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Crowley, Cara Leilani. "Bile salt induced stress response pathways." Diss., The University of Arizona, 2000. http://hdl.handle.net/10150/289231.
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Our lab has shown that the bile salt found in the highest concentration in human fecal water, sodium deoxycholate, induces apoptosis in several cell types including Jurkat cells as well as human colonic epithelial cells. We have also found that cells within the normal appearing flat mucosa of patients with a history of colon cancer are relatively resistant to apoptosis induced by NaDOC. The current studies test the hypothesis that sodium deoxycholate induces multiple stress response pathway s that protect against apoptosis. I have tested this hypothesis by developing and analyzing cell lines that are resistant to sodium deoxycholate-induced apoptosis and focusing on two stress-response proteins known to be activated by sodium deoxycholate, poly(ADP-ribose) polymerase (PARP) and the redo-sensitive transcription factor nuclear factor-kappa B (NF-κB). I found that PARP is protective against NaDOC-induced apoptosis, and by independently inhibiting the individual subunits of NF-κB, I found that the p65 subunit is protective, while the p50 subunit is not. Development and subsequent characterization of the NaDOC-resistant HCT-116 cell lines identified several proteins that may be responsible for the development of apoptosis resistance. These proteins will be further tested in future studies.
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Bouman, Lena. "A role of parkin in stress response pathways." Diss., lmu, 2010. http://nbn-resolving.de/urn:nbn:de:bvb:19-120918.
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Mutavchiev, Delyan Rumenov. "Regulation of fission yeast cell polarity by stress-response pathways." Thesis, University of Edinburgh, 2017. http://hdl.handle.net/1842/29006.
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Cell polarisation is a key biological process crucial for the functioning of essentially all cells. Regulation of cell polarity is achieved through various processes determined by both internal and external factors. An example of the latter is that cell polarity can be disrupted or lost as a consequence of a variety of external stresses. When facing such stresses, cells adapt to unfavourable conditions by activating a range of molecular signalling pathways, collectively termed ‘stress response’. Despite the connections between external stress and cell polarity, whether stress-response signalling regulates cell polarisation and what the molecular basis for such regulation remains an open question. The fission yeast Schizosaccharomyces pombe presents an excellent biological platform to study the complexity of cell polarity regulation on a systematic level. This study is aimed at understanding the functional relationship between stress-response signalling and maintenance of cell polarity in this model organism. The findings presented in this thesis set the basis for establishing a functional link between the activation of the S.pombe stress-response pathway and the activity of the master regulator of cell polarity- the Rho GTPase Cdc42. Here, I describe experiments that identify an active involvement of the stress-response mitogen-activated kinase (MAPK) Sty1 in the dispersal of active Cdc42 from the sites of growth. This new role for Sty1 occurs independently from its involvement in transcription regulation and other previously identified signalling pathways involving Sty1. Furthermore, I also find that Sty1’s involvement in Cdc42 regulation has direct implications for fission yeast physiology as it is essential for the maintenance of cellular quiescence upon nitrogen starvation. This thesis also focuses on identifying the targets of Sty1 orchestrating the active Cdc42 disruption. Here, I describe a candidate-based approach, where I investigate the role of proteins from the Cdc42 regulatory network during Sty1 activation. Additionally, I present a global phospho-proteomics approach to identify novel targets of Sty1 and offer preliminary findings which might explain Sty1’s involvement in Cdc42 regulation.
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Chalmers, Fiona. "Improving protein yield from mammalian cells by manipulation of stress response pathways." Thesis, University of Glasgow, 2016. http://theses.gla.ac.uk/7666/.
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Monoclonal antibodies are a class of therapeutic that is an expanding area of the lucrative biopharmaceutical industry. These complex proteins are predominantly produced from large cultures of mammalian cells; the industry standard cell line being Chinese Hamster Ovary (CHO) cells. A number of optimisation strategies have led to antibody titres from CHO cells increasing by a hundred-fold, and it has been proposed that a further bottleneck in biosynthesis is in protein folding and assembly within the secretory pathway. To alleviate this bottleneck, a CHO-derived host cell line was generated by researchers at the pharmaceutical company UCB that stably overexpressed two critical genes: XBP1, a transcription factor capable of expanding the endoplasmic reticulum and upregulating protein chaperones; and Ero1α, an oxidase that replenishes the machinery of disulphide bond formation. This host cell line, named CHO-S XE, was confirmed to have a high yield of secreted antibody. The work presented in this thesis further characterises CHO-S XE, with the aim of using the information gained to lead the generation of novel host cell lines with more optimal characteristics than CHO-S XE. In addition to antibodies, it was found that CHO-S XE had improved production of two other secreted proteins: one with a simple tertiary structure and one complex multi-domain protein; and higher levels of a number of endogenous protein chaperones. As a more controlled system of gene expression to unravel the specific roles of XBP1 and Ero1α in the secretory properties of CHO-S XE, CHO cells with inducible overexpression of XBP1, Ero1α, or a third gene involved in the Unfolded Protein Response, GADD34, were generated. From these cell lines, it was shown that more antibody was secreted by cells with induced overexpression of XBP1; however, Ero1α and GADD34 overexpression did not improve antibody yield. Further investigation revealed that endogenous XBP1 splicing was downregulated in the presence of an abundance of the active form of XBP1. This result indicated a novel aspect of the regulation of the activity of IRE1, the stress-induced endoribonuclease responsible for XBP1 splicing. Overall, the work described in this thesis confirms that the overexpression of XBP1 has an enhancing effect on the secretory properties of CHO cells; information which could contribute to the development of host cells with a greater capacity for antibody production.
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Richards, Siân Louise. "The involvement of Arabidopsis thaliana Annexin 1 in abiotic stress response pathways." Thesis, University of Cambridge, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648626.
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Edwards, Clare B. "The effects of supplemented metabolites on lifespan and stress response pathways in Caenorhabditis elegans." Scholar Commons, 2015. http://scholarcommons.usf.edu/etd/5681.
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Understanding how metabolites contribute to anaplerosis, antioxidant effects, and hormetic pathways during aging is fundamental to creating supplements and dietary habits that may decrease age-associated disease and decline, thus improving the quality of life in old age. In order to uncover metabolic pathways that delay aging, the effects of large sets of metabolites associated with mitochondrial function on lifespan were investigated.
Malate, the tricarboxylic acid (TCA) cycle metabolite, increased lifespan and thermotolerance in C. elegans. Addition of fumarate and succinate also extended lifespan and all three metabolites activated nuclear translocation of the cytoprotective DAF-16/FOXO transcription factor and protected from paraquat-induced oxidative stress. The increased longevity provided by malate addition did not occur in fumarase (fum-1), glyoxylate shunt (gei-7), succinate dehydrogenase flavoprotein (sdha-2), or soluble fumarate reductaseF48E8.3 RNAi knockdown worms. Therefore, to increase lifespan, malate must be first converted to fumarate, then fumarate must be reduced to succinate by soluble fumarate reductase and the mitochondrial electron transport chain complex II. Lifespan extension induced by malate depended upon the longevity regulators DAF-16 and SIR-2.1. Malate supplementation did not extend the lifespan of long-lived eat-2 mutant worms, a model of dietary restriction. Malate and fumarate addition increased oxygen consumption, but decreased ATP levels and mitochondrial membrane potential suggesting a mild uncoupling of oxidative phosphorylation.
Each of the twenty amino acids was individually supplemented to C. elegans and the effects on lifespan were determined. All amino acids except phenylalanine were found to extend lifespan at least to a small extent at one or more of the 3 concentrations tested with serine, histidine, and proline showing the largest effects. In most cases, amino acid supplementation did not extend lifespan in eat-2 worms, a model of dietary restriction or in daf-16, sir-2.1, rsks-1 (S6 kinase), or aak-2 (AMPK) longevity pathway mutants or in worms fed RNAi to skn-1, the C. elegans Nrf2 homolog. Serine and tryptophan addition further protected worms from Alzheimer’s amyloid-beta toxicity. Tryptophan and its catabolites nicotinic acid, picolinic acid, and NAD further induced a broad heat shock response. These results indicate that dietary amino acid imbalance and amino acid catabolism affect organismal longevity.
The ketone body beta-hydroxybutyrate (βHB) is a histone deacetylase (HDAC) inhibitor and has been shown to be protective in many disease models, but its effects on aging are not well studied. Therefore we determined the effect of βHB supplementation on the lifespan of C. elegans. βHB supplementation extended mean lifespan by approximately 20%. RNAi knockdown of HDACs hda-2 or hda-3 also increased lifespan and further prevented βHB-mediated lifespan extension. βHB-mediated lifespan extension required the DAF-16/FOXO and SKN-1/Nrf longevity pathways, the sirtuin SIR-2.1, and the AMP kinase subunit AAK-2. βHB did not extend lifespan in a genetic model of dietary restriction indicating that βHB is likely functioning through a similar mechanism. βHB addition also upregulated ΒHB dehydrogenase activity and increased oxygen consumption in the worms. RNAi knockdown of F55E10.6, a short chain dehydrogenase and SKN-1 target gene, prevented the increased lifespan and βHB dehydrogenase activity induced by βHB addition, suggesting that F55E10.6 functions as an inducible βHB dehydrogenase. Furthermore, βHB supplementation delayed Alzheimer's amyloid-beta toxicity and decreased Parkinson's alpha-synuclein aggregation. The results indicate that D-βHB extends lifespan through inhibiting HDACs and through the activation of conserved stress response pathways.
Aging is a progressive disease caused by the time dependent decline of an organism and is the primary risk factor for many human ailments, including heart disease, cancer, and Alzheimer’s disease. Uncovering metabolic pathways and metabolites that delay the onset of age-related decline was the primary drive of this investigation.
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Ryan, Ellis Louise. "Investigating the role of TAB182 in the DNA damage response and replication stress pathways." Thesis, University of Birmingham, 2016. http://etheses.bham.ac.uk//id/eprint/6741/.
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It is well established that adenoviruses degrade components of the cellular DNA damage response, such as p53, DNA ligase IV and Mre11, in order to avoid detection from the host cell and thus, promote viral replication. Here we show TAB182, a protein of previously unknown function, is degraded following adenovirus serotype 5 and 12 infection. Similarly to other DNA damage proteins, together with the cellular Cullin 5 (during Ad5 infection) and Cullin 2 (during Ad12 infection). Interestingly, siRNA-mediated knockdown of TAB182 appears to be beneficial for adenovirus infection, as denoted by an increased expression of the adenoviral E1A protein and Cyclin E during adenovirus infection. Together with other studies, we confirm that TAB182 interacts with the large, multi-subunit CNOT complex. This complex has no defined function in mammalian cells, but is known to play a role in gene regulation in yeast. Interestingly, components of the CNOT complex are also degraded during adenovirus infection, whether adenovirus degrades TAB182 as well as CNOT for the same advantage is currently unknown. Cells deficient in TAB182 are hypersensitive to agents that induce DNA replication stress and also exhibit abnormal replication dynamics following release from hydroxyurea-induced fork stalling. In particular, they display increased fork restart and elevated new origin firing following release from hydroxyurea treatment, suggesting that TAB182 prevents fork recovery and suppresses new origin firing following replication stress. Depletion of some components of the CNOT complex is able to rescue the phenotypes observed in TAB182 deficient cells, suggesting that TAB182 and the CNOT complex may act in concert at the replication fork. TAB182 deficient cells display less DNA gaps and breaks but increased levels of 53BP1 bodies in G1 and micronuclei, which are markers of genome instability, following replication stress. Whether TAB182 acts directly at the replication fork, or in conjunction with other proteins known to be involved in replication restart such as helicases, nucleases, or chromatin remodelling complexes, remains to be elucidated.
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Weatherbee, Jessica L. "Exploiting DNA Repair and ER Stress Response Pathways to Induce Apoptosis in Glioblastoma Multiforme: A Dissertation." eScholarship@UMMS, 2016. https://escholarship.umassmed.edu/gsbs_diss/865.
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Glioblastoma multiforme (GBM) is a deadly grade IV brain tumor characterized by a heterogeneous population of cells that are drug resistant, aggressive, and infiltrative. The current standard of care, which has not changed in over a decade, only provides GBM patients with 12-14 months survival post diagnosis. We asked if the addition of a novel endoplasmic reticulum (ER) stress inducing agent, JLK1486, to the standard chemotherapy, temozolomide (TMZ), which induces DNA double strand breaks (DSBs), would enhance TMZ’s efficacy. Because GBMs rely on the ER to mitigate their hypoxic environment and DNA repair to fix TMZ induced DSBs, we reasoned that DSBs occurring during heightened ER stress would be deleterious.
Treatment of GBM cells with TMZ+JLK1486 decreased cell viability and increased cell death due to apoptosis. We found that TMZ+JLK1486 prolonged ER stress induction, as indicated by elevated ER stress marker BiP, ATF4, and CHOP, while sustaining activation of the DNA damage response pathway. This combination produced unresolved DNA DSBs due to RAD51 reduction, a key DNA repair factor. The combination of TMZ+JLK1486 is a potential novel therapeutic combination and suggests an inverse relationship between ER stress and DNA repair pathways.
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Melgar, Katelyn M. "A polypharmacologic strategy for overcoming adaptive therapy resistance in AML by targeting immune stress response pathways." University of Cincinnati / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1571061798761171.
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Weatherbee, Jessica L. "Exploiting DNA Repair and ER Stress Response Pathways to Induce Apoptosis in Glioblastoma Multiforme: A Dissertation." eScholarship@UMMS, 2008. http://escholarship.umassmed.edu/gsbs_diss/865.
