Academic literature on the topic 'TET protease'
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Journal articles on the topic "TET protease"
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Ishikawa, Chika, Tatsuya Tsuda, Hiroe Konishi, Noboru Nakagawa, and Kiyofumi Yamanishi. "Tetracyclines Modulate Protease-Activated Receptor 2-Mediated Proinflammatory Reactions in Epidermal Keratinocytes." Antimicrobial Agents and Chemotherapy 53, no. 5 (March 2, 2009): 1760–65. http://dx.doi.org/10.1128/aac.01540-08.
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ABSTRACT In addition to their antibiotic effects, tetracyclines have anti-inflammatory action that is often beneficial in the control of inflammatory skin disorders. In this study, we examined the effects of tetracycline (TET) and two of its derivatives, doxycycline (DOX) and minocycline (MIN), on the production of interleukin-8 (IL-8) elicited by the activation of protease-activated receptor 2 (PAR2) in normal human epidermal keratinocytes (NHEK). In NHEK, the production of IL-8 stimulated by an agonist peptide of PAR2, SLIGKIV-NH2, at 100 μM was significantly reduced by TET, DOX, or MIN at 5 and 10 μM, concentrations that are noncytotoxic. The tumor necrosis factor alpha (TNF-α)-induced production of IL-8 was synergistically augmented by SLIGKIV-NH2, and that synergistic increase in the production of IL-8 was suppressed by 100 nM PAR2-specific small interfering RNA. It was also suppressed by TET, DOX, or MIN but not by the 14-membered-ring macrolide antibiotics erythromycin, roxithromycin, and clarithromycin, which also have anti-inflammatory activities, at 10 μM. These results suggest that tetracyclines attenuate the PAR2-IL-8 axis in keratinocytes and thereby effectively modulate proinflammatory responses in the skin.
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Wang, Chun-hui, Jia-min Yang, Yu-bo Guo, Jing Shen, and Xiao-hua Pei. "Anticancer Activity of Tetrandrine by Inducing Apoptosis in Human Breast Cancer Cell Line MDA-MB-231 In Vivo." Evidence-Based Complementary and Alternative Medicine 2020 (June 30, 2020): 1–11. http://dx.doi.org/10.1155/2020/6823520.
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Tetrandrine (TET) is an alkaloid extracted from a traditional Chinese medicinal plant. It exerts remarkable anticancer activity and induces apoptotic cell death in various human cancer cells. The present study aimed to investigate the effects of TET on the inhibition of tumor growth and the induction of apoptosis in MDA-MB-231 breast cancer in xenograft mice. Tumor weight and volume were measured. The histopathological changes in the tumor tissue were observed. Immunohistochemistry analysis of Bcl-2-associated X protein (Bax) and B-cell lymphoma/leukemia-2 (Bcl-2) was carried out. The expression of apoptosis-associated genes and proteins, such as cysteine aspartic acid-specific protease-3 (Caspase-3), Survivin, Bax, Bcl-2, BH3-interacting domain death agonist (Bid), and poly ADP-ribose polymerase (PARP), was measured by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, respectively. TET inhibited tumor growth and induced apoptosis in TNBC cell line MDA-MB-231. The mechanism underlying this effect might be mediated by TET-upregulated Caspase-3, Bax, and Bid and downregulated by Bcl-2, Survivin, and PARP. Taken together, this study supported the fact that TET is a promising therapeutic agent for the treatment of TNBC, thereby providing experimental evidence for its use in the treatment of breast cancer.
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Borissenko, Ljudmila, and Michael Groll. "Crystal Structure of TET Protease Reveals Complementary Protein Degradation Pathways in Prokaryotes." Journal of Molecular Biology 346, no. 5 (March 2005): 1207–19. http://dx.doi.org/10.1016/j.jmb.2004.12.056.
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Chow, W. A., S. Guo, and F. Valdes-Albini. "HIV protease inhibitor (PI) therapy for liposarcoma." Journal of Clinical Oncology 24, no. 18_suppl (June 20, 2006): 9564. http://dx.doi.org/10.1200/jco.2006.24.18_suppl.9564.
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9564 Background: Liposarcomas are the second most common soft-tissue sarcoma. Highly-active anti-retroviral therapy (HAART) with HIV PIs results in “HIV-1 protease inhibitor associated lipodystrophy syndrome,” characterized by peripheral fat wasting, central fat accumulation, insulin resistance, and hyperlipidemia. Based upon this syndrome, we hypothesized that HIV PIs might represent a novel liposarcoma therapy. Methods: SW872, LiSa-2, and FU-DDLS-1 liposarcoma, and control 293 embryonic kidney and HT1080 fibrosarcoma cell lines were treated with HIV PIs and subjected to cellular and molecular assays. Results: Clonogenic assays with SW872 cells using HIV PIs (saquinavir, ritonavir, indinavir, nelfinavir, and amprenavir) were performed. Nelfinavir demonstrated the most potent clonogenic inhibition without affecting 293 and HT1080 clonogenicity, and was studied further. Nelfinavir inhibited SW872 and LiSa-2 proliferation dose-dependently, and HT1080 proliferation at the highest concentration, without affecting FU-DDLS-1 nor 293 proliferation. Nelfinavir induced a G1 cell cycle arrest in SW872 and HT1080, but not in 293 cells. It also induced dose-dependent apoptosis in SW872, but not in 293 nor HT1080 cells. Western analyses for sterol regulatory element binding protein-1 (SREBP-1) expression, a key transcriptional regulator of fatty acid and cholesterol synthesis, were performed. Nelfinavir induced expression of SREBP-1 in nelfinavir-sensitive SW872 and LiSa-2 cells, and modestly in HT1080 cells, but not in insensitive FU-DDLS-1 nor 293 cells. Additionally, nelfinavir reduced protein expression of proliferating cell nuclear antigen (PCNA) in sensitive SW872 and LiSa-2 cells, and induced expression of the anti-proliferative protein, p21, as well as pro-apoptotic proteins, Bax and Fas, in a dose-dependent manner. Finally, forced expression of SREBP-1 with a Tet-On inducible SW872 cell line, in the absence of nelfinavir, induced expression of p21, Bax, Fas, reduced expression of PCNA, and inhibited cell proliferation. Conclusions: These studies demonstrate that nelfinavir inhibits cellular proliferation, and induces apoptosis in sensitive-liposarcoma cells through upregulation of SREBP-1. These studies validate nelfinavir as a potential, novel targeted therapy for liposarcoma. No significant financial relationships to disclose.
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Chen, S., D. O. Henry, and M. K. Wong. "The biologic therapy of prostate cancer using plasminogen activator inhibitor-1 (PAI-1)." Journal of Clinical Oncology 24, no. 18_suppl (June 20, 2006): 14596. http://dx.doi.org/10.1200/jco.2006.24.18_suppl.14596.
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14596 Background: Treating prostate cancer through the expression of intrinsic biologic modifiers is a relatively unexplored aspect of prostate cancer therapy. Plasminogen Activator Inhibitor-1 (PAI-1) is expressed at low levels in prostate cancer cells. PAI-1 is both an anti-angiogenesis agent, and also potently inhibits tumor proteases responsible for tumor invasion and metastases such as uPA and tPA. Thus we hypothesized that stimulation of tumor endogenous PAI-1 would result in a particularly powerful and profound prostate cancer regression. We present proof-of-concept from our experimental models that demonstrate significant tumor regression in experimental prostate tumors and supports this hypothesis. Methods and Results: Human prostate adenocarcinoma (PC3 cell line) xenograft tumors engineered to conditionally express either PAI-1 or Green Fluorescent Protein (GFP, control) were used to test our hypothesis. Stable cell lines were created that conditionally express either GFP or PAI-1 under the regulation of a doxycycline-responsive promoter (Tet-On). Thus gene expression is switched on in the presence of doxycycline. PC3 tumors were inoculated and allowed to reach at least 200 mm3 in size whereby the tumor-bearing mice were given doxycycline-doped drinking water. Genes were significantly turned on within 48 hours as monitored by the appearance of a GFP signal in control mice. The induction of PAI-1 results in significant inhibition of tumor growth as compared to GFP control. Importantly, in vitro induction of PAI-1 expression in PC-3 has no direct effects on cell growth as compared to any PC-3 control. Histological analysis of these tumors revealed a rich nexus of fine angiogenic vessels at the interface between control tumors and surrounding stroma. PAI-1 secreting tumors were significantly smaller and were pale, bland, and lacked peritumoral vessels. Protease activity measured by in-situ zymography directly on these tumors revealed that this was significantly reduced in PAI-1 expressing tumors as compared to GFP controls. Conclusion: PAI-1 expression results in tumor inhibition through direct anti-angiogenic effects and inhibition of tumor protease activity. No significant financial relationships to disclose.
