    USGS Publications Warehouse

    Boudinot, Pierre; Zou, Jun; Ota, Tatsuya; Buonocore, Francesco; Scapigliati, Giuseppe; Canapa, Adriana; Cannon, John; Litman, Gary; Hansen, John D.

    2014-01-01

    The recent availability of both robust transcriptome and genome resources for coelacanth (Latimeria chalumnae) has led to unique discoveries for coelacanth immunity such as the lack of IgM, a central component of adaptive immunity. This study was designed to more precisely address the origins and evolution of gene families involved in the initial recognition and response to microbial pathogens, which effect innate immunity. Several multigene families involved in innate immunity are addressed, including: Toll-like receptors (TLRs), retinoic acid inducible gene 1 (RIG1)-like receptors (RLRs), the nucleotide-binding domain and leucine-rich repeat containing proteins (NLRs), diverse immunoglobulin domain-containing proteins (DICP) and modular domain immune-type receptors (MDIRs). Our analyses also include the tripartite motif-containing proteins (TRIM), which are involved in pathogen recognition as well as the positive regulation of antiviral immunity. Finally, this study addressed some of the downstream effectors of the antimicrobial response including IL-1 family members, type I and II interferons (IFN) and IFN-stimulated effectors (ISGs). Collectively, the genes and gene families in coelacanth that effect innate immune functions share characteristics both in content, structure and arrangement with those found in tetrapods but not in teleosts. The findings support the sister group relationship of coelacanth fish with tetrapods.

  16. Characterization and DNA-Binding Specificities of Ralstonia TAL-Like Effectors

    PubMed Central

    Mahfouz, Magdy M.

    2013-01-01

    Transcription activator-like effectors (TALEs) from Xanthomonas sp. have been used as customizable DNA-binding modules for genome-engineering applications. Ralstonia solanacearum TALE-like proteins (RTLs) exhibit similar structural features to TALEs, including a central DNA-binding domain composed of 35 amino acid-long repeats. Here, we characterize the RTLs and show that they localize in the plant cell nucleus, mediate DNA binding, and might function as transcriptional activators. RTLs have a unique DNA-binding architecture and are enriched in repeat variable di-residues (RVDs), which determine repeat DNA-binding specificities. We determined the DNA-binding specificities for the RVD sequences ND, HN, NP, and NT. The RVD ND mediates highly specific interactions with C nucleotide, HN interacts specifically with A and G nucleotides, and NP binds to C, A, and G nucleotides. Moreover, we developed a highly efficient repeat assembly approach for engineering RTL effectors. Taken together, our data demonstrate that RTLs are unique DNA-targeting modules that are excellent alternatives to be tailored to bind to user-selected DNA sequences for targeted genomic and epigenomic modifications. These findings will facilitate research concerning RTL molecular biology and RTL roles in the pathogenicity of Ralstonia spp. PMID:23300258

  17. 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 Fc?RIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and Fc?RIIa. 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

  18. 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 Fc?RIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and Fc?RIIa. 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

  19. Multi-modal representation of effector modality in frontal cortex during rule switching

    PubMed Central

    Hodgson, Timothy L.; Parris, Benjamin A.; Benattayallah, Abdelmalek; Summers, Ian R.

    2015-01-01

    We report a functional magnetic resonance imaging (fMRI) study which investigated whether brain areas involved in updating task rules within the frontal lobe of the cerebral cortex show activity related to the modality of motor response used in the task. Participants performed a rule switching task using different effector modalities. In some blocks participants responded with left/right button presses, whilst in other blocks left/right saccades were required. The color of a Cue event instructed a left or right response based upon a rule, followed by a Feedback which indicated whether the rule was to stay the same or “Flip” on the next trial. The findings revealed variation in the locus of activity within the ventrolateral frontal cortex dependent upon effector modality. Other frontal areas showed no significant difference in activity between response epochs but changed their pattern of connectivity with posterior cortical areas dependent upon response. Multivariate analysis revealed that the pattern of activity evoked by Flip rule Feedbacks within an apparently supra modal frontal region (dorsolateral frontal cortex) discriminated between response epochs. The results are consistent with the existence of multi-modal representations of stimulus-response (SR) rules within the frontal cerebral cortex. PMID:26441588

  20. Altered effector functions of NK cells in chronic hepatitis C are associated with IFNL3 polymorphism.

    PubMed

    Rogalska-Taranta, Magdalena; Markova, Antoaneta A; Taranta, Andrzej; Lunemann, Sebastian; Schlaphoff, Verena; Flisiak, Robert; Manns, Michael P; Cornberg, Markus; Kraft, Anke R M; Wedemeyer, Heiner

    2015-08-01

    Interferon ?-mediated effector functions of NK cells may contribute to the control of HCV replication and the pathogenesis of liver disease. The single-nucleotide polymorphism rs12979860 near IFNL3 (previously known as IL28B) is important in response to IFN-? treatment and in spontaneous resolution of acute hepatitis C. The role of the IFNL3 polymorphism in NK cell function is unclear. Thus, we investigated the role of IFNL3 polymorphism in type I IFN-dependent regulation of NK cell functions in patients with cHC and healthy control subjects. We demonstrated a marked polarization of NK cells toward cytotoxicity in response to IFN-? stimulation in patients with hepatitis C. That TRAIL up-regulation was present, particularly in patients with the IFNL3-TT allele, was supported by a shift in the pSTAT-1:pSTAT-4 ratios toward pSTAT-1. In patients bearing the IFNL3-TT allele, NK cell effector function correlated with liver disease activity. In contrast, higher cytokine production of NK cells was observed in healthy individuals with the IFNL3-CC genotype, which may support spontaneous HCV clearance in acute infection. Overall, these findings show that the role of NK cells may differ in chronic infection vs. early antiviral defense and that the IFNL3 genotype differentially influences NK cell function. PMID:26034208