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Glioblastoma multiforme (GBM) is a deadly grade IV brain tumor characterized by a heterogeneous population of cells that are drug resistant, aggressive, and infiltrative. The current standard of care, which has not changed in over a decade, only provides GBM patients with 12-14 months survival post diagnosis. We asked if the addition of a novel endoplasmic reticulum (ER) stress inducing agent, JLK1486, to the standard chemotherapy, temozolomide (TMZ), which induces DNA double strand breaks (DSBs), would enhance TMZ’s efficacy. Because GBMs rely on the ER to mitigate their hypoxic environment and DNA repair to fix TMZ induced DSBs, we reasoned that DSBs occurring during heightened ER stress would be deleterious.
Treatment of GBM cells with TMZ+JLK1486 decreased cell viability and increased cell death due to apoptosis. We found that TMZ+JLK1486 prolonged ER stress induction, as indicated by elevated ER stress marker BiP, ATF4, and CHOP, while sustaining activation of the DNA damage response pathway. This combination produced unresolved DNA DSBs due to RAD51 reduction, a key DNA repair factor. The combination of TMZ+JLK1486 is a potential novel therapeutic combination and suggests an inverse relationship between ER stress and DNA repair pathways.
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Diedrichs, Danilo Roberto. "A mathematical model of the unfolded protein response to stress in the endoplasmic reticulum of mammalian cells." Diss., University of Iowa, 2012. https://ir.uiowa.edu/etd/3284.
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The unfolded protein response (UPR) is a cellular mechanism whose primary functions are to sense perturbations in the protein-folding capacity of the endoplasmic reticulum and to take corrective steps to restore homeostasis. Although the UPR is conserved across all eukaryotic cells, it is considerably more complex in mammalian cells, due to the presence of three interconnected pathways triggered by separate sensor proteins, a translation attenuation mechanism, and a negative feedback loop. The mechanisms of these interacting biochemical pathways in the mammalian UPR allow for a better fine-tuning of the response than in the case of lower eukaryotes, such as yeasts.
The present thesis develops a quantitative mathematical model for the dynamics of the UPR in mammalian cells, which incorporates all the proteins and interactions between them that are known to play a role in this response. This model can be used to provide quantitative information about the levels of its components throughout the response, and to analyze the ramifications of perturbations of the UPR. The model uses a system of ordinary nonlinear differential equations based on biochemical rate equations to describe the dynamics of the UPR as a network of interacting proteins and mRNAs. An early model is presented as a first attempt to investigate the UPR network and construct an inclusive wiring diagram, as well as suggesting a framework to model the differential equations. Then, a refined, quantitative model is designed based on experimental data collected on Mouse Embryonic Fibroblasts treated with Thapsigargin to induce stress and trigger the UPR. The model defines the differential equations and determines the unknown kinetic parameters by optimizing the fit of the system's solution to the experimental data. It includes the UPR's intrinsic feedback loops and allows for the integration of various forms of external stress signals. To the best of our knowledge, it is the first, data-validated, quantitative model in the literature for the UPR in mammalian cells.
The last chapters of the thesis address, from a modeling point of view, two important questions for the UPR: (1) cell survival versus apoptosis; and (2) incompleteness of the biological wiring diagram. Recent experimental results show that the UPR is capable of producing qualitatively different results leading to cell survival or death depending on the nature, strength, and persistence of the inducing stress. This thesis proposes several approaches by which the equations can be modified to model the transition from adaptation to apoptosis as a dynamic switch, while taking into account the various hypotheses of cell death mechanisms. Finally, we use recently-developed computational algebra techniques to infer an optimal structure of the UPR network, based solely on the experimental data; the resulting wiring diagram provides insights on elements of the structure of the model that may have been overlooked during the classical (mechanistic) approach to our original data-based model.
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Viana, Tiago Monteiro Lomba. "In search of role of yeast regulatory pathways during the last stages of wine fermentation." Doctoral thesis, ISA/UL, 2014. http://hdl.handle.net/10400.5/7321.
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Doutoramento em Engenharia Alimentar - Instituto Superior de Agronomia<br>While searching for roles of Saccharomyces cerevisiae regulatory pathways during the late stages of wine fermentation, a peculiar H+ homeostasis was found in an industrial wine strain, while fermenting natural grape must (NGM). Intracellular pH (pHi) of the population remained in the range 6.0 – 6.4, decreasing by the end of glucose fermentation (pHi 5.2 at 15oC), although cells remained viable. The plasma membranes of late stationary cells became almost impermeable to H+. To ensure replicability of subsequent experiments, a synthetic grape must was designed (ISA-SGM), in which we obtained similar fermentative profiles as in NGM for the wine strain and for laboratory strains. Fluorescence Ratio Imaging Microscopy (FRIM) was used to estimate pHi of individual cells of strain BY4741. Even at the end of fermentation, we found subpopulations with “healthy” cytosolic pH (i.e. pHi 6-7). Sixty-two single-deletion mutants of BY4741 were screened to identify genes required for optimal wine fermentation performance, leading to the identification of 10 determinants whose absence improved fermentation performance, while 37 were deleterious. Principal Component Analysis revealed that deletion of some genes involved in transcription, cell cycle and stress response led to improved performance, while reduced performance was associated mainly with genes involved in membrane functions.
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Gomez, Vargas Javier Alejandro. "Regulation of the signal transduction pathways of the unfolded protein response during chronic and physiological ER stresses." Diss., University of Iowa, 2016. https://ir.uiowa.edu/etd/5760.
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The unfolded protein response (UPR) is activated by protein misfolding stress in the endoplasmic reticulum (ER). The UPR is a transcriptional program that aims to maintain ER folding capacity, where imbalances between protein load and processing ability is termed ER stress. Signal transduction of the UPR begins with 3 ER-resident transmembrane sensors: PERK, IRE1 and ATF6. All sensors initiate downstream signaling cascades which culminate in improved protein folding, transcriptional upregulation of genes encoding ER chaperones, and mechanisms to reduce translational and transcriptional ER load, therefore re-establishing ER homeostasis. The signaling cascades of each sensor are distinct but cooperative, and involve a significant amount of crosstalk, feedback and overlap. Indeed, there are many pathological and physiological conditions have an effect on ER protein burden, and therefore on activation of the UPR. Increases in protein load in professional secretory cells, hypoxic conditions in a tumor mass, obesity all induce cause changes in the ER folding environment.
Although we understand how the UPR contributes to relieve ER stress under acute conditions (e.g. pharmacological treatment) much less is understood about the contributions to physiological processes and chronic stress conditions. Our overall goal was to understand how the UPR is activated during physiological settings, the mechanisms it uses to maintain folding capacity under these setting and the specific components responsible for adapting the response to various stresses.
We first decided to understand a chronic stress from a transgenic approach. By creating a knockout mouse, the genetic deletion functions as a stress and we can understand its physiological role. By compounding two genetic deletions in UPR components (ATF6α and p58IPK) we provide evidence for the developmental role these components play. Homozygous deletion ATF6α bears no gross histological phenotype yet causes synthetic lethality when combined with p58IPK deletion. This also reveals that the UPR is able to adapt to genetic impairment of protein folding in vivo.
Next, to better understand these chronic states, we established an experimentally tractable chronic stress treatment in vivo. Our treatment suppressed ATF6α dependent chaperone expression through an mRNA degradative mechanism, which led to long term changes in UPR expression. We determined that chronic conditions can change the sensitivity of the UPR to ER stress, potentially as an adaptive consequence. We also showed that sensitivity to ER stress can be changed during chronic stress.
Finally we simulated the UPR in a computational ordinary differential equation (ODE) model in order to determine how various stresses and component interactions determine the output of the UPR. We built a series of equations to describe the UPR signaling network, entrained it on experimental data and refined it through the use of transgenic knockout cells. Our model was robust enough to recreate experimental measurements of UPR components when tested in parallel with knockout cells. We found that stress sensitivity is dependent on the crosstalk and negative feedback connections of the UPR.
This study has enhanced our understanding of activation of the UPR under non-acute settings. It demonstrates that the UPR is a signaling hub with a broad output range that is capable of handling a variable degree of insults because of the intrinsic properties of the signaling network. This provides a better understanding for the contributions of the UPR to physiological stresses and certain chronic diseases.
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Platara, Maria. "Signaling pathways regulating the transcriptional response of the sodium ATPase ENA1 to saline and alkaline stress in the yeast Saccharomyces cerevisiae." Doctoral thesis, Universitat Autònoma de Barcelona, 2008. http://hdl.handle.net/10803/3586.
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La respuesta de adaptación de la levadura Saccharomyces cerevisiae a la alcalinización ambiental provoca una remodelación de su expresión génica. Una diana clave es el gen ENA1 que codifica una ATPasa de sodio, y cuya inducción por pH alcalino está mediada por las vías de calcineurina y el Rim101.<br/>En un estudio previo se identificaron dos regiones del promotor de ENA1 responsables de su respuesta al álcali, la ARR1 (Alkaline Responsive Region 1) que es calcineurina-dependiente y ARR2 que es calcineurina-independiente. En este trabajo restringimos la región responsable de la respuesta alcalina de ARR2 a un pequeño fragmento de 42 nucleótidos que denominamos MCIR (por Minimum Calcineurin Independent Response). MCIR contiene un elemento MIG, capaz de unir a los represores Mig1 y Mig2. Observamos que la respuesta a pH alcalino de la MCIR se anula en células que carecen de Snf1, la quinasa que regula la actividad represora de Mig1 en función de la disponibilidad de glucosa. En cambio, su respuesta se ve moderadamente reducida en cepas rim101, mientras que el doble mutante mig1 mig2 presenta altos niveles de expresión a pH alcalino. Además, la deleción de NRG1 resulta en una expresión elevada y la inducción de MCIR es marginal en el cuádruple mutante nrg1,2 mig1,2. También demostramos que Nrg1 se une al extremo 5' de la ARR2 in vitro e in vivo. Por lo tanto, la respuesta de ENA1 que es calcineurina independiente esta regulada por Rim101 (a través de Nrg1) y por Snf1 (a través de Nrg1 y Mig2). De esta manera, la inducción del promotor de ENA1 por pH alcalino en un mutante rim101snf1 en presencia del inhibidor químico de la calcineurina FK506 se anula totalmente. Por lo tanto, la respuesta transcripcional de ENA1 a estrés alcalino, integra tres vías de señalización, cuya importancia relativa es Snf1 > calcineurina > Rim101.<br/>La CK2 es una quinasa que está conservada en eucariotas y participa en diversos procesos celulares. En S. cerevisiae cepas que carecen de las subunidades reguladoras Ckb1 y/o Ckb2 de la CK2 son muy sensibles a cationes de litio y de sodio. <br/>En este estudio confirmamos observaciones anteriores que describían que la respuesta de ENA1 a estrés salino y alcalino está disminuida en células que carecen de Ckb1 y/o Ckb2. Además demostramos que los mutantes ckb son sensibles a pH alcalino. Las tres vías de señalización (Rim101, calcineurina, Snf1) responsables de la regulación de ENA1 en condiciones de estrés alcalino se examinaron para posibles interacciones con la CK2. Nuestros resultados sugieren que CK2 y calcineurina regulan la expresión de ENA1 de manera independiente. Además, mostramos que la deleción de RIM101 resulta en inducción de la expresión de ENA1, disminuida en condiciones de estrés salino, y que la deleción simultanea de CKB agrava solo ligeramente el defecto de las cepas rim101 en la expresión salina y alcalina de ENA1. Deleción del factor de transcripción Nrg1 en un fondo genético ckb resulta en niveles de expresión de ENA1 relativamente altos en condiciones de estrés salino y alcalino. Estos resultados, junto con datos anteriores que muestran que Nrg1 se fosforila por CK2 en estas condiciones de estrés, son compatibles con una supuesta interacción entre CK2 y la vía Rim101. Cabe destacar que la deleción de CKB repara el defecto que presentan las células snf1 en la expresión de ENA1 bajo estrés salino y alcalino y que los mutantes ckb1,2 snf1 presentan un crecimiento en litio mayor que la cepa salvaje, sugiriendo la existencia de una interacción compleja entre CK2 y Snf1.<br>The adaptive response of the yeast Saccharomyces cerevisiae to environmental alkalinization results in remodeling of gene expression. A key target is the gene ENA1, encoding a sodium ATPase, whose induction by alkaline pH was shown to integrate at least two different signals, mediated by the calcineurin and the Rim101 pathways.<br/>Early work in our laboratory identified two regions in the ENA1 promoter required for full response to alkalinization, the ARR1 (from Alkaline Responsive Region 1), whose response is calcineurin-dependent and ARR2, whose response is calcineurin-independent. In this work we have restricted the alkaline response of ARR2 to a smaller fragment of 42 nucleotides that we denominated MCIR (from Minimum Calcineurin Independent Response). MCIR contains a MIG element, able to bind Mig1 and Mig2 repressors. We observe that high pH-induced response driven from MCIR is largely abolished in cells lacking Snf1, the protein kinase that regulates repressor activity of Mig1 with respect to glucose availability, and results moderately reduced in a rim101 strain, whereas the double mig1 mig2 mutant presents high levels of expression upon alkaline stress. In addition, deletion of NRG1 results in increased expression and induction from the MCIR region is marginal in a quadruple nrg1,2 mig1,2 mutant. We also demonstrate that Nrg1 binds to the 5´-end of the ARR2 region in vitro and in vivo. Therefore, the calcineurin-independent response of the ENA1 gene is under the regulation of Rim101 (through Nrg1) and Snf1 (through Nrg1 and Mig2). Accordingly, induction by alkaline stress of the entire ENA1 promoter in a snf1 rim101 mutant in the presence of the calcineurin inhibitor FK506 is completely abolished. Thus, the transcriptional response to alkaline stress of the ENA1 gene integrates three different signaling pathways, whose relative potency is Snf1 > calcineurin > Rim101. <br/>CK2 is a well conserved kinase among eukaryotes that participates in many different cellular processes. In Saccharomyces cerevisiae, strains lacking the regulatory subunits Ckb1 and/or Ckb2 of this kinase are hypersensitive to sodium and lithium cations. However, the mechanism by which CK2 affects yeast salt tolerance is not known. <br/>In this study we confirm previous observations that the alkaline and saline response of ENA1 is decreased in strains lacking Ckb1 and/or Ckb2. Furthermore, we show that ckb mutants are sensible at alkaline pH. The three pathways (Rim101, calcineurin and Snf1) responsible for ENA1 regulation under alkaline stress conditions were examined for any possible interaction with CK2. Our results suggest that CK2 and calcineurin regulate ENA1 expression under alkaline and lithium stress in an independent fashion. Moreover, we show that deletion of RIM101 results in decreased ENA1 induction under lithium stress conditions and that simultaneous mutation of CKB only slightly aggravates the defect that presents the rim101 strain in ENA1 alkaline and saline expression. Mutation of the Nrg1 transcription factor in a ckb background leads to relatively high levels of ENA1 expression under alkaline and lithium stress conditions. These results, together with previous data showing that Nrg1 is phosphorylated by CK2 under these stress conditions, support a possible interaction between the CK2 and the Rim101 pathways. Remarkably, deletion of CKB partially counteracts the defect that snf1 mutants present in ENA1 saline and alkaline expression, and ckb1,2 snf1 mutants are as tolerant as wild type cells to lithium ions, revealing a complex interaction between CK2 and Snf1.