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Borissenko, Ljudmila, and Michael Groll. "Corrigendum to “Crystal Structure of TET Protease Reveals Complementary Protein Degradation Pathways in Prokaryotes” [J. Mol. Biol. (2005) 346, 1207–1219]." Journal of Molecular Biology 351, no. 1 (August 2005): 247. http://dx.doi.org/10.1016/j.jmb.2005.03.032.
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Byarugaba, Denis K., Godfrey Wokorach, Stephen Alafi, Bernard Erima, Florence Najjuka, Edison A. Mworozi, Hannah Kibuuka, and Fred Wabwire-Mangen. "Whole Genome Sequencing Reveals High Genetic Diversity, Diverse Repertoire of Virulence-Associated Genes and Limited Antibiotic Resistance Genes among Commensal Escherichia coli from Food Animals in Uganda." Microorganisms 11, no. 8 (July 25, 2023): 1868. http://dx.doi.org/10.3390/microorganisms11081868.
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Commensal Escherichia coli with broad repertoire of virulence and antimicrobial resistance (AMR) genes pose serious public health risks as reservoirs of AMR and virulence. This study undertook whole genome characterization of commensal E. coli from food-producing animals in Uganda to investigate their genome variability (resistome and virulome). We established that the E. coli had high genomic diversity with 38 sequence types, 24 FimH types, and 33 O-antigen serotypes randomly distributed within three phylogroups (A, B1, and E). A greater proportion (≥93.65%) of the E. coli were resistant to amoxicillin/clavulanate and ampicillin antibiotics. The isolates were AmpC beta-lactamase producers dominated by blaEC-15 (71.88%) and tet(A) (20.31%) antimicrobial resistant genes besides a diverse armory of virulence-associated genes in the class of exotoxin, adhesins, iron uptake, and serine protease autotransporters which varied by host species. Cattle were found to be the major source of E. coli carrying Shiga toxin genes, whereas swine was the main source of E. coli carrying colicin-like Usp toxin gene. The study underscores the importance of livestock as the carrier of E. coli with antimicrobial resistance and a large repertoire of virulence traits with a potential of causing disease in animals and humans by acquiring more genetic traits.
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Maghsoudi, Nader, Narges Kh Tafreshi, Fariba Khodagholi, Zahra Zakeri, Mitra Esfandiarei, Hamid Hadi-Alijanvand, Marjan Sabbaghian, et al. "Targeting enteroviral 2A protease by a 16-mer synthetic peptide: Inhibition of 2Apro-induced apoptosis in a stable Tet-on HeLa cell line." Virology 399, no. 1 (March 2010): 39–45. http://dx.doi.org/10.1016/j.virol.2009.12.017.
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Aprikyan, Andrew A. G., Tomas Vaisar, Vahagn Makaryan, and Jay Heinecke. "Cellular Model of Severe Congenital Neutropenia Reveals the Molecular Mechanism of Mutant Elastase-Mediated Agranulocytosis." Blood 104, no. 11 (November 16, 2004): 1453. http://dx.doi.org/10.1182/blood.v104.11.1453.1453.
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Abstract Severe congenital neutropenia (SCN) or Kostmann’s syndrome defines an inheritable hematopoietic disorder of an impaired neutrophil production in the bone marrow due to a “maturation arrest” at promyelocytic stage of differentiation in the marrow. SCN patients have recurring severe infections and may evolve to develop leukemia. We reported accelerated apoptosis and cell cycle arrest of bone marrow-derived myeloid progenitor cells in SCN patients with acquired, autosomal dominant, as well as autosomal recessive inheritance. We also reported that approximately 80% of these patients have heterozygous mutations in the neutrophil elastase (NE) gene encoding a serine protease normally targeted to azurophil granules. Neither knock-in of mutant elastase nor a knock-out of normal neutrophil elastase caused severe neutropenia in mice. Therefore, we established a cellular model of SCN with tetracycline-regulated expression of mutant NE in human promyelocytic tet-off HL-60 cells. Induced expression of mutant elastase in these cells led to a reduced mitochondrial membrane potential and subsequent caspase-independent apoptosis and cell cycle arrest as determined by flow cytometry. Block of differentiation of DMSO-treated cells was also observed in tet-off HL-60 cells with induced expression of mutant NE, similar to that in SCN patients. This cellular model of SCN very closely recapitulates the human severe neutropenia phenotype. To elucidate the molecular mechanisms of mutant NE-mediated neutropenia, we employed various proteomics methods, including ICAT analysis, two-dimensional gel electrophoresis, immunoprecipitation, LC-MS/MS and identified a number of proteins with altered level of expression including apoptosis and cell cycle regulatory gene products, transcription factor and cytoskeleton proteins. Changes in the protein expression profiles revealed the abnormal molecular events underlying the impaired cell survival and cell cycle arrest and suggested additional pathways implicated in pathogenesis of SCN. Immunostaining of control cells with phalloidin revealed a weak F-actin polymerization in cell periphery, which helps to maintain cell shape flexibility, and intracellular long F-actin filaments supporting the normal cytoskeleton. In contrast, cells expressing mutant NE, exhibited an increased polymerization of F-actin in the periphery, with apparent increase in lamellipodia that may contribute to an increased phosphatydilserine exposure in apoptotic cells expressing mutant elastase. The hyperpolymerization of F-actin, which appears to stem from an elevated level of RhoA, a member of the RAS-superfamily of GTPases, makes the cells more rigid and less flexible thus contributing to impaired cell cycle progression in cells expressing mutant NE. Impaired cell survival in these cells is associated with a significant reduction in the level of phosphorylated PKB/Akt, which in turn appears to be a consequence of a decreased level of PI3kinase in response to expression of mutant NE. Interestingly, G-CSF treatment of mutant NE expressing cells resulted in restoration of the level of phospho-Akt to near normal level comparable to that in control cells with normal NE expression. These data may explain the anti-apoptotic effect of G-CSF in SCN. Thus, these data demonstrate that the cellular model of SCN based on tet-off HL-60/mutNE cells with inducible expression of mutant elastase is useful to unravel the cellular and molecular mechanisms of mutant NE-mediated severe neutropenia.
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Aprikyan, Andrew A., Tomas Vaisar, Vahagn Makaryan, and Jay Heinecke. "A Model of Severe Congenital Neutropenia Reveals Signaling Pathways Mediating Mutant Elastase-Triggered Agranulocytosis." Blood 106, no. 11 (November 16, 2005): 3070. http://dx.doi.org/10.1182/blood.v106.11.3070.3070.
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Abstract Severe congenital neutropenia (SCN; Kostmann’s syndrome or infantile genetic agranulocytosis) defines an inheritable hematopoietic disorder of impaired neutrophil production due to a “maturation arrest” at the promyelocytic stage of differentiation in the bone marrow. SCN patients have recurring severe infections and often develop acute myelogenous leukemia. We and others reported accelerated apoptosis and cell cycle arrest of bone marrow-derived myeloid progenitor cells in SCN patients with autosomal dominant and autosomal recessive inheritance. Heterozygous mutations in the neutrophil elastase (NE) gene encoding a serine protease, are present in a majority of SCN patients, but not in healthy members of the family, thus indicating a key role of mutant NE in pathogenesis of this disorder. To date, there are no animal or cellular models of SCN as both the knock-in of mutant NE as well as the knock-out of normal NE failed to result in neutropenia phenotype in mice. The molecular mechanisms of mutant NE-mediated severe neutropenia remain largely unknown. We hypothesized that mutations in NE expose the protease to a new range of substrates. To explore this proposal, we established a cellular model of SCN based on tetracycline-regulated expression of mutant NE in human promyelocytic tet-off HL-60 cells that very closely recapitulated the human phenotype. Mutant NE expression resulted in a characteristic block of myeloid differentiation - the cellular hallmark of SCN. Expression of the mutant product was associated with a significant reduction in phosphatidylinosytol-3-kinase and phosphorylated PKB/Akt levels and an imbalance of anti-apoptotic Bcl-2 and pro-apoptotic Bax. These alterations contributed to observed dissipation of mitochondrial membrane potential as determined by FACS analysis, aberrant release of cytochrome C, and accelerated apoptosis. Marked changes in actin cytoskeleton that made the cells more rigid appeared to stem from a reduced level of alpha-actinin and elevated level of Rho GTPase. Immunoprecipitation of cell lysates with elastase-specific monoclonal antibodies followed by mass spectrometric analysis revealed that NE interacted with histone H2B, one of the key components of the nucleosome core of the chromatin. Interestingly, the expression level of histone H2B was substantially reduced in cells expressing mutant NE, therefore supporting the notion of altered substrate specificity of mutant NE. Thus, these observations provide the first evidence that mutant NE affects specific signaling pathways that lead to alterations in cytoskeleton and chromatin reorganization, subsequent apoptosis, and a block of myeloid differentiation in SCN. This cellular model of SCN should provide an invaluable tool for screening potential therapeutic agents capable of preventing maturation arrest and leukemogenesis in subjects suffering from severe congenital neutropenia.