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Xing, Qikun. "Deciphering the oxylipin signaling pathways during defense responses in brown macroalgae." Thesis, Sorbonne université, 2021. http://www.theses.fr/2021SORUS536.
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Les oxylipines sont des composés oxygénés dérivés d'acides gras polyinsaturés (AGPI), présents chez de nombreux organismes. Chez les plantes et les animaux, ils ont des rôles signaux dans la croissance et en réponse à divers stress. Les algues brunes ont développé des voies de biosynthèse des oxylipines uniques, à partir des AGPI en C18 et C20. Cependant, le rôle et la régulation de ces voies y sont encore peu connus. Au cours de ma thèse, j'ai mené des analyses transcriptomiques à large échelle chez deux espèces d’algues brunes, Saccharina latissima et Laminaria digitata, lors d'interactions biotiques, en relation avec l'induction des voies des oxylipines. Ces analyses ont été conduites sur des algues cocultivées avec des endophytes et en réponse à l’élicitation par les oligoguluronates. Plusieurs gènes liés à la biosynthèse des oxylipines montrent des régulations différentes. Après l’élicitation, un gène codant potentiellement pour un cytochrome P450 (CYP) est surexprimé à 1 h et 12 h, suggérant une induction séquentielle des voies des oxylipines chez S. latissima. Des profilages métaboliques montrent l’accumulation précoces d’oxylipines issues de la voie des AGPI en C18 et suggèrent le rôle alternatif de composés dérivés des AGPI en C20. Enfin, le rôle biologique de plusieurs oxylipines et aldéhydes a été étudié sur la régulation de certains gènes cibles ou l'accumulation de composés putatifs de défense chez S. latissima et L. digitata. Les résultats de ces travaux ont permis de reconstruire partiellement les voies des oxylipines et de mieux comprendre leur régulation lors des réponses de défense chez les algues brunes<br>Oxylipins are oxygenated compounds derived from polyunsaturated fatty acids (PUFAs), found in many organisms. In land plants and animals, they are known to have regulating roles in growth and in response to various stresses. Brown algae have developed unique oxylipin pathways, using both C18- and C20- type PUFAs. However, the role and regulation of oxylipin pathways during defense responses are largely unclear in brown algae. In my PhD thesis, I conducted large transcriptomic analyses in order to decipher the molecular responses of two kelps species, Saccharina latissima and Laminaria digitata during biotic interactions, in relationships with the induction of oxylipin pathways. The transcriptomic analysis was performed on a kelp-endophyte co-cultivation bioassay and oligoguluronate elicitation. Among differentially expressed genes, several genes putatively involved in oxylipin pathways were identified with diverse expression patterns. During the elicitation, one putative cytochrome P450 (CYP)-encoding gene was induced at 1 h and 12 h, suggesting that oxylipin pathways in S. latissima might be induced at least two times. The result of metabolite profiling showed an early production of putative C18 oxylipins and the putative alternative role of C20 oxylipin pathways during the elicitation. Finally, the biological function of several oxylipins and aldehydes were tested on targeted gene or metabolite inductions in S. latissima and L. digitata. Altogether, these results led to the partial reconstruction of oxylipin pathways and provide a better understanding of their regulation during defense responses in kelps
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Packer, John. "Identification of signalling pathways involved in the oxidative stress response triggered by Low Temperature Plasma in prostate epithelial cells and the assessment of tumour-associated allelic expression in prostate cancer." Thesis, University of York, 2018. http://etheses.whiterose.ac.uk/22237/.
Full textAbstract:
Pairing of cancer genome and transcriptome data has revealed that heterozygous mutations aren’t always expressed in cells. The potential for point mutation or genomic rearrangement to alter tumour allelic expression has implications for understanding cellular heterogeneity and application of treatments. Mutation of SPOP, PTEN and IDH-1 was assessed in 51 primary prostate cancer cultures to establish allelic heterozygosity and ascertain whether oncogenic change to coding regions altered allelic expression. No mutations were detected in the three genes, although 18% of tested cultures had loss of heterozygosity in PTEN. The TMPRSS2-ERG fusion, present in half of all prostate cancers, is selectively expressed at an allelic level by cancer stem cells. Monoallelic expression didn’t correlate with TMPRSS2 promoter hypermethylation. Prostate cultures expressed fusion transcript, however epigenetic features of monoallelically expressed genes were not investigated in the epithelial subpopulations. Understanding of allelic chromatin states may inform treatment strategies that permit tumour suppressor expression or oncogenic protein repression. Inability to predict indolent or aggressive progression of organ-confined prostate cancers has created the problem of surgical overtreatment. Focal therapies targeting the tumour core are being met with increasing rates of recurrence, necessitating development of novel treatments. The anti-cancer properties of Low Temperature Plasma (LTP) are being explored in prostate models where it produces autophagy and necrosis through generation of reactive species. Initial gene expression response to LTP and the activation of upstream transcription factors were analysed. LTP activated Nrf2, AP-1 and Notch signalling in patient matched prostate normal and cancer cultures. The progenitor-containing cell fraction was more responsive to LTP than differentiated epithelial cells in both transcription of response genes and nuclear accumulation of active Notch1. When linked to cell-fate outcomes, these immediate molecular responses of prostate cancer to LTP could be used as hallmarks of resistance or treatment efficacy in patients.
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Koren, John. "The Role of Hsp70 in Cancer: A Study of the Hsp70 / Akt Relationship." Scholar Commons, 2012. http://scholarcommons.usf.edu/etd/4105.
Full textAbstract:
The Hsp70 family of molecular chaperones is essential for
protein folding, re-folding misfolded client proteins, clearance
of aberrant client proteins, and can also inhibit programmed
cell death. There are two major cytosolic members of this
family: the constitutive Hsc70, and the inducible Hsp72. Under
stress conditions the Hsp70 family protects the cell from
protein related damage by the induction of Hsp72. Hsc70 and
Hsp72 are highly homologous with minor differences in
substrate binding. In cancers, Hsp72 is commonly induced and
this induction is thought to aid in cancer cell survival. In these
studies we demonstrate the differential regulation of the prosurvival
kinase Akt by Hsc70 and Hsp72. We demonstrate that
of the two cytosolic forms, Hsp72 is the primary Akt regulator.
Using a phenothiazine class inhibitor of Hsp70-family activity,
methylene blue, we demonstrate dose dependent decreases in
the levels of Akt; produced breast cancer specific cell death.
This cell death could be rescued by the use of an Hsp70 family ATPase stimulating compound, SW02. We also demonstrate a
similar phenotype with a rhodacyanine class Hsp70 family
inhibitor, YM-1, also capable of reducing Akt and causing
cancer specific cytotoxicity. The resulting Akt decreases were
sufficient to block a tamoxifen-resistance pathway, allowing
previously resistant cells to regain sensitivity to tamoxifen.
These results demonstrate the capabilities of Hsp70 family
inhibitors as potent compounds for the treatment of breast
cancer.
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Naim, Adnan. "The Role of G3BPs in the Stress Response Pathway." Thesis, Griffith University, 2016. http://hdl.handle.net/10072/367499.
Full textAbstract:
The ras-GTPase SH3-domain Binding Proteins (G3BP) are a family of RNA-binding proteins that have been implicated in multiple cellular activities ranging from signal transduction to regulation of messenger RNA (mRNA). G3BPs were named after their interaction with the SH3 domain of Ras-GTPase-activating protein; however recent research did not find this interaction. All three members of the G3BPs family, G3BP1, G3BP2a and G3BP2b, share structural similarities with each other by having four distinct regions (1) the Nuclear Transporting Factor 2, (NTF2) domain at the N-terminal, (2) the acidic and proline-rich domain in the centre, (3) the RNA recognition motif (RRM) and (4) the arginine glycine (RGG)-rich region rich at the C-terminal. The presence of the NTF2 domain in its structure suggests G3BP might play a role in nucleocytoplasmic transportation, which was observed after serum stimulation where G3BP1 was translocated to the nucleus from the cytoplasm. The RNA recognition motif (RRM) region plays a vital role in its interaction with the target RNA. The RGG-rich box is a region rich in arginine and glycine residues, which plays a role assisting RRM in interactions with protein or RNA.
G3BP1 is found to be overexpressed in many cancers, including breast cancer, and head and neck tumours, as well as cell lines derived from human lung, prostrate, colon, thyroid and breast cancer. G3BPs have also been implicated in translational control within differentiating neurons, suggesting that G3BP may play several roles in controlling the translational fate of its cargo and that its role may be cell-specific. G3BP1 has also been found in β-integrin- induced adhesion complexes. This information highlights G3BPs as a dynamic protein that is involved in several biological functions.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>School of Natural Sciences<br>Science, Environment, Engineering and Technology<br>Full Text
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Nalam, Vamsi J. "9-Lipoxygenase Oxylipin Pathway in Plant Response to Biotic Stress." Thesis, University of North Texas, 2012. https://digital.library.unt.edu/ark:/67531/metadc115127/.
Full textAbstract:
The activity of plant 9-lipoxygenases (LOXs) influences the outcome of Arabidopsis thaliana interaction with pathogen and insects. Evidence provided here indicates that in Arabidopsis, 9-LOXs facilitate infestation by Myzus persicae, commonly known as the green peach aphid (GPA), a sap-sucking insect, and infection by the fungal pathogen Fusarium graminearum. in comparison to the wild-type plant, lox5 mutants, which are deficient in a 9-lipoxygenase, GPA population was smaller and the insect spent less time feeding from sieve elements and xylem, thus resulting in reduced water content and fecundity of GPA. LOX5 expression is induced rapidly in roots of GPA-infested plants. This increase in LOX5 expression is paralleled by an increase in LOX5-synthesized oxylipins in the root and petiole exudates of GPA-infested plants. Micrografting experiments demonstrated that GPA population size was smaller on plants in which the roots were of the lox5 mutant genotype. Exogenous treatment of lox5 mutant roots with 9-hydroxyoctadecanoic acid restored water content and population size of GPA on lox5 mutants. Together, these results suggest that LOX5 genotype in roots is critical for facilitating insect infestation of Arabidopsis. in Arabidopsis, 9-LOX function is also required for facilitating infection by F. graminearum, which is a leading cause of Fusarium head blight (FHB) disease in wheat and other small grain crops. Loss of LOX1 and LOX5 function resulted in enhanced resistance to F. graminearum infection. Similarly in wheat, RNA interference mediated silencing of the 9-LOX homolog TaLpx1, resulted in enhanced resistance to F. graminearum. Experiments in Arabidopsis indicate that 9-LOXs promote susceptibility to this fungus by suppressing the activation of salicylic acid-mediated defense responses that are important for basal resistance to this fungus. the lox1 and lox5 mutants were also compromised for systemic acquired resistance (SAR), an inducible defense mechanism that is systemically activated throughout a plant in response to a localized infection. the lox1 and lox5 mutants exhibited reduced cell death and delayed hypersensitive response when challenged with an avirulent strain of the bacterial pathogen Pseudomonas syringae pv tomato. LOX1 and LOX5 functions were further required for the synthesis as well as perception of a SAR-inducing activity present in petiole exudates collected from wild-type avirulent pathogen-challenged leaves. Taken together, results presented here demonstrate that 9-LOX contribute to host susceptibility as well as defense against different biotic stressors.
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Gomes, Susana Isabel Lopes. "Effect assessment of nanoparticles toxicity in the terrestrial compartment." Doctoral thesis, Universidade de Aveiro, 2013. http://hdl.handle.net/10773/12578.
Full textAbstract:
Doutoramento em Biologia<br>Over 11 million tons of nanomaterials (NMs) have been produced in 2012 and predictions point the increase in production. Despite predictions and extended usage via consumer products and industry, the understanding of the potential impact of these materials on the environment is virtually absent. The main aim of this thesis is to understand how a selected group of nanomaterials (metal based particles) may impact soil invertebrates, with special focus on the
mechanisms of response. Since a case-by-case Environmental Risk Assessment (ERA) of all the emerging contaminants (particularly NMs) is impossible, among others due to time and cost reasons, to gain understanding on the mechanism of action and response is very important to reach a common paradigm. Understanding the modes of action provides predictive characters in cross particle extrapolation. Besides, it also provides insight for the production of new and sustainable materials.
Overall, the effects of the selected NMs (Copper and Silver, Titanium and Zirconium oxides) and the respective salt forms, were investigated at the gene
expression (using high-throughput tools, microarray and qPCR technology), biochemical (using enzymatic assays for analysis of oxidative stress markers)
and organism (survival and reproduction as in OECD test guidelines) levels, this using standard soil species (Enchytraeus albidus, Enchytraeus crypticus,
Eisenia fetida).
Gene expression analysis provided valuable information on the mechanisms affected by each of the NMs. The gene expression profile highlighted a
(nano)material signature and the effect of the duration of exposure. The functional analyses integrated with the biochemical and organism data,
revealed a good understanding power. The biochemical parameters (oxidative stress related) were distinct across the materials and also influenced by
duration of exposure and concentration. The standardized organismal responses differed the least between the various materials.
The overall outcome is that, in this context of NMs effect assessment, gene expression and enzymatic assays introduced a very important knowledge gap,
which could not had been achieved by the standard organismal effects alone.
A reoccurring issue with some metal based NMs is the possible dissolution and subsequent release of ions that then causes toxicity e.g. Cu-NPs or Ag-NPs
release Cu2+ or Ag+. The oxidation state of the particles was investigated, although this was not the focus of the thesis. The study of fate, e.g. dissolution
of NPs, is also only in its beginning and the appropriate techniques are currently being developed. The results showed a specific nanoparticle effect.
The UV exposure with titanium dioxide nanoparticles increased its effect.<br>Em 2012 foram produzidas mais de 11 milhões de toneladas de nanomateriais (NMs) e as perspetivas apontam para um aumento na produção. Apesar das previsões e o uso extensivo em produtos de consumo e indústria, o conhecimento é praticamente inexistente no que diz respeito ao potencial
impacto destes materiais no ambiente. O principal objetivo desta tese é compreender o impacto de um grupo de NMs selecionados (NMs de base metálica) em invertebrados de solo, com especial incidência nos mecanismos de resposta. Uma vez que a avaliação de risco ambiental feita caso-a-caso para todos os contaminantes emergentes (particularmente NMs) é impossível, devido, entre outros fatores, ao tempo e custos necessário, a compreensão dos mecanismos de ação é muito importante para alcançar paradigmas comuns. A compreensão dos modos de ação fornece os caracteres com valor preditivo para a extrapolação entre partículas. Além disso, também fornece informação para a produção de novos materiais sustentáveis.
Em suma, os efeitos dos NMs selecionados (Cobre e Prata, Óxido de Titânio e Zircónio) e do respetivo sal, foram investigados ao nível dos genes (utilizando a ferramentas de alto varrimento, tecnologia de “microarrays” e PCR em tempo real), bioquímico (utilizando ensaios enzimáticos para a análise de marcadores de stress oxidativo) e do organismo (sobrevivência e reprodução, tal como nos protocolos OCDE), utilizando espécies modelo ecotoxicológicas (Enchytraeus albidus, Enchytraeus crypticus, Eisenia fetida).
A análise da expressão de genes forneceu informação importante sobre os mecanismos afetados por cada um dos NMs testados. Os perfis de expressão genéticos evidenciaram uma assinatura do (nano)material e o efeito do tempo de exposição. A análise funcional integrada com os dados bioquímicos e de organismo revelou um bom poder de entendimento. As respostas dos parâmetros bioquímicos (relacionados com stress oxidativo) foram distintas entre os materiais testados e também influenciados pelo tempo de exposição e concentrações testadas. As respostas padronizadas ao nível do organismo
foram as que mostraram menor diferenciação entre os vários materiais testados.
De um modo geral, e neste contexto de avaliação de efeitos de NMs, a expressão de genes e ensaios enzimáticos, apresentaram um papel muito importante no preenchimento de lacunas que não podería ter sido alcançado através dos efeitos no organismo isoladamente.
Um assunto recorrente relativo a alguns NMs de base metálica tem a ver com a possível dissolução e subsequente libertação de iões que a posteriori causam toxicidade, p.e. Cu-NPs ou Ag-NPs libertam Cu2+ ou Ag+. O estado de oxidação das partículas foi investigado, apesar deste não ser o foco da tese. O
estudo do destino, p.e. dissolução de NPs, está ainda apenas no seu início e as técnicas apropriadas estão presentemente a ser desenvolvidas. Os resultados mostraram um efeito específico das nanopartículas. A exposição UV com o dióxido de titânio aumentou o seu efeito.
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Johnson, Shane Benjamin. "An analysis of prefrontal cortex pathways and their assembly of stress coping responses." Diss., University of Iowa, 2019. https://ir.uiowa.edu/etd/6965.
Full textAbstract:
Stress is characterized by the deployment of response systems to promote adaptation in the face of threats. Among these, the neuroendocrine hypothalamic-pituitary-adrenal (HPA) axis has received considerable attention due to the potent acute and chronic effects of its glucocorticoid end-products, including cortisol in humans and corticosterone (CORT) in rodents. Stress also simultaneously elicits conserved behavioral responses that may be key to understanding how animals and humans cope with ongoing threats. Both neuroendocrine and behavioral responses to psychological stress are thought to originate from, and are modulated by, complex neurocircuitry residing within the limbic forebrain. However, to date these responses have largely been studied in functional and neuroanatomical isolation. The experiments here described are intended to shed light on the circuitry underlying the dual modulation of behavioral and HPA output.
Chapter 2 investigates a pathway from the prelimbic subfield (PL) of the medial prefrontal cortex (mPFC) to the anteroventral bed nuclei of the stria terminalis (avBST). Using an optogenetic approach, we found that this pathway simultaneously suppresses both immobility behavior and HPA output during an acute psychological stressor (tail suspension, TS). We go on to show that this pathway also suppresses behavioral passivity in the shock probe defensive burying test (SPDB), a test of coping behavior. Furthermore, endogenous activity in this pathway, as measured by Fos immunoreactivity in avBST–projecting PL neurons, was negatively correlated with passive coping behavior in the SPDB. Follow-up experiments found that PL axonal terminals within avBST were glutamatergic and photoexcitation of these terminals produced excitatory post-synaptic potentials in avBST neurons. Next, a downstream pathway from avBST to the ventrolateral periaqueductal gray (vlPAG) was investigated as a candidate mediator of the observed effects on passive coping. Photoinhibition of avBST terminals in vlPAG recapitulated the effects of PL–avBST photoinhibition. Finally, avBST terminals within vlPAG were found to be GABAergic, consistent with a role for avBST inputs in inhibiting passive coping-related activity in this region.
Chapter 3 expands on the role of avBST and its output pathways in modulating behavioral and neuroendocrine stress responses. Photoinhibition of avBST cell bodies during TS produced a marked increase in both immobility and HPA output while photoexcitation was sufficient to suppress the neuroendocrine stress axis. Follow-up studies found that the HPA-modulatory effects of avBST cell body manipulations were likely mediated by direct avBST inputs to the paraventricular nucleus of the hypothalamus (PVH). We found that avBST terminals within PVH terminated in close proximity to putatively neurosecretory corticotropin releasing factor (CRF)-immunoreactive neurons. Photoinhibition of avBST terminals in PVH during TS produced elevations in HPA output that were comparable to those observed follow avBST cell body inhibition. Finally, photoinhibition of avBST terminals in vlPAG was associated with increased immobility during both TS and acute exposure to the forced swim test, consistent with a role for this pathway in suppressing passive behaviors across a variety of behavioral tests.
Chapter 4 studies parallel pathways from mPFC to distinct cell columns within the periaqueductal gray (PAG). The PAG is a highly conserved region of the midbrain that surrounds the cerebral aqueduct and has been implicated in the regulation of defensive behaviors. Prior work suggests that ventrolateral aspects of the structure promote passive defensive behaviors (e.g., freezing and immobility), whereas activation of the dorsal (d) cell column produces active behavior (threat confrontation or flight). In these experiments, we again utilized the SPDB; rats are exposed to an electrified probe mounted on their cage wall, whereby after receiving electric shock, they display both active (probe burying with cage bedding) and passive (immobility) coping behavior. Consistent with previous reports, we found that rostral mPFC provided dense innervation of ventrolateral PAG, whereas caudal mPFC provided innervation of dorsal PAG. Using an optogenetic approach we found that photoinhibition enhanced, and photoexcitation of the rostral mPFC–ventrolateral PAG pathway diminished passive coping during the SPDB, but active coping behavior was unaffected. Next, we investigated the contributions of the caudal mPFC–dorsal PAG pathway during the SPDB. Here, pathway photoexcitation enhanced probe burying behavior, the primary measure of active coping, while other behaviors remained unaffected. This result suggested that activation of a single pathway was sufficient to drive active coping. Finally, we tested the effects of caudal mPFC–dorsal PAG pathway photoexcitation under conditions where active coping behavior is prohibited, by removal of the cage bedding to prevent rats from the ability to bury the shock probe. In control animals acutely deprived of bedding during the SPDB, we observed increased immobility behavior and ultrasonic vocalizations, as well as autonomic and HPA output, while each of these were decreased in bedding-deprived animals that received caudal mPFC–dPAG pathway photoexcitation. This final series of experiments implicate separate prefrontal-PAG pathways in either the suppression of passive, or promotion of active coping behavior. They further suggest that the caudal mPFC-dorsal PAG pathway provides a neural basis linking active coping with stress-buffering effects, marked by decreases in displacement behavior and neuroendocrine activation.
These results show that separate pathways from the medial prefrontal cortex to the bed nuclei of the stria terminalis and periaqueductal gray are simultaneously and differentially able modulate passive and active coping in response to aversive stimuli in rats. The prefrontal–avBST pathway coordinates the inhibition of something akin to a “passive response set” – i.e., by gating passive coping behavior and restraining neuroendocrine activation. Complementary, parallel prefrontal–periaqueductal gray pathways are able to independently support either the suppression of passive, or promotion of active coping behavior. The discussion will consider the naturalistic contexts accounting for how activity in mPFC may provide for the cooperative engagement of an active behavioral response set, and how differentially engaging these pathways may promote distinct adaptative strategies as based upon changing environmental conditions. Finally, we will consider how these data offer a neural basis linking active coping with stress-buffering effects, and how perturbations in these circuits may lead to chronic stress-related dysfunction of multiple systems and inform disease susceptibility in humans.
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Morancho, Armisen Beatriz. "Regulation of NFAT5 by signaling pathways involved in osmotic stress responses and cell growth." Doctoral thesis, Universitat Pompeu Fabra, 2008. http://hdl.handle.net/10803/77906.
Full textAbstract:
NFAT5 is the main regulator of an osmoprotective gene program that is switched on by osmotic stress. Hypertonicity causes a harmful increase in the intracellular concentration of inorganic ions and NFAT5-regulated genes allow the accumulation of organic osmolytes and chaperones in order to protect the cells. However, little is known about the physiopathologic tonicity thresholds that trigger NFAT5 transcriptional activity, and which signaling pathways are involved, in primary cells.
We have studied the regulation of NFAT5 transcriptional activity in several types of primary cells obtained from transgenic mice carrying the 9xNFAT-Luc reporter developed by the Molkentin laboratory. Our results indicate that NFAT5 is able to respond to pathological tonicity levels in primary cells and it requires a combination of signaling mediators, some of which are more relevant in specific cell types.
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Al-Bader, Nadia. "The role of the trehalose biosynthesis pathway in «Aspergillus fumigatus» development and stress response." Thesis, McGill University, 2009. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=40760.