Dissertations / Theses on the topic "TET protease"
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Marty, Vincent. "Adaptation de l'Archaea halophile halobacterium salinarum aux stress environnementaux : mécanismes de survie et rôle de la protéolyse intracellulaire." Thesis, Grenoble, 2011. http://www.theses.fr/2011GRENV087/document.
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Les systèmes moléculaires décrits chez les Archaea mettent en évidence un caractère primitif et une simplicité, comparativement à leurs homologues eucaryotes. Par ailleurs, leur caractère extrêmophile a pour conséquence une hyper-robustesse qui rend leur manipulation in vitro et les études structurales beaucoup plus aisées. Ainsi les Archaea représentent de bons modèles pour comprendre les fonctions cellulaires complexes, particulièrement celles qui mettent en jeu de grandes machineries moléculaires, comme celles impliquées dans la protéolyse. Mon travail de thèse a consisté à comprendre les mécanismes de résistance et l'importance des systèmes de protéolyse dans l'adaptation des Archaea halophiles aux stress environnementaux. Les Archaea halophiles accumulent des concentrations multi-molaires de KCl/NaCl dans leur cytosol (3.4M KCl / 1.1M NaCl chez Halobacterium salinarum). Ainsi, les protéines de ces organismes sont elles-mêmes halophiles et ne sont solubles et repliées que dans des conditions de salinité extrêmes (de 2 à 5M).Nous avons étudié la réponse de l'Archaea halophile stricte H. salinarum à des stress, dont les stress à basse salinité, en se focalisant en particulier sur les modifications de la dynamique moléculaire du protéome in vivo (diffusion neutronique). Au cours de ce travail, il a été mis en évidence un phénomène de survie à la basse salinité associé à des modifications morphologiques.Un autre objectif de ma thèse a été de contribuer à la compréhension du rôle dans la protéolyse intracellulaire du complexe aminopeptidasique TET, dans les conditions de stress mises en place dans les études précédentesMolecular systems described for Archaea show primitive and simple characteristics, compared to their homologous eukaryotes. In addition, extremophilic characteristic results in an hyper-robust which makes in vitro manipulation and structural studies much easier. Thus, Archaea represent good models for understanding complex cellular functions, particularly those that involve large molecular machines, such as those involved in proteolysis. My thesis consisted in understanding the resistance mechanisms and the importance of proteolytic systems in the adaptation of halophilic Archaea to environmental stresses. Halophilic Archaea accumulate multi-molar concentrations of KCl / NaCl in their cytosol (3.4M KCl / NaCl 1.1M for Halobacterium Salinarum). This requires a very special biochemistry that allows operation in solvents where free water is scarce. Thus, the proteins of these organisms are themselves halophilic and are soluble and folded only in extreme salinity conditions (2 to 5 M). This particular biochemistry partly explain the extraordinary ability of halophilic Archaea to resist physical and chemical stress (temperature, radiation, dehydration). We study the response of the halophilic Archaea strict H. salinarum at low-salinity stress. Indeed, beyond the osmotic shock, the fall of the environment salt concentration causes a decrease in the intracellular KCl concentration, which should have a direct effect on the folding state of intracellular protein, as in case of heat stress. In the first part of this thesis, a study was conduct to determine viability limits and cytosolic modifications, associated with a salinity decrease. These studies involve intracellular salt dosages, viability studies (microscopic counts, color live / dead), induction of chaperone proteins linked to stress response and biophysical neutron experiments, to evaluate the effect of stress on proteins folding. In this work, a phenomenon of survival at low salinity linked to morphological changes was revealed. To describe this phenomenon, this second study involves confocal microscopy experiences, fluorescence microscopy, viability tests, counting on box, scanning electron microscopy, electron microscopy by negative staining, salt intracellular dosages and proteins separation experiments, to study the overall proteome composition during low salinity stress. In this study, a fall of the intracellular K $^+$ concentration and the proteome clarification during stress was revealed. Low salt concentrations causes halophiles proteins denaturation, the accumulation of misfolded proteins in the cytoplasm involves chaperones systems and intracellular proteolysis machinery. In this context, another objective of my thesis was to contribute to the understanding of the intracellular proteolysis role in the PAN-proteasome system and in the aminopeptidase TET complex, in stress conditions established in previous studies. This part of the thesis involves experiments of endopeptidase activity assay, aminopeptidase activity assay, quantification of mRNA genic expression by Northern blot, immunoprecipitation, proteins separation by sucrose gradient and proteasome chemical inhibition (drug). We show the role of the PAN-proteasome system in stress response and we deepen our understanding of the aminopeptidase TET role in vivo. This protease appears to have an independent role of the proteasome complex. The protease TET seems to participate at the amino acids treatment in cells to maintain the metabolic activities in nutritional deficiencies
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Tonnaire, Thierry. "Conception et synthese d'inhibiteurs contraints de la protease du vih-1 (doctorat : pharmacochimie)." Paris 11, 1999. http://www.theses.fr/1999PA114843.
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Chan, Ka-ho John, and 陳家豪. "Effects of Chinese green tea on cigarette smoke-induced oxidative stress, inflammation and proteases/anti-proteases in rat lung in vivo." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2010. http://hub.hku.hk/bib/B45199334.
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The Best PhD Thesis in the Faculties of Architecture, Arts, Business &Economics, Education, Law and Social Sciences (University of HongKong), Li Ka Shing Prize, 2008-2009published_or_final_version
Medicine
Doctoral
Doctor of Philosophy
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CARRON, CLEMENCE. "Etude fonctionnelle de la proteine tel et des onco-proteines derivees tel-pdgfr et tel-jak2." Paris 6, 1999. http://www.theses.fr/1999PA066554.
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Les regulateurs transcriptionnels de la famille ets participent a divers processus oncogeniques chez l'homme. Ils sont impliques sous forme de proteines de fusion issues de translocations chromosomiques specifiques. Tel appartient a la famille ets. Tel est remanie dans plusieurs leucemies humaines. Dans ces remaniements chromosomiques, tel est fusionne a differents partenaires (pdgfr, abl, jak2, tkc, aml1). Le plus souvent, c'est la partie 5 de tel qui est fusionnee a la portion 3 des genes partenaires. Cette region de tel code, notamment, un domaine conserve entre certaines proteines ets, le domaine b. Les proprietes biochimiques et fonctionnelles du domaine b n'etaient pas connues au moment ou nous avons debute nos travaux. Nous avons etudie le role du domaine b de tel dans la fonction de la proteine tel sauvage ainsi que dans les proteines de fusion tel-pdgfr et tel-jak2. Nous avons montre que le domaine b de tel est un domaine d'interaction proteique homotypique. Cette propriete est specifique du domaine b de tel car le domaine b d'autres proteines ets ne presente pas cette capacite d'auto-association. Nous avons montre que tel-pdgfr forme des homo-oligomeres in vivo. Cette auto-association permet l'activation constitutive de l'activite tyrosine kinase intrinseque de tel-pdgfr. Les proprietes mitogeniques de tel-pdgfr reposent sur la capacite du domaine b de tel a induire son auto-association. Par ailleurs, nous avons montre que, contrairement a la majorite des proteines ets qui sont des activateurs transcriptionnels, tel est un represseur. Cette propriete repose en partie sur la capacite de tel a s'auto-associer. Enfin, nous avons teste les proprietes oncogeniques de tel-jak2 in vivo en generant et en etudiant des souris transgeniques exprimant cette proteine dans le lignage lymphoide. Ces animaux developpent des leucemies t fatales. Ces resultats ont permis de definir l'implication du domaine b de tel dans le fonctionnement de tel sauvage et dans les proprietes transformantes des proteines derivees tel-pdgfr et tel-jak2. Les animaux transgeniques que nous avons generes seront utiles pour analyser in vivo l'importance des voies de signalisation de jak2 dans les processus oncogeniques. Ces animaux ouvrent aussi la perspective d'un developpement d'experimentations a but therapeutique.