Full textAbstract:
Trehalose is a non-reducing dissacharide of glucose involved in the metabolism of bacteria, plants, insects and fungi that is not present in mammals. In fungi, trehalose serves as source of energy for reproduction and development, and it can protect cells from environmental stress and nutrient limitations. It has also been linked with virulence in pathogenic yeasts. Trehalose biosynthesis in fungi involves a protein complex consisting of trehalose-6-phosphate synthase (Tps), trehalose-6-phosphate phosphatase (Tpp), and regulatory proteins. In the pathogenic mold Aspergillus fumigatus, we located four putative trehalose-6-phosphate synthases: tpsA, tpsB, tpsC, and tpsD. We found that trehalose accumulated during the developmental life cycle of A. fumigatus, throughout which both tpsA and tpsB were significantly expressed. In stress, trehalose accumulated only during heat shock, and this accumulation correlated with an increase in expression of tpsA. We therefore created strains of A. fumigatus in which tpsA, tpsB, or both were disrupted. Only the ΔtpsAB double mutant failed to accumulate trehalose under any conditions. The ΔtpsAB mutant was also delayed in germination at normal growth temperature, suggesting a developmental defect. With heat stress, most ΔtpsAB spores were found to be non-viable, and those that were viable were severely delayed in germination, growth and subsequent sporulation. No clear correlation could be made between ΔtpsAB and susceptibility to oxidative stress; however ΔtpsAB may have an abnormal cell wall. Further studies in mice are underway to determine if the ΔtpsAB mutant is impaired in virulence, potentially making the trehalose synthase complex a promising new target for antifungal therapy.<br>Le tréhalose est un disaccharide de glucose qui fait partie du métabolisme d’organismes comme les bactéries, les plantes, les insectes et les champignons, mais il est absent chez les mammifères. Chez les champignons, le tréhalose sert de source d’énergie pour la reproduction et la croissance, alors qu’il peut conférer une protection aux cellules en conditions de stress environnemental. En plus, l’accumulation de tréhalose a été liée à la virulence chez les levures pathogènes. Normalement, la biosynthèse de tréhalose est effectuée par un complexe de protéines comprenant une tréhalose-6-phosphate synthéase, une tréhalose-6-phosphate phosphatase et des protéines régulatrices. Chez la moisissure pathogène Aspergillus fumigatus, on a identifié quatre gènes potentiels de tréhalose-6-phosphate synthéases : tpsA, tpsB, tpsC et tpsD. Nous avons observé une accumulation de tréhalose pendant le cycle de vie de A. fumigatus et elle a été accompagnée par une expression considérable de tpsA et tpsB. Le tréhalose s’est accumulé suite à un choc thermique et cette accumulation correspondait à une expression accrue de tpsA. On a donc fait une disruption de tpsA, tpsB et de ces deux gènes chez A. fumigatus. Seulement le double mutant ΔtpsAB n’a pas accumulé de tréhalose. Ce mutant démontrait un délai de germination à température de croissance normale. Ce délai était plus marqué à température élevée à laquelle les spores de ΔtpsAB ont démontré un défaut grave de viabilité, de croissance et de sporulation subséquente. En plus, on a des preuves que ΔtpsAB a un défaut dans la paroi cellulaire. Des études sont en cours chez un modèle de souris pour déterminer si ΔtpsAB a un défaut de virulence, ce qui pourrait faire des tréhalose synthéases une nouvelle cible pour la thérapie antifongique.
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Wong, Ee Tsin. "The impact of cell architecture on activation and output of the p53 stress response pathway." Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2006. http://wwwlib.umi.com/cr/ucsd/fullcit?p3225303.
Full textAbstract:
Thesis (Ph. D.)--University of California, San Diego, 2006.<br>Title from first page of PDF file (viewed September 21, 2006). Available via ProQuest Digital Dissertations. Vita. Includes bibliographical references.
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Prochnik, Simon Edward. "The role of the fission yeast Wis1 pathway in stress response and cell cycle control." Thesis, University of Edinburgh, 1998. http://hdl.handle.net/1842/11284.
Full textAbstract:
This thesis consists of work on the <I>wis1</I> pathway: the analysis of some of the upstream components and the isolation and characterisation of genes that lie downstream of <I>wis1</I>. The <I>mcs4, win1 </I>and <I>wis4</I> genes had already been shown to lie upstream of <I>wis1</I>. Strains were constructed with different combinations of mutations in these genes. <I>fbp1</I> transcription was assayed in these strains. An additive effect was seen in <I>win1 wis4</I> double mutants, suggesting that <I>win1</I> and<I> wis4</I> act in parallel. To identify functionally-related genes downstream of <I>wis1</I>, the stress sensitivity of <I>wis1Δ</I> cells was exploited. A screen for extragenic suppressing mutations was carried out. Several hundred heat resistant mutants were isolated. Some also presented the salt sensitivity and/or cell length defect of <I>wisΔ</I>. Twelve such <I>sow</I> (for suppressor of <I>wis1Δ</I>) mutants, each of which containing a single suppressing mutation, were analysed further. They fell into two linkage groups: <I>sow1</I> (nine strains) and <I>sow2</I> (three strains). When the <I>sow </I>mutations were crossed into a <I>wis1<SUP>+</SUP></I> background, both <I>sow1</I> and <I>sow2</I> were able to grow at temperatures above the usual range for <I>S. pombe</I>. In addition, <I>sow1</I> strains divide at a shorter length than wild type, indicating a mitotic advance, and <I>sow2</I> cells have a slightly aberrant morphology. To determine whether the <I>sow</I> mutations corresponded to any known genes, crosses were carried out between the <I>sow</I> mutants and mutants in the following genes: <I>wis1</I> pathway genes (<I>sty1, atf1</I>, <I>ppa1, ppa2 </I>and <I>ppe1</I>), cAMP pathway genes (<I>cyr1</I>, <I>pka1</I>) cell cycle regulation genes (<I>cdc2, cdc25, wee1, cdc13</I>), a heat shock protein (hsp90) gene (<I>swo1</I>) and a gene required for maintenance of the mitotic cell cycle (<I>pat1</I>).
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Farabaugh, Kenneth Thomas kt. "Insights into a Novel Signaling Pathway that Determines Cell Fate in Response to Hyperosmotic Stress." Case Western Reserve University School of Graduate Studies / OhioLINK, 2019. http://rave.ohiolink.edu/etdc/view?acc_num=case1571692276973131.
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Botha, Cornelis Petrus. "Simulation of the human energy system / Cornelis Petrus Botha." Thesis, North-West University, 2002. http://hdl.handle.net/10394/9623.
Full textAbstract:
Preface -
Biotechnology is generally accepted to be the next economical wave of the future. In order to attain
the many benefits associated with this growing industry simulation modelling techniques have to be
implemented successfully. One of the simulations that ne' ed to be performed is that of the human
energy system.
Pharmaceutical companies are currently pouring vast amounts of capital into research regarding
simulation of bodily processes. Their aim is to develop cures, treatments, medication, etc. for major
diseases. These diseases include epidemics like diabetes, cancer, cardiovascular diseases, obesity,
stress, hypertension, etc. One of the most important driving forces behind these diseases is poor
blood sugar control.
The blood glucose system is one of the major subsystems of the complete human energy system. In
this study a simulation model and procedure for simulating blood glucose response due to various
external influences on the human body is presented.
The study is presented in two parts. The first is the development of a novel concept for quantifying
glucose energy flow into, within and out of the human energy system. The new quantification unit
is called ets (equivalent teaspoons sugar). The second part of the study is the implementation of the
ets concept in order to develop the simulation model.
Development of the ets concept -
In the first part of the study the ets concept, used for predicting glycaemic response, is developed
and presented.
The two current methods for predicting glycaemic response due to ingestion of food are discussed,
namely carbohydrate counting and the glycaemic index. Furthermore, it is shown that it is currently
incorrectly assumed that 100% of the chemical energy contained in food is available to the human
energy system after consumption. The ets concept is derived to provide a better measure of
available energy from food.
In order to verify the ets concept, two links with ets are investigated. These are the links with
insulin response prediction as well as with endurance energy expenditure. It is shown that with both
these links linear relationships provide a good approximation of empirical data. It is also shown that
individualised characterisation of different people is only dependent on a single measurable variable
for each link.
Lastly, two novel applications of the ets concept are considered. The first is a new method to use the
ets values associated with food and energy expenditure in order to calculate both short-acting and
long-acting insulin dosages for Type 1 diabetics. The second application entails a new
quantification method for describing the effects of stress and illness in terms of ets.
Development of the blood glucose simulation model -
The second part of the study presents a literature study regarding human physiology, the
development for the blood glucose simulation model as well as a verification study of the
simulation model.
Firstly, a brief overview is given for the need and motivation behind simulation is given. A
discussion on the implementation of the techniques for construction of the model is also shown. The
procedure for solving the model is then outlined.
During the literature study regarding human physiology two detailed schematic layouts are
presented and discussed. The first layout involves the complex flow pathways of energy through the
human energy system. The second layout presents a detailed discussion on the control system
involved with the glucose energy pathway.
Following the literature review the model for predicting glycaemic response is proposed. The
design of the component models used for the simulations of the internal processes are developed in
detail as well as the control strategies implemented for the control system of the simulation model.
Lastly, the simulation model is applied for glycaemic response prediction of actual test subjects and
the quality of the predictions are evaluated. The verification of the model and the procedure is
performed by comparing simulated results to measured data. Two evaluations were considered,
namely long-term and short-term trials. The quality of both are determined according to certain
evaluation criteria and it is found that the model is more than 70% accurate for long-term
simulations and more than 80% accurate for short-term simulations.
Conclusion -
In conclusion, it is shown that simplified simulation of the human energy system is not only
possible but also relatively accurate. However, in order to accomplish the simulations a simple
quantification method is required and this is provided by the ets concept developed in the first part
of this study. Some recommendations are also made for future research regarding both the ets
concept and the simulation model.
Finally, as an initial endeavour the simulation model and the ets concept proposed in this study may
provide the necessary edge for groundbreaking biotechnological discoveries.<br>PhD (Mechanical Engineering) North-West University, Potchefstroom Campus, 2003
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Bain, Peter A., and n/a. "Gene Expression Profiling of Cylindrospermopsin Toxicity." Griffith University. School of Biomolecular and Physical Sciences, 2007. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20080404.145834.
Full textAbstract:
Cylindrospermopsin (CYN) is a toxic alkaloid produced by several freshwater cyanobacterial species, the most prevalent in Australian waters being Cylindrospermopsis raciborskii. The occurrence of CYN-producing cyanobacteria in drinking water sources worldwide poses a potential human health risk, with one well-documented case of human poisoning attributed to the toxin. While extensive characterisation of CYN-induced toxicity has been conducted in rodents both in vivo and in primary cell cultures, little is known about mechanisms of toxicity in human cell types. This thesis describes studies undertaken to further define the molecular mechanisms of CYN toxicity in human cells. Concentration-response relationships were determined in various cultured human cell types using standard toxicity assays. As expected, CYN caused dose-dependent decreases in the growth of three cell lines, HepG2, Caco-2 and HeLa, and one primary cell type, human dermal fibroblasts, according to tetrazolium reduction assays. CYN treatment did not disrupt cellular membranes according to the lactate dehydrogenase release assay in HepG2 or Caco-2 cells after 24, 48 or 72 h exposure, but did cause membrane disruption in fibroblasts after 72 h exposure to relatively high concentrations of the toxin. Apoptosis occurred more readily in HeLa cells than HepG2 cells or fibroblasts, with 72 h exposure to 1 &mug/mL required before statistically significant rates of apoptosis occurred in the latter cell types. CYN did not appear to directly affect the structure of actin filaments or microtubules under the conditions used in the present study. The major portion of the work presented in this thesis comprises a large-scale interrogation of changes in gene expression induced by the toxin in cultured cells. To assess the effects of CYN on global gene expression, relative messenger RNA (mRNA) levels in human dermal fibroblasts and HepG2 cells after 6 h and 24 h exposure to 1 &mug/mL CYN were determined using oligonucleotide microarrays representing approximately 19 000 genes. Overall, the number of transcripts significantly altered in abundance was greater in fibroblasts than in HepG2 cells. In both cell types, mRNA levels for genes related to amino acid biosynthesis, carbohydrate metabolism, and protein folding and transport were reduced after CYN treatment, while transcripts representing genes for apoptosis, RNA biosynthesis and RNA processing increased in abundance. More detailed data analyses revealed the modulation of a number of stress response pathways—genes regulated by NF-&kappaB were induced, DNA damage response pathways were up-regulated, and a large number of genes involved in endoplasmic reticulum stress were strongly down-regulated. Genes for the synthesis and processing of mRNA, tRNA and rRNA were strongly up-regulated, indicating that CYN treatment may increase the turnover of all forms of cellular RNA. A small group of genes were differentially expressed in HepG2 cells and fibroblasts, revealing cell-specific responses to the toxin. Selected changes in transcript level were validated using real-time quantitative reverse transcriptase PCR (qRT-PCR). The modulation of stress response pathways by CYN, indicated by microarray analysis, was further investigated using other methods. The role of tumour suppressor protein p53 in CYN-mediated gene expression was confirmed by measuring the expression of known p53-regulated genes following CYN treatment of HepG2 cells and human dermal fibroblasts using qRT-PCR. Western blotting of protein extracts from CYNtreated cells showed that p53 protein accumulation occurred in HepG2 cells, providing additional evidence of the activation of the p53 pathway by CYN in this cell line. The immediate-early genes JUN and FOS were found to be induced by CYN in a concentration-dependent manner, and MYC was induced to a lesser extent. The mitogen-activated protein kinase c-Jun NH2-terminal kinase, implicated in the ribotoxic stress response initiated by damage to ribosomal RNA, appeared to become phosphorylated in HeLa cells after CYN exposure, suggesting that ribotoxic stress may occur in response to CYN in at least some cell types. The expression of a reporter gene under the control of a response element specific for NF-&kappaB was induced at the mRNA level but inhibited at the protein level. This shows that while transcription factors such as p53 and NF-&kappaB are apparently activated in response to the toxin, transactivation of target genes may not necessarily manifest a corresponding increase at the protein level. The current work contributes significantly to the current understanding of cylindrospermopsin toxicity in human-derived cell types, and provides further insight into putative modes of action.
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Bain, Peter A. "Gene Expression Profiling of Cylindrospermopsin Toxicity." Thesis, Griffith University, 2007. http://hdl.handle.net/10072/367068.
Full textAbstract:
Cylindrospermopsin (CYN) is a toxic alkaloid produced by several freshwater cyanobacterial species, the most prevalent in Australian waters being Cylindrospermopsis raciborskii. The occurrence of CYN-producing cyanobacteria in drinking water sources worldwide poses a potential human health risk, with one well-documented case of human poisoning attributed to the toxin. While extensive characterisation of CYN-induced toxicity has been conducted in rodents both in vivo and in primary cell cultures, little is known about mechanisms of toxicity in human cell types. This thesis describes studies undertaken to further define the molecular mechanisms of CYN toxicity in human cells. Concentration-response relationships were determined in various cultured human cell types using standard toxicity assays. As expected, CYN caused dose-dependent decreases in the growth of three cell lines, HepG2, Caco-2 and HeLa, and one primary cell type, human dermal fibroblasts, according to tetrazolium reduction assays. CYN treatment did not disrupt cellular membranes according to the lactate dehydrogenase release assay in HepG2 or Caco-2 cells after 24, 48 or 72 h exposure, but did cause membrane disruption in fibroblasts after 72 h exposure to relatively high concentrations of the toxin. Apoptosis occurred more readily in HeLa cells than HepG2 cells or fibroblasts, with 72 h exposure to 1 µg/mL required before statistically significant rates of apoptosis occurred in the latter cell types. CYN did not appear to directly affect the structure of actin filaments or microtubules under the conditions used in the present study. The major portion of the work presented in this thesis comprises a large-scale interrogation of changes in gene expression induced by the toxin in cultured cells. To assess the effects of CYN on global gene expression, relative messenger RNA (mRNA) levels in human dermal fibroblasts and HepG2 cells after 6 h and 24 h exposure to 1 µg/mL CYN were determined using oligonucleotide microarrays representing approximately 19 000 genes. Overall, the number of transcripts significantly altered in abundance was greater in fibroblasts than in HepG2 cells. In both cell types, mRNA levels for genes related to amino acid biosynthesis, carbohydrate metabolism, and protein folding and transport were reduced after CYN treatment, while transcripts representing genes for apoptosis, RNA biosynthesis and RNA processing increased in abundance. More detailed data analyses revealed the modulation of a number of stress response pathways—genes regulated by NF-?B were induced, DNA damage response pathways were up-regulated, and a large number of genes involved in endoplasmic reticulum stress were strongly down-regulated. Genes for the synthesis and processing of mRNA, tRNA and rRNA were strongly up-regulated, indicating that CYN treatment may increase the turnover of all forms of cellular RNA. A small group of genes were differentially expressed in HepG2 cells and fibroblasts, revealing cell-specific responses to the toxin. Selected changes in transcript level were validated using real-time quantitative reverse transcriptase PCR (qRT-PCR). The modulation of stress response pathways by CYN, indicated by microarray analysis, was further investigated using other methods. The role of tumour suppressor protein p53 in CYN-mediated gene expression was confirmed by measuring the expression of known p53-regulated genes following CYN treatment of HepG2 cells and human dermal fibroblasts using qRT-PCR. Western blotting of protein extracts from CYNtreated cells showed that p53 protein accumulation occurred in HepG2 cells, providing additional evidence of the activation of the p53 pathway by CYN in this cell line. The immediate-early genes JUN and FOS were found to be induced by CYN in a concentration-dependent manner, and MYC was induced to a lesser extent. The mitogen-activated protein kinase c-Jun NH2-terminal kinase, implicated in the ribotoxic stress response initiated by damage to ribosomal RNA, appeared to become phosphorylated in HeLa cells after CYN exposure, suggesting that ribotoxic stress may occur in response to CYN in at least some cell types. The expression of a reporter gene under the control of a response element specific for NF-?B was induced at the mRNA level but inhibited at the protein level. This shows that while transcription factors such as p53 and NF-?B are apparently activated in response to the toxin, transactivation of target genes may not necessarily manifest a corresponding increase at the protein level. The current work contributes significantly to the current understanding of cylindrospermopsin toxicity in human-derived cell types, and provides further insight into putative modes of action.<br>Thesis (PhD Doctorate)<br>Doctor of Philosophy (PhD)<br>School of Biomolecular and Physical Sciences<br>Faculty of Science, Environment, Engineering and Technology<br>Full Text
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Schwaiger, Marion [Verfasser], and Markus [Akademischer Betreuer] Heinrichs. "Long-term consequences of childhood adversity – exploring stress responsive molecular pathways and a psychobiological intervention model." Freiburg : Universität, 2016. http://d-nb.info/1124005110/34.
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Wang, Qunwei. "ATM kinase and DNA-PKcs pathways regulate the expression of IL-23 in dendritic cells : interaction with ER-stress pathways and impact on Th17 responses." Thesis, University of Nottingham, 2014. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.718993.
Full textAbstract:
Dendritic cells (DCs) are key members of the immune system as they detect danger and reflect this during presentation of antigen to T cells. DC produce cytokines that influence both the innate and adaptive immune system. Notably they produce the IL- 12-family of cytokines known for their influence on T helper (Th) cell priming. Interleukin-23 is one such cytokine that is recognised for its roles in supporting Thl7 cell differentiation characterised by IL-17 and IL-22 production, in driving IL-17 secretion by y6 T cells, and in its contribution to anti-bacterial immunity and autoimmunity. A body of evidence exists that Dectin-l/TLR-2 Syk- CARD9-NFkB signalling pathways, p38 MAPK/Erk- NFk-B signalling pathways and PI3K/Akt pathways are involved in IL-23 production in their respective settings. However, little is known at the molecular level regarding the regulation of IL-23 secretion in human DC. Here we show for first time that the Ataxia Telangiectasia Mutated (ATM) pathway and DNA-Protein kinase catalytic subunit (DNA-PKcs), involved in DNA-damage-sensing and DNA double strand break (DSB) repair, act as repressors of IL-23 in DC. Inhibition of ATM with the highly-selective antagonist KU55933, or DNA-PKcs with NU7441, markedly increased IL-23 secretion human monocyte-derived DC (moDC) and freshly isolated CDlc+ myeloid DC (myDC). In contrast, inhibiting the closely related mammalian target of rapamycin (mTOR) had no effect on IL-23. Priming naive CD4+ T- cells in the presence of supernatant from ATM or DNA-PKcs-inhibited DC resulted in increased Thl7 responses. Whilst ATM or DNA-PKcs-blockade increased the abundance of pl9, p35 and p40 mRNA, IL- 12p70 secretion was much less affected. In order to further examine a role for ATM and DNA-PKcs in IL-23 regulation we exposed DC to low doses of ionizing radiation. Exposure of DC to X-rays resulted in a rapid increase (15min) in ATM phosphorylation and a corresponding depression of IL-23 production. Importantly, ATM-inhibition with KU55933 prevented radiation-induced ATM phosphorylation and abrogated the capacity of X-rays to suppress IL-23. Interestingly, the viability of DC exposed to X-rays after DNA-Pkcs inhibition was substantially affected but unaffected in DC exposed to IR after ATM inhibition, suggesting an important role for DNA-PKcs in the resistance of DC to DNA-damage. To explore how IL-23 was repressed by ATM or DNA-PKcs we examined the unfolded protein response (UPR) pathways by measuring generation of the spliced form of X-box protein-1 (XBPls), a key ER-stress transcription factor. When ATM or DNA-Pkcs were inhibited the abundance of XBPls mRNA increased and was followed 3hr later by increased p l9 transcription and subsequent IL-23 release. Unexpectedly, in DC deficient for the UPR-related CHOP transcription factor, inhibition of ATM or DNA-PKcs continued to secrete high levels of IL-23. However, inhibition of the PERK arm of the UPR reduced IL-23 production by matured DC, and importantly partially suppressed ATM- or DNA-PKcs-dependent IL-23. In summary, in addition to their canonical roles in DNA-repair, ATM and DNA-PKcs also regulate immune-activation by DC. Whilst their activation further represses IL-23-dependent responses, their inhibition permits markedly high levels of IL-23 to be generated. These pathways thus represent new therapeutic targets in autoimmunity and vaccine development.
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Wei, Yu-Chen. "Pct1 regulates phosphatidylcholine synthesis in response to changes in surface curvature elastic stress sensed on the inner nuclear membrane." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/287931.
Full textAbstract:
Cell and organelle membranes consist of a complex mixture of phospholipids that determine their size, shape, and function. Among the distinct types of phospholipids found in membranes of living organisms, phosphatidylcholine (PC) is the most abundant. The rate-limiting step of the predominant pathway for PC synthesis in eukaryotic cells is catalysed by the enzyme, CTP: phosphocholine cytidylyltransferase α (CCTα or PCYT1A). CCTα has a critical role in lipid metabolism and also has direct clinical relevance as mutations in CCTα result in an interesting spectrum of human diseases, such as lipodystrophy with fatty liver, growth plate dysplasia and cone-rod related dystrophy. Numerous biochemical and structural studies on purified CCTα have revealed its membrane-bound activation and suggested that it acts as a lipid compositional sensor, yet the in vivo mechanism of how CCTα senses and regulates PC levels in membranes remains unclear. Here I show that in budding yeast Saccharomyces cerevisiae, Pct1, the yeast homolog of CCTα, is intranuclear and translocates to the nuclear membrane in response to changes in membrane properties and the need for membrane PC synthesis. By aligning imaging with lipidomic analysis and data-driven modelling, Pct1 membrane association is demonstrated to correlate with membrane stored curvature elastic stress estimates. Furthermore, this process occurs inside the nucleus, although nuclear localization signal mutants can compensate for the loss of endogenous Pct1. These data suggest an ancient mechanism by which CCTα senses lipid packing defects and regulates phospholipid homeostasis from the nucleus. Additionally, I identified the importance of mammalian CCTα in early adipogenesis and investigated the enzymatic function of PCYT1A mutants in fibroblasts from lipodystrophic patients. The allele Val142Met is evaluated to be the main cause of loss-of-function in the compound heterozygous mutations by using yeast survival assay. These results collectively provide preliminary evidence for the pathogenicity of PCYT1A mutations in adipose tissue. From yeast to humans, this study uncovers the critical role of Pct1/CCTα in maintaining the internal membrane environment.
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Evans, Joanne Susan. "A role for the stress-activated MAPK pathway of Schizosaccharomyces pombe in the cellular response to ionising radiation." Thesis, University of Oxford, 2004. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.410679.
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Bourougaa, Karima. "Diversifying the p53 pathway in response to Endoplasmic Reticulum stress via alternative translation initiation of the p53 mRNA." Paris 7, 2009. http://www.theses.fr/2009PA077128.
Full textAbstract:
La protéine p53 est le produit d'un gène muté dans plus de 50% des cancers humains. P53 est un facteur de transcription capable d'induire, en réponse à différents stress, l'arrêt du cycle cellulaire en Gl et en G2, la réparation de l'ADN, ou, si le dommage est trop sévère, la mort cellulaire. Un grand nombre de gènes répondent à p53 mais il reste quand même difficile de comprendre comment les cellules parviennent à différencier et à intégrer l'activation de p53 en une réponse bien spécifique. Récemment, différents isoformes de p53 ont été identifiés mais leur rôle physiologique reste encore inconnu. L'isoforme p53/47 ne possède pas les 39 premiers acides aminés de p53 correspondant au premier domaine de transactivation et au site de liaison du régulateur négatif Mdm2. D'abord décrite comme un antagoniste de p53, p53/47 présente des vertus pro-apoptotiques. Nous avons précédemment montré que p53/47 pouvait être exprimée de façon indépendante de celle de p53 spécifiquement en réponse aux stress associés à l'hypoxie tumorale. De plus, p53 peut s'oligomériser avec p53/47. Les différents complexes formés présentent des activités et des propriétés biochimiques différentes comparés à ceux formés uniquement par p53. Nos résultats montrent que l'activation de la protéine PERK lors du stress du réticulum endoplasmique conduit à une surexpression de p53/47. Par ailleurs, ,ce stress induit une tétramérisation sélective de p53/47 qui est associée à la régulation du gène 14-3-3a conduisant à un arrêt du cycle cellulaire en G2. Ces résultats montrent comment en réponse à un stress spécifique, c'est un isoforme particulier qui est activé conduisant à une réponse cellulaire définie<br>The p53 protein is the product of a tumour suppressor gene mutated in 50% of human tumours and it has been suggested that all tumours carry defects in the p53 pathways. The p53 protein is a tetrameric transcription factor which is activated in response to a large panel of stresses. P53 downstream pathways control Gl and G2 cell cycle arrest, DNA repair or apoptosis. Even though a large number of p53 response genes have been identified it is still not clear how cells can differentiate the cell biological outcome of p53 activation. More recently, several p53 isoforms hâve been identified and the physiological roles of these are now starting to unravel. The p53/47 isoform lacks the first 39 amino acids including the Mdm2 binding site and trans-actiyation domain I. In consequence, p53/47 homo and hetero-oligomer complexes exhibit altered activity and biochemical properties. The results presented here show that activation of the PERK kinase in response to endoplasmic reticulum (ER) stress induces cap-independent translation of p53/47 via an Internai Ribosome Entry Site (IRES) located within the first 120 nt of the p53 encoding sequence. ER stress promotes selective homo-oligomerisation of p53/47 which induces 14-3-3a and a G2 arrest. At the same time, p53/47 prevents p53-induced expression of p21 and Gl arrest. This thesis shows how IRES-dependent mRNA translation in response to a specific cellular stress pathway alters the expression of p53/47 allowing the cell to differentiate the cell biological outcome of p53 activation. This study can then help to unravel how thé cells can differentiate the cell biological response to different types of cell stress
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Machingura, Marylou. "Activity of the beta-cyanoalanine synthase pathway is associated with the response to abiotic stress by Arabidopsis thaliana." OpenSIUC, 2012. https://opensiuc.lib.siu.edu/dissertations/628.