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Bengtsson, Martin. "PURIFICATION AND CLEAVAGE OF FUSION PROTEIN CONTAINING THE PHOTOSYSTEM I SUBUNIT PSI-N USING AFFINITY CHROMATOGRAPHY AND TEV PROTEASE." Thesis, Halmstad University, School of Business and Engineering (SET), 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:hh:diva-1749.
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A method describing the expression and purification of PSI-N together with fusion protein, using affinity chromatography and TEV protease. Although the method proved successful, optimization is still needed due to partial degradation of PSI-N.
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DE, VUYST Guillaume. "LA PROTEINE MC1 D'ARCHAEABACTERIE : RECONNAISSANCE DE SEQUENCES PARTICULIERES." Phd thesis, Université d'Orléans, 2004. http://tel.archives-ouvertes.fr/tel-00009918.
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La protéine MC1 est une petite protéine structurale très abondante chez Methanosarcina thermophila (archaeabactérie). Nous avons utilisé la méthode SELEX (Systematic Evolution of Ligands by EXponential enrichment) pour rechercher ses séquences préférentielles de fixation. Après 10 cycles de sélection, une séquence consensus avec une affinité 50 fois plus forte que celle pour une séquence aléatoire a été déterminée. Cette séquence a été étudiée par simulation de dynamique moléculaire. Le mode de fixation de MC1 sur l?ADN a ensuite été déterminé par des empreintes moléculaires (DMS, OH·). Les résultats montrent une fixation par une face et dans le petit sillon de l?ADN. Une reconnaissance par lecture indirecte des séquences est probable. L?oxydation de certains acides aminés (Trp, Met) par les rayons gamma entraîne une modification des propriétés de la protéine : perte de la reconnaissance de structures et séquences particulières et diminution de sa capacité à courber l?ADN.
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Malnoë, Alizée. "A genetic suppressor approach to the biogenesis, quality control and function of photosynthetic complexes in Chlamydomonas reinhardtii." Phd thesis, Université Paris Sud - Paris XI, 2011. http://tel.archives-ouvertes.fr/tel-01057821.
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Central in oxygenic photosynthesis, the cytochrome b6f complex, couples electron transfer to proton translocation across the thylakoid membrane via its quinol:plastocyanin oxidoreductase activity, contributing to ATP formation. Cytochrome b6f complex differs from its respiratory homolog, the bc1 complex, by the presence of an additional heme, heme ci located within the quinone reduction site Qi and attached by a unique thioether bond. Mutants lacking heme ci show low accumulation of partially functional b6f complex and, hence, cannot grow phototrophically. This grounded a screen for suppressor mutations that would restore higher accumulation of b6f complexes whose function, even if compromised, would sustain phototrophic growth.The genetic suppressor approach undertook in Chlamydomonas reinhardtii during this PhD thesis led to the isolation and characterisation of the ftsh1-1 protease mutant (mutation R420C which should affect ATP hydrolysis). The mutant ftsh1-1 proved to be a versatile tool for the functional study of otherwise degraded proteins. The combination of genetic, biochemical, physiological and biophysical experiments demonstrated notably that: (i) a QiKO mutant, whose b6f complexes are devoid of both bh and ci hemes, can grow phototrophically despite a broken Q-cycle, (ii) the absence of covalently bound heme ci, in the Rccb2 mutant, triggers photosensivity enhanced in the presence of O2 supporting a role for heme ci in oxygen rich environment, (iii) FtsH is involved in the maintenance of the main photosynthetic complexes.
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Gummel, Jérémie. "STRUCTURES ET MECANISMES DE FORMATION DE COMPLEXES POLYELECTROLYTE-PROTEINE." Phd thesis, Université Paris Sud - Paris XI, 2006. http://tel.archives-ouvertes.fr/tel-00120315.
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Les mécanismes régissant la formation de complexes constitués de chaînes polymères chargées (polyélectrolytes) et de protéines, rencontrés dans l'agro-alimentaire, la pharmacologie ou la biologie, mènent à des structures mal connues jusqu'ici. Nous les avons étudiées par Diffusion de Neutrons aux Petits Angles (DNPA), associée au marquage par deutériation du polymère et à la variation de contraste dans des mélanges eau lourde - eau légère. La protéine est le lysozyme (chargé positivement à pH < 11) et le polyélectrolyte le polystyrène sulfonate (PSS, toujours chargé négativement) ; le rapport des charges apportées([-]/[+]) est un paramètre essentiel. Lorsqu'il est proche de 1, on obtient soit un gel de PSS réticulé par les protéines, qui contractent partiellement les chaînes de PSS, tout en les laissant en régime interpénétré (semi dilué), soit des globules denses (rayon ~10 nm) avec agrégation fractale à plus grande échelle lorsque, après contraction, les chaînes sont trop courtes et passent en régime désinterpénétré (dilué). Cette structure très bien définie a permis une étude fouillée: nous avons montré que le coeur des globules possède une charge nulle, que les espèces introduites en excès du point de vue électrostatique restent en solution, éventuellement dans des couronnes pour le PSS, et que leur taille finie est fixée par la force ionique. Une mesure spécifique de la localisation des contre-ions a prouvé qu'ils sont expulsés du coeur des complexes lors de leur formation. La conformation des chaînes au sein des complexes a été mesurée par marquage adapté. Enfin lorsque [-]/[+]>>1, un réseau transitoire fluide et limpide de protéines dénaturées par le PSS est obtenu.
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Jimenez, Clément. "Caracterisation de proteines synthetisees et secretees dans la tete de l'epididyme de souris dont l'expression est regulee par les androgenes." Clermont-Ferrand 1, 1989. http://www.theses.fr/1989CLF13824.
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Agez, Morgane. "ETUDE STRUCTURALE ET FONCTIONNELLE DE LA PROTEINE CHAPERON D'HISTONES ASF1." Phd thesis, Université Pierre et Marie Curie - Paris VI, 2008. http://tel.archives-ouvertes.fr/tel-00268886.
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Au sein des cellules eucaryotes, l'ADN est compacté sous la forme de chromatine dont l'unité de base est le nucléosome. Cette structure protéique protège l'ADN des agressions oxydantes et a un rôle essentiel dans la régulation des processus nucléaires. Ce travail a pour objectif de mieux comprendre les mécanismes de remodelage de la chromatine. Nous nous sommes plus particulièrement intéressés à l'interaction entre les protéines du nucléosome que sont les histones H3 et H4 et le chaperon d'histones Asf1. Nous avons résolu par RMN la structure du complexe entre Asf1 et les histones. Ce travail a constitué une base pour, d'une part, analyser la fonction de cette interaction in vivo et, d'autre part, initier la conception de peptides inhibiteurs spécifiques de cette interaction et anti-tumoraux potentiels. Au cours de ce travail, nous avons également développé une méthode in vitro d'étude des mécanismes moléculaires d'assemblage et de désassemblage de nucléosomes par Asf1.
Books on the topic "TET protease"
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Ten tea parties: Patriotic protests that history forgot. Philadelphia: Quirk Books, 2012.
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Eger, William Edmond. Analyzing the divisions in the Tea Party movement: The varieties of American political passion. Lewiston, N.Y: Edwin Mellen Press, 2012.
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Filiu, Jean-Pierre. The Arab revolution: Ten lessons from the democratic uprising. London: Hurst & Co, 2011.
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Kelm, Juan Ricardo. El Tractorazo: Crónica de una epopeya : la Asociación de Productores Agropecuarios de Misiones a 10 años de la gesta. Posadas, Misiones [Argentina]: Editorial Universitaria, Universidad Nacional de Misiones, 2013.
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Anderson, Stephanie Selene. Signs of freedom: The American Tea Party message. Fort Collins, Colo: Crowded Corner Press, 2009.
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Chuck, Asay, ed. Taxpayers' Tea Party: A manual for reclaiming our country. Riverdale, NY: Baen Pub. Enterprises, 2010.
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Cooper, Sharon. Taxpayers' Tea Party: A manual for reclaiming our country. Riverdale, NY: Baen Pub. Enterprises, 2010.
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Martino, Mazzonis, ed. Tea party: La rivolta populista e la destra americana. Venezia: Marsilio, 2012.