Full textAbstract:
Cyanide is produced throughout a plant's life cycle alongside the hormone ethylene by oxidation of 1-aminocyclopropane-1-carboxylic acid. Production increases during certain developmental stages such as seed germination, seedling elongation, fruit ripening and senescence. Abiotic stresses increase ethylene production giving rise to `stress cyanide'. Cyanide also comes from metabolism of cyanogenic compounds. Cyanide is however, a toxic chemical which readily binds to metallo-enzymes inhibiting primary metabolic processes. Plants have mechanisms to maintain cyanide homeostasis such as the β-cyanoalanine pathway whereby cysteine reacts with cyanide forming β--cyanoalanine, mediated by β-cyanoalanine synthase and cysteine synthase. A dual nitrilase 4 enzyme then converts the β-cyanoalanine into asparagine or aspartate and ammonium. Studies have suggested that the physiological function of the pathway is not restricted to detoxification and assimilation of excess cyanide. The overall research goal was to investigate the role of the pathway in plant tolerance to water deficit and exogenous cyanide exposure in Arabidopsis thaliana. The first objective was to investigate responsiveness of the pathway to duration and intensity of water deficit and cyanide exposure. The second was to investigate the contribution of enzymes associated with the pathway to cyanide metabolism. The questions addressed were whether there is enzymatic redundancy in enzymes associated with the first step of cyanide detoxification and whether there is pathway redundancy between the β-cyanoalanine and an alternative sulfurtransferase pathway. A. thaliana Col-0 and three SALK-line mutants with a T-DNA insertion for the genesAtCysA1, AtCysC1 and AtNIT4 were grown and exposed to water stress. Physiological and biochemical measurements were taken. The results showed a transient increase in cyanide concentration and β-cyanoalanine synthase activity on exposure to stress. The response pattern was similar regardless of intensity or duration of stress. Knocking out AtCysA1 or AtCysC1 did not impair the ability of plants to metabolize cyanide and tolerate stress i.e the enzymes were functionally redundant. The AtNIT4 mutant however, was impaired in cyanide metabolism and exhibited a sensitive phenotype under both stresses, suggesting that the cyanoalanine pathway is the sole pathway in cyanide detoxification. The results show that the pathway may be an important tool in improvement of plant tolerance to abiotic stress.
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Alvarenga, Santos Buiate Ester. "ESTABLISHMENT OF BIOTROPHY BY THE MAIZE ANTHRACNOSE PATHOGEN COLLETOTRICHUM GRAMINICOLA: USE OF BIOINFORMATICS AND TRANSCRIPTOMICS TO ADDRESS THE POTENTIAL ROLES OF SECRETION, STRESS RESPONSE, AND SECRETED PROTEINS." UKnowledge, 2015. http://uknowledge.uky.edu/plantpath_etds/17.
Full textAbstract:
Colletotrichum graminicola is a hemibiotrophic pathogen of maize that causes anthracnose leaf and stalk rot diseases. The pathogen penetrates the host and initially establishes an intracellular biotrophic infection, in which the hyphae are separated from the living host cell by a membrane that is elaborated by the host, apparently in response to pathogen signals. A nonpathogenic mutant (MT) of C. graminicola was generated that germinates and penetrates the host normally, but is incapable of establishing a normal biotrophic infection. The mutated gene is Cpr1, conserved in eukaryotes and predicted to encode a component of the signal peptidase complex. How can we explain why the MT is normal in culture and during early stages of pathogenicity, but is deficient specifically in the ability to establish biotrophy? To address this, first I characterized the insertion in the 3’ UTR of the MT strain in detail, something that had not been done before. The wild-type (WT) transcript did not differ from predictions, but the MT produced several aberrant transcript species, including truncated and non-spliced transcripts, and the normal one. Aberrant splicing of MT cpr1 was observed both in RNAseq transcriptome data and reverse-transcription polymerase chain reaction (RT-PCR), under different growth conditions and in planta. I also conducted a bioinformatic analysis of other conserved components of the secretory pathway in the MT and WT in planta. One explanation for nonpathogenicity of the MT is that it cannot cope with an increase in secretory activity during infection, and fails to produce necessary pathogenicity factors. With the transcriptome data, I was able to identify effector proteins that were expressed in the WT but not in the MT. Another possible explanation for the MT phenotype is that the MT can’t adapt to stress imposed by the plant. I developed a growth assay to characterize the effect of chemical stressors in vitro. The MT was more sensitive to most stressors, when compared to the WT. The transcriptome data indicates that the genes involved in different stress pathways are expressed in planta in both WT and MT, although very few genes are differentially expressed across the different growth stages.
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Flanagan, Marc David. "The roles and regulation of ubiquitin/ubiquitin-like protein conjugation pathways in responses to oxidative stress in Schizosaccharomyces pombe." Thesis, University of Newcastle Upon Tyne, 2013. http://hdl.handle.net/10443/1825.
Full textAbstract:
Ubiquitin and ubiquitin-like proteins (Ubls) are conjugated to proteins to regulate activity, stability, localisation or function. Ubiquitin/Ubl conjugation pathways are highly conserved in eukaryotes, and usually involve activating enzymes (E1s) and conjugating enzymes (E2s) specific for each ubiquitin/Ubl. Many studies have suggested that ubiquitin/Ubl conjugation pathways are important for oxidative stress resistance. However, there is much to learn regarding the roles of these pathways in oxidative stress responses. Additionally, limited studies in mammalian cells and yeast have indicated that certain E1s and E2s are redox-regulated, although how this relates to stress resistance is largely unclear, and these regulatory mechanisms have never been shown to be conserved in eukaryotes. Here, the roles and regulation of ubiquitin/Ubl conjugation pathways in responses to oxidative stress are investigated in Schizosaccharomyces pombe. Firstly, while our previous research has shown that the budding yeast E2, Cdc34, is redox-regulated, it was unclear whether ubiquitination is redox-regulated in other organisms. Results presented here show that the fission yeast Cdc34 homologue, Ubc15, is redox-regulated, suggesting that redox regulation of specific ubiquitination events may be conserved. Furthermore, Ubc15 is important for resistance to oxidative stress in S. pombe. Secondly, the Ubl Urm1 is found to be important for resistance to a range of stress conditions in S. pombe, as in S. cerevisiae, thus demonstrating for the first time that urmylation has conserved roles in stress resistance in eukaryotes. Additionally, urmylation controls the activation of a conserved mitogen-activated protein kinase during exposure to H2O2. Finally, although autophagic Ubl conjugation is not important for oxidative stress responses in S. pombe, these investigations have identified an E2 with roles in oxidative stress responses and cell cycle control. Taken together, these findings advance the study of the roles of ubiquitin/Ubl conjugation pathways in responses to oxidative stress, and offer exciting prospects for future investigations.
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Leitao, Beatriz Belchior. "The role of the SUMO pathway in the cellular responses to oxidative stress signals in human endometrium." Thesis, Imperial College London, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.544293.
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Wang, Hong. "Regulation of the plant one-carbon metabolic pathway and global gene responses in maize under salt stress." Diss., The University of Arizona, 2001. http://hdl.handle.net/10150/289752.
Full textAbstract:
One-carbon (C₁) metabolism. C₁ metabolism is central to all organisms, because C₁ units have essential roles in biosyntheses of basic materials for living cells, such as protein, nucleic acids, choline and its derivatives. One unique feature of plant C₁ metabolism is that it channels significant C₁ flux from primary carbon metabolism to methylated metabolites. Part I of this dissertation presents functional analysis of plant methyane-THF reductases (MTHFRs) in Arabidopsis and maize, and regulation of the plant C₁ metabolic pathway and glycine betaine (GlyBet) biosynthesis in maize GlyBet near isogenic lines (NILs). Plant MTHFRs were isolated from Arabidopsis and maize and functionally characterized in yeast. Unlike mammalian MTHFRs, the plant enzymes strongly prefer NADH to NADPH and are not inhibited by S-adenosyl-methionine. An NADH dependent MTHFR reaction could be reversible in the cytosol, supported by radiotracer labeling data using [methyl-¹⁴C]methyltetrahydrofolate. Systematic analyses of profiling transcript abundance of plant C₁ genes using microarrays indicated that the regulation of the plant C₁ metabolic pathway may be "fine-tuned", or at a downstream "outlet" point in maize. Comparison of GlyBet NILs through analyses of metabolites, [¹⁴C]formaldehyde labeling and northern blotting of phosphoethanolamine methyltransferase (P-EAMT) indicated that the regulation glycine betaine biosynthesis is at P-EAMT, whose transcript is probably repressed by its product, phospho-choline. Global gene responses to salt stress. To understand how genes respond to salt stress is very important for the genetic improvement of salt tolerance in plants. Part II of this dissertation presents a systematic analysis of global gene responses to salt stress in maize roots, using microarrays, that contained ∼8,000 ESTs. Microarray expression analysis revealed that 916 different ESTs were up- or down-regulated under salt stress (611 up-regulated, 305 down-regulated), representing 11% of ∼8,000 ESTs printed on slides. These up- or down-regulated ESTs were clustered into various subgroups based on repression or induction of the timing, amplitude and duration of their transcripts. Contig analysis assigned 916 up- or down-regulated ESTs into 472 tentative unique genes (51.6%). These responsive genes are involved in a broad range of cellular processes, biochemical pathways and signal transduction cascades etc. The functions of some regulated genes are discussed.
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Amarsaikhan, Nansalmaa. "The Role Of The Cross Pathway Control Protein In The Stress Response And Adaptation Of Aspergillus Species To Antifungals." Phd thesis, METU, 2010. http://etd.lib.metu.edu.tr/upload/12612308/index.pdf.
Full textAbstract:
In this study, the adaptation and response of Aspergillus nidulans and Aspergillus
fumigatus wild type and cpcA strains to antifungal compounds were studied using cultural,
genetic and proteomic methods. CpcA is the fungal cross pathway control protein which may
also have a role in the development of resistance to antifungal that has become a major problem
in human and plant fungal diseases and many studies are devoted to address the drug resistance
mechanisms. Cell adapts itself to stress when it is subjected to a stress repeatedly. The ancestor
of CpcA, ATF4 (CREB2) has recently been found to be important in the survival of tumor cells
after starvation and nutrient limitation and these findings are expected to open new insights into
the future antifungal therapy. Fungal cross pathway control system conserves similar mechanism
with the stress response pathway in humans as a response to amino acid starvation. Fungal
adaptation to antifungal agents was studied using the genetic model A. nidulans with the
experimentally induced adaptation setup. It was concluded that A. nidulans cells are able to
adapt to antifungal. In order to understand how cell becomes resistant to a previously susceptible
agent, it is important to investigate the process when the cell encounters the agent for the first
time. Fungal cellular response to antifungal drugs was studied using the human opportunistic
pathogen A. fumigatus at the protein level. This is the first proteomic study directed to
investigate the A. fumigatus response to voriconazole (VRC). The recently developed two
dimensional gel electrophoresis approach, Fluorescence 2-D Differential Gel Electrophoresis
(DIGE) method was applied to visualize differentially expressed proteins. It was concluded that,
about 150 proteins were differentially regulated as a response to stress exerted by azole group
antifungal drugs. cpcA strains of A. nidulans and A. fumigatus were compared to wild type
strains in terms of susceptibility to various stresses, adaptation potential also at the proteome
level. The results obtained in this study showed that CpcA was important in the response of
Aspergillus to oxidative, heat stress and in the adaptation of cells to VRC and that its absence
drastically changed the cellular response to VRC at the protein level by changing the expression
of about 80 proteins. Thus, this protein is a good candidate in future as a potential drug resistance
mediator and further characterization is needed to elucidate its mechanism of action on drug
resistance.
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Morris, Bethany H. "The Effect of Stress on Hedonic Capacity in Generalized Anxiety Disorder: A Prospective Experimental Study of One Potential Pathway to Depression." Scholar Commons, 2009. https://scholarcommons.usf.edu/etd/1716.
Full textAbstract:
A growing body of work links psychopathology to changes in hedonic capacity following stressors. This was the first experimental study of the effects of stress on hedonic capacity in an analog generalized anxiety disorder (GAD) sample (a high worry group). Specifically, we utilized an experimental manipulation of stress and a behavioral index of anhedonia to test the hypothesis that individuals with GAD, who are at higher risk for developing depression symptoms, exhibit greater stress-related deficits in hedonic capacity than do nonanxious controls. Further, this study assessed whether stress-induced hedonic deficits predicted future depression. Controls exhibited the expected reward learning pattern in the baseline condition, demonstrating intact hedonic responding, as well as the expected pattern of behavioral anhedonia under stress. Contrary to predictions, worriers demonstrated intact hedonic capacity under stress. The stress effect in worriers was modulated by past depression diagnostic status; whereas worriers with no past depression demonstrated blunted baseline hedonic capacity and heightened hedonic capacity under stress, worriers with past depression demonstrated the normative response pattern. Blunted baseline response bias predicted higher future depression in both groups. We discuss the differential stress effects on behavioral hedonic capacity found as a function of worry, the role of past depression as a moderator of stress effects among worriers, and the need for future work to further explicate the mechanisms that may modulate reward response under stress.