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Schmidt, Erin K. Tea and water pipe. [Rochester, Mich.]: Erin K. Schmidt, 2012.
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The Arab revolution: Ten lessons from the democratic uprising. London: Hurst & Co, 2011.
Find full textBook chapters on the topic "TET protease"
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Sun, Simon H. "The Hancocks’ Tea Trade and Origins of the American Revolution." In Protest in the Long Eighteenth Century, 141–58. Abingdon, Oxon ; New York, NY : Routledge, 2021. |: Routledge, 2021. http://dx.doi.org/10.4324/9780429275173-12.
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Polley, Tapasi, and Uma Ghosh. "Application of Used Tea as Solid Matrix for Immobilization of Alkaline Protease by OVAT Method." In Lecture Notes in Bioengineering, 219–29. Singapore: Springer Singapore, 2020. http://dx.doi.org/10.1007/978-981-15-7409-2_22.
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Jacomet, M., J. Breitenstein, R. Wälti, L. Winzenried, and M. Gysel. "ProTest: A Low Cost Rapid Prototypig and Test System for PCB, ASIC and FPGA." In Microelectronics Education, 161–64. Dordrecht: Springer Netherlands, 1998. http://dx.doi.org/10.1007/978-94-011-5110-8_38.
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"Protest." In A Time for Tea, 289–323. Duke University Press, 2001. http://dx.doi.org/10.1215/9780822380153-008.
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"8 Protest." In A Time for Tea, 289–324. Duke University Press, 2020. http://dx.doi.org/10.1515/9780822380153-010.
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"Aboriginal protest." In The Aboriginal Tent Embassy, 86–98. Routledge, 2013. http://dx.doi.org/10.4324/9780203771235-14.
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Campbell, Kimberlee Anne. "Introduction." In The Protean Text, 1–5. Routledge, 2019. http://dx.doi.org/10.4324/9780429061394-1.
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Campbell, Kimberlee Anne. "Historical Contexts." In The Protean Text, 7–13. Routledge, 2019. http://dx.doi.org/10.4324/9780429061394-2.
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Campbell, Kimberlee Anne. "Narrative Structures." In The Protean Text, 15–38. Routledge, 2019. http://dx.doi.org/10.4324/9780429061394-3.
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Campbell, Kimberlee Anne. "Generic Shift." In The Protean Text, 39–60. Routledge, 2019. http://dx.doi.org/10.4324/9780429061394-4.
Full textConference papers on the topic "TET protease"
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Guice, Justin, Caroline Best, Morgan Hollins, Kelly Tinker, and Sean Garvey. "Fungal Digestive Enzymes Promote Macronutrient Hydrolysis in the INFOGEST in vitro Simulation of Digestion." In 2022 AOCS Annual Meeting & Expo. American Oil Chemists' Society (AOCS), 2022. http://dx.doi.org/10.21748/agsn3911.
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Fungal enzymes are common ingredients in dietary supplements that support digestion. At adequate concentrations, exogenous enzymes may improve digestion by hydrolyzing macronutrients beyond acid-mediated hydrolysis, endogenous gastric pepsin, and pancreatic enzymes alone. The purpose of this study was to test the hydrolytic efficacy of 6 commercial fungal enzymes—three proteases, a lipase, an amylase, and a glucoamylase—in the INFOGEST static in vitro simulation of gastrointestinal digestion. The efficacy of 5 doses of each enzyme was determined by measuring free amino nitrogen (FAN), glycerol, maltose, and glucose concentrations following salivary-gastric (SG) and full salivary-gastric-intestinal (SGI) simulations of digestion of an oral nutritional supplement. In the SG simulation, the 3 proteases, lipase, and combination of amylase and glucoamylase promoted greater hydrolysis of dietary protein, fat, and carbohydrates, respectively, compared to control conditions. Acid protease (Aspergillus niger) treatment increased FAN concentrations than controls from 27% at the lowest dose to 142% greater than controls at the highest dose (p<0.0001). Fungal protease (A. oryzae) treatment increased FAN concentration at the highest dose (160,000 HUT, p=0.0115). Peptidase (A. melleus) treatment promoted higher FAN concentrations, up to 50% increase at the highest dose (160 LAPU, p<0.0001). Glycerol concentrations increased across all lipase treatments (p<0.0001), from 4.1-fold to 11.2-fold increases at the lowest and highest doses, respectively. All doses of amylase increased glucose concentrations (p<0.0001), and maltose concentrations started increasing at 4,000 SKB units (p=0.0010). In the SGI simulation, FAN concentrations following protease treatments were similar to control, suggesting little benefit beyond pancreatin alone in this static simulation of healthy digestion. Lipase and amylase/glucoamylase treatments, however, did increase glycerol (p<0.0001) and maltose/glucose concentrations (p<0.0001), respectively, compared to controls in the full SGI simulation. These data demonstrate that exogenous, fungal enzymes can improve macronutrient digestion under the acidic conditions of the gastric simulation, as well as the intestinal simulation.
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MAJEED, Huda Zuheir, Firas Nabeeh JAAFAR, Mohammed Twfeek ABID ALHUSAIN, Shatha Zuheir MAJEED, and Nadia Kamil BASHAR. "THE ANTIBACTERIAL EFFECTS OF GREEN TEA EXTRACT ON RESISTANT BACTERIA ISOLATED FROM HUMAN EYE INFECTIONS." In IV.International Scientific Congress of Pure,Appliedand Technological Sciences. Rimar Academy, 2022. http://dx.doi.org/10.47832/minarcongress4-28.
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Ocular infection is a world wide issue especially for public health field which could be a result to its own normal flora due to subjection to external factors (e.g. stress, getting older, hits, surgical operations, systemic diseases and losing commensal flora). Ocular pathogens could be healed by a group of topical antibiotics, but with time, drug resistance had been developed, which more magnified by wrong diagnosis and random use of antibiotics leading to unexpected complications e.g. visual problems, leading to blindness at last . Alternative therapy had been used to treat such infections including plant extracts like Green Tea (Camellia sinensis) . Eye swabs about (50) samples were gathered from people had ocular infections,then biochemical tests diagnosed (30) bacterial isolates. There were (17) isolates (6 isolates of Staphylococcus spp. and 11 isolates were Enterococcus) out of the (30) isolates showed multiple antibiotic resistance to nine antibiotics by disc-diffusion method,there were high complete resistance to Moxifloxacin and Bacitracin, in contrast to Ciprofloxacin and Chloramphenicol. The antibacterial effects of hot water, cold water,Acetone, Ethanol and Methanol Green tea extracts was examined against the (17) multiple antibiotic resistant isolates by agar-well diffusion method using. Only the Ethanol and Methanol green tea extract showed promising results, without any effect of the remaining green tea extracts. Green tea extracts were equal to Ciprofloxacin and Sulphamethoxazole in effectiveness against antibiotic resistant isolates . The (17) isolates were tested for production of biofilm and protease. (12) isolates were biofilm-producer but after subjection to Ethanol Green tea extract changed into non biofilmformer. (13) isolate were protease-producer but after subjection to Ethanol Green tea extract changed into non protease-former. Key words: Eye Swabs, Antibiotic Resistance, Alternative Therapy, Green Tea Extracts, Biofilm and Protease.
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Caselli, Tommaso, Osman Mutlu, Angelo Basile, and Ali Hürriyetoğlu. "PROTEST-ER: Retraining BERT for Protest Event Extraction." In Proceedings of the 4th Workshop on Challenges and Applications of Automated Extraction of Socio-political Events from Text (CASE 2021). Stroudsburg, PA, USA: Association for Computational Linguistics, 2021. http://dx.doi.org/10.18653/v1/2021.case-1.4.
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Ahmad, Sana. "Multienzyme System as detergent additive to improve cleaning potential." In INTERNATIONAL CONFERENCE ON BIOLOGICAL RESEARCH AND APPLIED SCIENCE. Jinnah University for Women, Karachi,Pakistan, 2022. http://dx.doi.org/10.37962/ibras/2022/8-9.
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The detergent industry represents approximately 30% of the industrial contribution of enzymes. Detergents are widely used in dishwashing liquids, laundry materials, domestic, industrial and institutional cleaning agents. Various harmful effects has been associated with incomplete degradation of synthetic detergents after utilization. Enrichment of the detergents with enzymes significantly influence the growth and development of detergent industry1. The enzyme-added detergents are ecofriendly, economic, and highly efficient in cleaning action. Hydrolytic enzymes have been extensively exploited for removing the strains2. In present research, bacterial protease and lipase produced separately and washing potential of crude enzymes had been investigated. Lipase and protease used separately and with detergents in eleven different combinations to remove tea and oil stains. According to the results, combination of these hydrolases along with detergent has been shown to possess the highest cleaning action3. The diameter of the tea stain reduced to 64 % and 72% reduction was observed in oil stain along with significant decline in its visibility. These findings suggests the potential use of lipase and protease combination in biodetergent formulation.
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Bambhaniya, Abhimanyu Rajeshkumar, Yangyu Chen, Anshuman, Rohan Banerjee, and Tushar Krishna. "Proteus : HLS-based NoC Generator and Simulator." In 2023 Design, Automation & Test in Europe Conference & Exhibition (DATE). IEEE, 2023. http://dx.doi.org/10.23919/date56975.2023.10137173.
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Rammadan ABDUL, Fatima, Ihsan Ali RAHEEM, and Batool Abd Al Ameer BAQER. "DETECTION OF BIOFILM FORMATION AND RESISTANCE TO SOME ANTIBIOTICS OF ESCHERICHIA COLI." In VI.International Scientific Congress of Pure,Applied and Technological Sciences. Rimar Academy, 2022. http://dx.doi.org/10.47832/minarcongress6-19.
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The study included twenty isolates of Escherichia coli isolated from different disease samples. It was diagnosis by microscopic methods, culture and biochemical tests and confirmed by Vitek 2 system. The ability of the isolates to produce protease showed that (9) isolates out of a total of (20) isolates with a percentage of 45% were protease -producing and 55% not producing. Biofilm formation by MTP method (15) isolates showed a percentage of 75% of a positive result for biofilm formation, while the percentage of negative isolates was 25%. The sensitivity study was conducted for the isolates against five antibiotics, and it was found that the isolates have multi drug resistance. All clinical isolates of E.coli were shown ( 100 %) resistant to piperacillin and ceftazidime. Bacteria for the rest of the antibiotics showed the least resistance to meropenem . All ten isolates were subjected to a beta-lactamase production test. Six were enzyme-producing, while four were non-enzyme producing. In addition, the minimum inhibitory concentration (MICs) for isolates were determined to five antibiotics included (Piperacillin, Ceftazidime, Azithromycin, imipenem and meropenem). The values of (MICs) for the two antibiotics ranged between (64-512) µg/ml, Piperacillin and Ceftazidime, While the meropenem(MIC) value ranged between(4-32) µg/ml
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Liu, Ze, Min Xiong, Debbie Liao, Dan Lv, Ralph A. Reisfeld, Wolfgang Wrasidlo, Si Chen, et al. "Abstract 5614: Legumain protease substrate modified TAT-liposome cargo: an efficient tool for targeting malignant diseases." In Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1538-7445.am2013-5614.
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Radovanović, Marko, Ignjat Filipović, Maja Đukić, Marija Ristić, Ivan Jakovljević, and Zoran D. Matović. "Molecular docking study of ruthenium-p-cymene complexes with isothiazole derivatives as SARS-CoV-2 main protease inhibitors." In 2nd International Conference on Chemo and Bioinformatics. Institute for Information Technologies, University of Kragujevac, 2023. http://dx.doi.org/10.46793/iccbi23.387r.
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Since proper treatment for COVID-19 still has not been developed, exploration of novel options is required. Activities of different metal complexes, promising results gained from examining different thiazole derivatives, and research in the field of natural products like p-cymene, produced an idea to test piano stool ruthenium p-cymene complexes with isothiazole derivatives as ligands. In silico methods are often used as the first step in a series of experiments during the development of new drugs, and docking simulations are a quick way to determine the feasibility of novel compounds as potential inhibitors of target enzymes. Existing compounds of ruthenium with published crystal structures were tested against the main protease of SARS-CoV-2. All of the tested compounds show a potential ability to bind to the target enzyme, while the compound with phenyl and morpholinyl substituents in isothiazole ligand shows the best activity among tested compounds. Authors feel confident that further research on this topic will yield compounds with even better potential activities against the main protease of the SARS-CoV-2.
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Schapira, M., B. Waeber, H. R. Brunner, R. Crystal, and M. Courtney. "PROTECTION BY α1-ANTITRYPSIN ALA-357 ARG-358 AGAINST ARTERIAL HYPOTENSION INDUCED BY FACTOR XII FRAGMENT." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642801.
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The specificity of serine protease inhibitors belonging to the serpin superfamily depends on the nature of the reactive center amino acid residues. For example, Met→Arg mutation at the reactive center P1 residue (position 358) alters the specificity of α1-antitrypsin (AT) from the Met-specific enzyme neutrophil elastase to the Arg-specific proteases thrombin, plasma kallikrein (K) and activated Factor XII fragment (XIIf). To obtain an inhibitor species which would inhibit K and XIIf but not thrombin, we now have produced by site-directed mutagenesis of cloned AT cDNA an AT variant having Arg at P1 and Ala at P2. This modification at P2 was made because C1-inhibitor, the major inhibitor of K and XIIf, also has Ala at P2. In purified systems, AT Ala-357 Arg-358 inactivated thrombin, K and XIIf with 2nd-order rate constants of 1.1, 21.8 and 0.6 μM-1 min-1 whereas values of 8.5, 4.2 and 2.1 μM-1 min-1 were found with AT Arg-358. Thus, when compared to AT Arg-358, AT Ala-357 Arg-358 was 5.2 times more efficient for inhibiting K but 7.7 times less efficient for inhibiting thrombin. In vivo, AT Ala-357 Arg-358 (0.7 mg i.v.) did not modify the thrombin time of male Wistar rats while a 2-fold prolongation was seen with 0.7 mg AT Arg-358. However, AT Ala-357 Arg-358 (0.7 mg i.v.) partially prevented the kinin-mediated circulatory collapse induced by XIIf (0.1 μg i.v.) since rats (n=4) treated with this double mutant had a blood pressure fall of 14 ±3 (meaniSD) mmHg while control animals (n = 8) receiving saline or AT Val-358 (0.7 mg i.v.) had a decrease of 27 ± 3 mmHg (p<0.01 by t test). AT Ala-357 Arg-358 has therapeutic potential for disease states with activation of the plasma kinin-forming system such as angioedema attacks or septic shock.
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Chan, Ka H., Sze C. Yeung, Mary S. Ip, Ricky Y. K. Man, and Judith C. Mak. "Effects Of Chinese Green Tea On Cigarette Smoke-induced Lung Inflammation, Oxidative Stress And Protease Activity In Rats." In American Thoracic Society 2010 International Conference, May 14-19, 2010 • New Orleans. American Thoracic Society, 2010. http://dx.doi.org/10.1164/ajrccm-conference.2010.181.1_meetingabstracts.a5062.
Full textReports on the topic "TET protease"
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Stanley, Stephanie. The Tea Party and Occupy Wall Street Movements: Populism and Protest. Portland State University Library, January 2012. http://dx.doi.org/10.15760/honors.2.
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Connaway, H. M., and C. H. Lee. Preliminary Analysis of the Transient Reactor Test Facility (TREAT) with PROTEUS. Office of Scientific and Technical Information (OSTI), November 2015. http://dx.doi.org/10.2172/1227442.
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Manulis, Shulamit, Christine D. Smart, Isaac Barash, Guido Sessa, and Harvey C. Hoch. Molecular Interactions of Clavibacter michiganensis subsp. michiganensis with Tomato. United States Department of Agriculture, January 2011. http://dx.doi.org/10.32747/2011.7697113.bard.
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Clavibacter michiganensis subsp. michiganensis (Cmm), the causal agent of bacterial wilt and canker of tomato, is the most destructive bacterial disease of tomato causing substantial economic losses in Israel, the U.S.A. and worldwide. The molecular strategies that allow Cmm, a Gram-positive bacterium, to develop a successful infection in tomato plants are largely unknown. The goal of the project was to elucidate the molecular interactions between Cmmand tomato. The first objective was to analyze gene expression profiles of susceptible tomato plants infected with pathogenic and endophytic Cmmstrains. Microarray analysis identified 122 genes that were differentially expressed during early stages of infection. Cmm activated typical basal defense responses in the host including induction of defense-related genes, production of scavenging of free oxygen radicals, enhanced protein turnover and hormone synthesis. Proteomic investigation of the Cmm-tomato interaction was performed with Multi-Dimensional Protein Identification Technology (MudPIT) and mass spectroscopy. A wide range of enzymes secreted by Cmm382, including cell-wall degrading enzymes and a large group of serine proteases from different families were identified in the xylem sap of infected tomato. Based on proteomic results, the expression pattern of selected bacterial virulence genes and plant defense genes were examined by qRT-PCR. Expression of the plasmid-borne cellulase (celA), serine protease (pat-1) and serine proteases residing on the chp/tomA pathogenicity island (chpCandppaA), were significantly induced within 96 hr after inoculation. Transcription of chromosomal genes involved in cell wall degradation (i.e., pelA1, celB, xysA and xysB) was also induced in early infection stages. The second objective was to identify by VIGS technology host genes affecting Cmm multiplication and appearance of disease symptoms in plant. VIGS screening showed that out of 160 tomato genes, which could be involved in defense-related signaling, suppression of 14 genes led to increase host susceptibility. Noteworthy are the genes Snakin-2 (inhibitor of Cmm growth) and extensin-like protein (ELP) involved in cell wall fortification. To further test the significance of Snakin -2 and ELP in resistance towards Cmm, transgenic tomato plants over-expressing the two genes were generated. These plants showed partial resistance to Cmm resulting in a significant delay of the wilt symptoms and reduction in size of canker lesion compared to control. Furthermore, colonization of the transgenic plants was significantly lower. The third objective was to assess the involvement of ethylene (ET), jasmonate (JA) and salicylic acid (SA) in Cmm infection. Microarray and proteomic studies showed the induction of enzymes involved in ET and JA biosynthesis. Cmm promoted ET production 8 days after inoculation and SIACO, a key enzyme of ET biosynthesis, was upregulated. Inoculation of the tomato mutants Never ripe (Nr) impaired in ET perception and transgenic plants with reduced ET synthesis significantly delayed wilt symptoms as compared to the wild-type plants. The retarded wilting in Nr plants was shown to be a specific effect of ET insensitivity and was not due to altered expression of defense related genes, reduced bacterial population or decrease in ethylene biosynthesis . In contrast, infection of various tomato mutants impaired in JA biosynthesis (e.g., def1, acx1) and JA insensitive mutant (jai1) yielded unequivocal results. The fourth objective was to determine the role of cell wall degrading enzymes produced by Cmm in xylem colonization and symptoms development. A significance increase (2 to 7 fold) in expression of cellulases (CelA, CelB), pectate lyases (PelA1, PelA2), polygalacturonase and xylanases (XylA, XylB) was detected by qRT-PCR and by proteomic analysis of the xylem sap. However, with the exception of CelA, whose inactivation led to reduced wilt symptoms, inactivation of any of the other cell wall degrading enzymes did not lead to reduced virulence. Results achieved emphasized the complexity involved in Cmm-tomato interactions. Nevertheless they provide the basis for additional research which will unravel the mechanism of Cmm pathogenicity and formulating disease control measures.
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Dolja, Valerian V., Amit Gal-On, and Victor Gaba. Suppression of Potyvirus Infection by a Closterovirus Protein. United States Department of Agriculture, March 2002. http://dx.doi.org/10.32747/2002.7580682.bard.
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The plant virus family Polyviridae is the largest and most destructive of all plant viruses. Despite the continuous effort to develop resistant plant varieties, there is a desperate need for novel approaches conferring wide-range potyvirus resistance. Based on experiments with the tobacco etch potyvirus (TEV)-derived gene expression vector, we suggested approach for screening of the candidate resistance genes. This approach relies on insertion of the genes into a virus vector and evaluation of the phenotypes of the resulting recombinant viruses. The genes which suppress infection by the recombinant virus are selected as candidates for engineering transgenic resistance. Our analysis of the TEV variants expressing proteins of the beet yellows closterovirus (BYV) revealed that one of those, the leader proteinase (L-Pro), strongly and specifically interfered with the hybrid TEV infection. Since closterovirus L-Pro is evolutionary related to potyviral helper component-proteinase (HC-Pro), we suggested that the L-Pro interfered with HC-Pro function via a trans-dominant inhibitory effect. Based on these findings, we proposed to test two major hypotheses. First, we suggested that L-Pro-mediated suppression of potyvirus infection is a general phenomenon effective against a range of potyviruses. The second hypothesis stated that the suppression effect can be reproduced in transgenic plants expressing L-Pro, and can be utilized for generation of resistance to potyviruses. In accord with these hypotheses, we developed two original objectives of our proposal: A) to determine the range of the closterovirus-derived suppression of potyviral infection, and B) to try and utilize the L-Pro-mediated suppression for the development of transgenic resistance to potyviruses. In the first phase of the project, we have developed all major tools and technologies required for successful completion of the proposed research. These included TEV and ZYMV vectors engineered to express several closteroviral L-Pro variants, and generation of the large collection of transgenic plants. To our satisfaction, characterization of the infection phenotypes exhibited by chimeric TEV and ZYMV variants confirmed our first hypothesis. For instance, similar to TEV-L- Pro(BYV) chimera, ZYMV-L-Pro(LIYV) chimera was debilitated in its systemic spread. In contrast, ZYMV-GUS chimera (positive control) was competent in establishing vigorous systemic infection. These and other results with chimeric viruses indicated that several closteroviral proteinases inhibit long-distance movement of the potyviruses upon co-expression in infected plants. In order to complete the second objective, we have generated ~90 tobacco lines transformed with closteroviral L-Pro variants, as well as ~100 lines transformed with BYV Hsp70-homolog (Hsp70h; a negative control). The presence and expression of the trans gene in each line was initially confirmed using RT-PCR and RNA preparations isolated from plants. However, since detection of the trans gene-specific RNA can not guarantee production of the corresponding protein, we have also generated L-Pro- and Hsp70h-specific antisera using corresponding synthetic peptides. These antisera allowed us to confirm that the transgenic plant lines produced detectable, although highly variable levels of the closterovirus antigens. In a final phase of the project, we tested susceptibility of the transgenic lines to TEV infection. To this end, we determined that the minimal dilution of the TEV inoculum that is still capable of infecting 100% of nontransgenic plants was 1:20, and used 10 plants per line (in total, ~2,000 plants). Unfortunately, none of the lines exhibited statistically significant reduction in susceptibility. Although discouraging, this outcome prompted us to expand our experimental plan and conduct additional experiments. Our aim was to test if closteroviral proteinases are capable of functioning in trans. We have developed agroinfection protocol for BYV, and tested if co- expression of the L-Pro is capable of rescuing corresponding null-mutant. The clear-cut, negative results of these experiments demonstrated that L-Pro acts only in cis, thus explaining the lack of resistance in our transgenic plants. We have also characterized a collection of the L-Pro alanine- scanning mutants and found direct genetic evidence of the requirement for L-Pro in virus systemic spread. To conclude, our research supported by BARD confirmed one but not another of our original hypotheses. Moreover, it provided an important insight into functional specialization of the viral proteinases and generated set of tools and data with which we will be able to address the molecular mechanisms by which these proteins provide a variety of critical functions during virus life cycle.
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Chamovitz, Daniel A., and Albrecht G. Von Arnim. eIF3 Complexes and the eIF3e Subunit in Arabidopsis Development and Translation Initiation. United States Department of Agriculture, September 2009. http://dx.doi.org/10.32747/2009.7696545.bard.
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The original working hypothesis of our proposal was that The “e” subunit of eIF3 has multiple functions from both within the nucleus and in the cytoplasm. Within this model, we further hypothesized that the “e” subunit of eIF3 functions in translation as a repressor. We proposed to test these hypotheses along the following specific aims: 1) Determine the subcellular localization of the interaction between eIF3e and other eIF3 subunits, or the COP9 signalosome. 2) Elucidate the biological significance of the varied subcellular localizations of eIF3e through generating Arabidopsis eIF3e alleles with altered subcellular localization. 3.) Purify different eIF3e complexes by tandem affinity purification (TAP). 4) Study the role of eIF3e in translational repression using both in vitro and in planta assays. eIF3 is an evolutionarily ancient and essential component of the translational apparatus in both the plant and animal kingdoms. eIF3 is the largest, and in some ways the most mysterious, of the translation factors. It is a multi-subunit protein complex that has a structural/scaffolding role in translation initiation. However, despite years of study, only recently have differential roles for eIF3 in the developmental regulation of translation been experimentally grounded. Furthermore, the roles of individual eIF3 subunits are not clear, and indeed some, such as the “e” subunit may have roles independent of translation initiation. The original three goals of the proposal were technically hampered by a finding that became evident during the course of the research – Any attempt to make transgenic plants that expressed eIF3e wt or eIF3e variants resulted in seedling lethality or seed inviability. That is, it was impossible to regenerate any transgenic plants that expressed eIF3e. We did manage to generate plants that expressed an inducible form of eIF3e. This also eventually led to lethality, but was very useful in elucidating the 4th goal of the research (Yahalom et al., 2008), where we showed, for the first time in any organism, that eIF3e has a repressory role in translation. In attempt to solve the expression problems, we also tried expression from the native promoter, and as such analyzed this promoter in transgenic plants (Epel, 2008). As such, several additional avenues were pursued. 1) We investigated protein-protein interactions of eIF3e (Paz-Aviram et al., 2008). 2) The results from goal #4 led to a novel hypothesis that the interaction of eIF3e and the CSN meets at the control of protein degradation of nascent proteins. In other words, that the block in translation seen in csn and eIF3e-overexpressing plants (Yahalom et al., 2008) leads to proteasome stress. Indeed we showed that both over expression of eIF3e and the csn mutants lead to the unfolded protein response. 3) We further investigated the role of an additional eIF3 subunit, eIF3h, in transalational regulation in the apical meristem (Zhou et al., 2009). Epel, A. (2008). Characterization of eIF3e in the model plant Arabidopsis thaliana. In Plant Sciences (Tel Aviv, Tel Aviv University). Paz-Aviram, T., Yahalom, A., and Chamovitz, D.A. (2008). Arabidopsis eIF3e interacts with subunits of the ribosome, Cop9 signalosome and proteasome. Plant Signaling and Behaviour 3, 409-411. Yahalom, A., Kim, T.H., Roy, B., Singer, R., von Arnim, A.G., and Chamovitz, D.A. (2008). Arabidopsis eIF3e is regulated by the COP9 signalosome and has an impact on development and protein translation. Plant J 53, 300-311. Zhou, F., Dunlap, J.R., and von Arnim, A.G. The translation initiation factor subunit eIF3h is .1 involved in Arabidopsis shoot apical meristem maintenance and auxin response. (submitted to Development).
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Grumet, Rebecca, and Benjamin Raccah. Identification of Potyviral Domains Controlling Systemic Infection, Host Range and Aphid Transmission. United States Department of Agriculture, July 2000. http://dx.doi.org/10.32747/2000.7695842.bard.
Full textAbstract:
Potyviruses form one of the largest and most economically important groups of plant viruses. Individual potyviruses and their isolates vary in symptom expression, host range, and ability to overcome host resistance genes. Understanding factors influencing these biological characteristics is of agricultural importance for epidemiology and deployment of resistance strategies. Cucurbit crops are subject to severe losses by several potyviruses including the highly aggressive and variable zucchini yellow mosaic virus (ZYMV). In this project we sought to investigate protein domains in ZYMV that influence systemic infection and host range. Particular emphasis was on coat protein (CP), because of known functions in both cell to cell and long distance movement, and helper component-protease (HC-Pro), which has been implicated to play a role in symptom development and long distance movement. These two genes are also essential for aphid mediated transmission, and domains that influence disease development may also influence transmissibility. The objectives of the approved BARD project were to test roles of specific domains in the CP and HC-Pro by making sequence alterations or switches between different isolates and viruses, and testing for infectivity, host range, and aphid transmissibility. These objectives were largely achieved as described below. Finally, we also initiated new research to identify host factors interacting with potyviral proteins and demonstrated interaction between the ZYMV RNA dependent RNA polymerase and host poly-(A)-binding protein (Wang et al., in press). The focus of the CP studies (MSU) was to investigate the role of the highly variable amino terminus (NT) in host range determination and systemic infection. Hybrid ZYMV infectious clones were produced by substituting the CP-NT of ZYMV with either the CP-NT from watermelon mosaic virus (overlapping, but broader host range) or tobacco etch virus (TEV) (non- overlapping host range) (Grumet et al., 2000; Ullah ct al., in prep). Although both hybrid viruses initially established systemic infection, indicating that even the non-cucurbit adapted TEV CP-NT could facilitate long distance transport in cucurbits, after approximately 4-6, the plants inoculated with the TEV-CPNT hybrid exhibited a distinct recovery of reduced symptoms, virus titer, and virus specific protection against secondary infection. These results suggest that the plant recognizes the presence of the TEV CP-NT, which has not been adapted to infection of cucurbits, and initiates defense responses. The CP-NT also appears to play a role in naturally occurring resistance conferred by the zym locus in the cucumber line 'Dina-1'. Patterns of virus accumulation indicated that expression of resistance is developmentally controlled and is due to a block in virus movement. Switches between the core and NT domains of ZYMV-NAA (does not cause veinal chlorosis on 'Dina-1'), and ZYMV-Ct (causes veinal chlorosis), indicated that the resistance response likely involves interaction with the CP-NT (Ullah and Grumet, submitted). At the Volcani Center the main thrust was to identify domains in the HC-Pro that affect symptom expression or aphid transmissibility. From the data reported in the first and second year report and in the attached publications (Peng et al. 1998; Kadouri et al. 1998; Raccah et al. 2000: it was shown that: 1. The mutation from PTK to PAK resulted in milder symptoms of the virus on squash, 2. Two mutations, PAK and ATK, resulted in total loss of helper activity, 3. It was established for the first time that the PTK domain is involved in binding of the HC-Pro to the potyvirus particle, and 4. Some of these experiments required greater amount of HC-Pro, therefore a simpler and more efficient purification method was developed based on Ni2+ resin.
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Tucker, Mark L., Shimon Meir, Amnon Lers, Sonia Philosoph-Hadas, and Cai-Zhong Jiang. Elucidation of signaling pathways that regulate ethylene-induced leaf and flower abscission of agriculturally important plants. United States Department of Agriculture, January 2012. http://dx.doi.org/10.32747/2012.7597929.bard.
Full textAbstract:
The Problem: Abscission is a highly regulated process, occurring as a natural terminal stage of development, in which various organs are separated from the parent plant. In most plant species, the process is initiated by a decrease in active auxin in the abscission zone (AZ) and an increase in ethylene, and may be accelerated by postharvest or environmental stresses. Another potential key regulator in abscission is IDA (Inflorescence Deficient in Abscission), which was identified as an essential peptide signal for floral organ abscission in Arabidopsis. However, information is still lacking regarding the molecular mechanisms integrating all these regulators. In our previous BARD funded research we made substantial progress towards understanding these molecular events in tomato, and the study is still in progress. We established a powerful platform for analysis of genes for regulatory proteins expressed in AZ. We identified changes in gene expression for several transcription factors (TFs) directly linked to ethylene and auxin signaling and several additional regulatory proteins not so obviously linked to these hormones. Moreover, we demonstrated using a virus-induced gene silencing (VIGS) assay that several play a functional role in the onset of abscission. Based on these results we have selected 14 genes for further analysis in stably transformed tomato plants. All 14 genes were suppressed by RNA interference (RNAi) using a constitutive promoter, and 5 of them were also suppressed using an abscission-specific promoter. Transformations are currently at different stages of progress including some lines that already display an abscission phenotype. Objectives: We propose here to (1) complete the functional analysis of the stably transformed tomato plants with T2 lines and perform transcriptome analysis using custom abscission-specific microarrays; (2) conduct an indepth analysis of the role of IDA signaling in tomato leaf and flower abscission; (3) perform transcriptome and proteome analyses to extend the earlier gene expression studies to identify transcripts and proteins that are highly specific to the separation layer (i.e., target cells for cell separation) prior to the onset of abscission; (4) extend and compliment the work in tomato using a winnowed set of genes in soybean. Methodology: Next Generation Sequencing (NGS) of mRNA will be used to further increase the list of abscission-associated genes, and for preparation of a custom tomato abscission microarray to test altered gene expression in transgenic plants. Tandem mass spectrometry (LC-MS/MS) of protein extracts from leaf petiole, flower pedicel and their AZ tissues will be used to identify the proteome of the AZ before and during abscission. AZ-specific gene promoters will be used in stably transformed tomato plants to reduce non-target phenotypes. The bean pod mottle virus (BPMV) plasmid vectors will be used for VIGS analysis in soybean. Expected Contribution: Our study will provide new insights into the regulation of ethylene-induced abscission by further revealing the role of key regulators in the process. This will permit development of novel techniques for manipulating leaf and flower abscission, thereby improving the postharvest performance of agriculturally important crops.