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Oun, Rabiaa R. Salah. "The Rad9.M50 variant of the DNA damage checkpoint protein Rad9 regulates the MAP kinase pathway in the response to heat stress." Thesis, Bangor University, 2016. https://research.bangor.ac.uk/portal/en/theses/the-rad9m50-variant-of-the-dna-damage-checkpoint-protein-rad9-regulates-the-map-kinase-pathway-in-the-response-to-heat-stress(507c63ee-2f27-4f3d-a39b-e7dba7201f95).html.
Full textAbstract:
Environmental stress activates the MAP kinase pathway to regulate transcription and cell division. In this pathway, a MAP kinase kinase kinases (MAPKKK) activates the MAP kinase kinase (MAPKK) which in turn phosphorylates the MAP kinase (MAPK) simultaneously at a serine or threonine residue and a tyrosine residue in close vicinity. In the fission yeast Schizosaccharomyces pombe, the dual specific MAPKK Wis1 phosphorylates the MAPK Sty1/Spc1 at threonine 171 and tyrosine 173. Active Sty1 phosphorylates and activates the transcription factor Atf1. Sty1 is closely related to Hog1 in the budding yeast S.cerevisiae and p38 in human cells. This thesis reveals a novel regulation of Sty1 by the alternative translation product of the DNA damage checkpoint protein Rad9. Alternative translation from the internal AUG codon at position 50 produces an N-terminally truncated protein variant (Rad9-M50), expression of which is restricted to dividing cells. The basal level of this variant is low and increases in the response to heat stress and DNA alkylation by the DNA damaging drug methyl-methanesulfonate (MMS). The key finding reported here is the ability of Rad9 variant to shield Sty1 from dual specific kinases other than Wis1. Loss of Wis1 eliminates the phosphorylation of Sty1 at T171 and Y173. However, removal of the Rad9 variant (rad9-M50A) restores Sty1 phosphorylation in the absence of Wis1 at high temperature and in the presence of oxidative stress. This aberrant stimulation enables Sty1 to phosphorylate Atf1. A model is presented in which Rad9 variant shields Sty1 from an alternative dual-specific kinase, possibly Wee1, Hhp1 (CK1) or Mps1/TTK. The thesis also reports the requirement of the DEAD-box RNA helicase Ded1 for the expression of Rad9 variant and presents genetic evidence linking Rad9 variant with the phosphatase Ptc1 in the response to heat stress and MMS induced DNA damage.
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Weerakoon, Tasmeen Shiny. "Investigation of a putative mitochondrial Twin Arginine Translocation pathway in Arabidopsis thaliana." Miami University / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=miami1501256746410956.
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Keiten-Schmitz, Jan [Verfasser], Stefan [Akademischer Betreuer] Müller, Volker [Gutachter] Dötsch, and Stefan [Gutachter] Müller. "Characterizing the SUMO-targeted ubiquitin ligase pathway in the proteotoxic stress response / Jan Keiten-Schmitz ; Gutachter: Volker Dötsch, Stefan Müller ; Betreuer: Stefan Müller." Frankfurt am Main : Universitätsbibliothek Johann Christian Senckenberg, 2021. http://d-nb.info/1229989196/34.
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Cornes, Maragliano Eric 1987. "A functional study of the conserved LSM proteins in C. elegans reveals their involvement in the stress response of metazoans." Doctoral thesis, Universitat Pompeu Fabra, 2015. http://hdl.handle.net/10803/315473.
Full textAbstract:
Lsm proteins regulate RNA metabolism and are conserved in the three domains of life, typically functioning as RNA-binding complexes involved in a wide range of post-transcriptional mechanisms. Generally, their functions have been explored in unicellular models using biochemical approaches; however their physiological roles in multicellular organisms remain unknown. This gap in knowledge is biomedically relevant since alterations of individual LSM proteins functions have been related to cancer development.
We performed a functional study of the eleven LSM proteins encoded in the C. elegans genome. We found that although lsm-1 and lsm-3 genes are not essential for the viability of the organism, they are required for wild type healthspan. In addition LSM-1 and LSM-3 proteins function in stress responses by promoting cytoplasmic LSM foci formation and influencing Insulin/IGF-like signaling pathway, a major regulator of development and stress response in metazoans. This study uncovers a physiological role for the LSM proteins in multicellular organisms as essential players for healthspan maintenance and stress adaptation.<br>Las proteínas de la familia Lsm están conservadas desde bacterias a humanos y participan en el metabolismo de ARN. Aunque el estudio de sus funciones ha sido generalmente abordado mediante aproximaciones bioquímicas en modelos unicelulares, sus funciones en organismos multicelulares son desconocidas. Su estudio en organismos modelo es de especial relevancia biomédica ya que la alteración específica de ciertas proteínas Lsm ha sido relacionada con el desarrollo del cáncer.
Mediante el estudio funcional de las once proteínas Lsm presentes en C. elegans, mostramos cómo los genes lsm-1 y lsm-3, pese a no ser esenciales para la viabilidad del organismo, son necesarios para el mantenimiento de su salud. Además, LSM-1 y LSM-3 funcionan durante la respuesta a estrés promoviendo la agregación citoplasmática de proteínas LSM y contribuyendo a la correcta señalización a través de la vía de la Insulina/IGF-like, importante en la regulación del metabolismo y la respuesta a estrés en metazoos. Este estudio destaca el importante papel fisiológico de las proteínas LSM en el desarrollo y la adaptación al estrés de un organismo multicelular.
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Ivanova, Petkova Mima. "Functional, genomic and molecular characterisation of Mtl1, an element of the CWI pathway of Saccharomyces cerevisiae with a role in the oxidative stress response." Doctoral thesis, Universitat de Lleida, 2011. http://hdl.handle.net/10803/51089.
Full textAbstract:
Saccharomyces cerevisiae és un microorganisme eucariota que s’utilitza com a
model per l’estudi de les vies de transducció de senyal implicades en la resposta a estrès
oxidatiu. Fins l’actualitat, no s’ha descrit l’existència d’una via específica de senyals
d’oxidació. Mtl1 es un membre de la via CWI (Cell Wall Integrity: via d’integritat cel.lular) que
funciona com un sensor transmembrana que dectecta l’estrès oxidatiu.
En aquest estudi es demostra que Mtl1 es esencial en el procés de senyalització de
l’estrès oxidatiu i quiescència cap a la via CWI i cap al factor general de resposta a estrès
Msn2/Msn4. En aquest últim cas, la senyalització es duu a terme a través de Rom2 i Rho1 (i
probablement de Pkc1) cap a l’inhibició de les funcions Tor1 i Ras2.
La funció de Mtl1 es necessaria per: i) la repressió de la transcripció de gens
ribosomals, ii) l’inducció transcripcional Msn2/Msn4 i iii) l’activació la via CWI en resposta a
estrès oxidatiu i dejú de glucosa.
En la segona part d’aquesta tesi, es demostra que el domini citoplasmàtic de Mtl1
interacciona físicament amb Rom2, la GEF (GTP Exchange Factor: factor intercanviador de
GTP). Les nostres dades suggereixen que l’activitat de Slt2 es important per la
supervivència en condicions de quiescència. No obstant, Msn2/Msn4 contribueixen de
manera significativa a la supervivència cel.lular davant condicions oxidatives. A més a més,
l’absència de TOR1 o RAS2 es suficient com per induir l’activació de Slt2 de manera
independent de Mtl1, en les condiciones d’estrès anomenades prèviament. Tot això
suggereix que entre CWI, TOR i RAS-cAMP s’estableixen un seguit de reaccions creuades
encaminades a asegurar que les cèl.lules siguin capaces d’adaptar el seu creixement i la
seva maquinaria metabòlica adecuadament.
Mtl1 es N-glicosila i es O-manosila, principalment per la manosil transferasa Pmt2.
Mtl1 es localitza preferentment i de manera homogènia en la perifèria cel.lular, gemma,
septe i extrem apical del shmoo. La manosilació de Mtl1 es important per la localització de
Mtl1 de manera regular en la perifèria i en l’extrem apical del shmoo. La O-manosilació
catalizada per Pmt2 en general, i en particular la O-manosilació de Mtl1, poseeixen una gran
relevancia en: a) la resposta a estrès oxidatiu; b) davant el bloqueix de la via TOR; i c) en
l’extensió cronològica de la vida.<br>Saccharomyces cerevisiae es un microorganismo eucariota que se utiliza como
modelo de estudio de las vías de transducción de señal implicadas en la respuesta a estrés
oxidativo. Hasta el momento no se ha descrito la existencia de una ruta específica de
señales de oxidación. Mtl1 es un miembro de la ruta CWI (Cell Wall Integrity: vía de
integridad celular) que funciona como un sensor transmembrana que detecta el estrés
oxidativo.
En el presente estudio se demuestra que Mtl1 es esencial en el proceso de
señalización del estrés oxidativo y quiescencia hacia la ruta CWI y hacia el factor general de
respuesta a estrés Msn2/Msn4. En este último caso la señalización ocurre a través de Rom2
y Rho1 (y probablemente también a través de Pkc1) hacia la inhibición de las funciones
Tor1 y Ras2.
La función Mtl1 se requiere para: i) la represión de la trascripción de genes
ribosomales, ii) la inducción del factor transcripcional Msn2/Msn4 y iii) activar la ruta CWI en
respuesta a estrés oxidativo y ayuno de glucosa.
En la segunda parte de la presente tesis se muestra que el dominio citoplasmático de
Mtl1 interacciona físicamente con Rom2, la GEF (GTP Exchange Factor: factor
intercambiador de GTP). Nuestros datos sugieren que la actividad Slt2 es importante para la
supervivencia en condiciones de quiescencia. Sin embargo, Msn2/Msn4 contribuyen de
manera más significativa a la supervivencia celular frente a condiciones oxidativas. Además,
la ausencia de TOR1 o RAS2 es suficiente como para inducir la activación de Slt2 de
manera independiente de Mtl1, en las condiciones de estrés mencionadas anteriormente.
Todo ello sugiere que entre CWI, TOR y RAS-cAMP se establecen una serie de reacciones
cruzadas encaminadas a asegurar que las células sean capaces de adaptar el crecimiento y
su maquinaria metabólica de manera adecuada.
Mtl1 se N-glicosila y se O-manosila, principalmente por la manosil transferasa Pmt2.
Mtl1 se localiza preferentemente y de manera homogénea en la periferia celular, yema,
septo y en la punta del shmoo. La manosilación de Mtl1 es importante para la localización de
Mtl1 de manera regular en la periferia y en la punta del shmoo. La O-manosilación
catalizada por Pmt2 en general, y en particular la O-manosilación de Mtl1, poseen una gran
relevancia en: a) la respuesta a estrés oxidativo; b) frente al bloqueo en la ruta TOR; y c) la
extensión cronológica de la vida.<br>The eukaryotic microorganism Saccharomyces cerevisiae serves as a model system
in which to study the signal transduction pathways involved in the oxidative stress response.
Up to date, there is no evidence of any MAPK cascade which is specific to oxidative signals.
Mtl1 is a member of the CWI pathway, which functions as a cell wall sensor for oxidative
stress.
In the present study, we propose an essential role for Mtl1 in signalling oxidative
stress and quiescence to the CWI pathway and to the general stress response through the
inhibition of either Tor1 or Ras2 functions.
The Mtl1 function is required i) to induce ribosomal gene repression, ii) to induce the
general stress response driven by the transcription factor Msn2/Msn4, and iii) to activate the
CWI pathway in response to both oxidative stress and glucose starvation. The signalling from
Mtl1 to Tor1 and/or Ras2 inhibition under these conditions occurs through Rom2 and Rho1,
and probably through Pkc1, at least that signal which target is the ribosomal gene
expression.
We demonstrate that the Mtl1 cytoplasmic domain physically interacts with the GEF
Rom2. Our data indicate that Slt2 activity is really essential in terms of cell survival in
quiescent conditions. However, in response to oxidative stress the contribution of
Msn2/Msn4 function is more significant. In addition, we demonstrate that deletion of either
TOR1 or RAS2 is sufficient to activate Slt2 upon the above mentioned stress conditions,
independently on Mtl1. These data suggested that CWI, TOR and Ras-cAMP provide diverse
cross talks in order to assure the cells to appropriately adapt metabolism and growth.
We demonstrate that Mtl1 is N-glycosylated and highly O-mannosylated mostly by
Pmt2 protein O-mannosyltransferase. Mtl1 localises to the cell periphery, the bud, the
septum, and to the tip of the shmoo. Mtl1 O-mannosylation confers its proper localisation.
We provide evidence for the importance of protein O-mannosylation in oxidative stress
response, through at least Mtl1. This is the first report suggesting a role of protein Omannosylation
in cell survival upon TOR blockage. Mtl1 O-mannosylation by Pmt2 is
required to elicit cellular responses to TOR inhibition. Both Pmt2 and Mtl1 play positive roles
in the chronological life span.
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Wu, Hui-Chen. "Molecular bases of the heat shock response in plants : identification of elements involved in HS transduction pathway and in the cross talk between HS and oxidative stress." Thesis, Montpellier 2, 2010. http://www.theses.fr/2010MON20125.
Full textAbstract:
Les plantes n'échappent pas à leur lieu de vie, elles doivent en permanence adapter leurs processus physiologiques pour répondre aux variations de leurs conditions environnementales. Durant ma thèse, j'ai étudié deux stress affectant le développement des plantes, les stress thermique (HS) et oxydant (OS), en ciblant des éléments clé de ces phénomènes (les protéines de choc thermique Hsp et Thiorédoxines TRX) afin d'apporter des éléments de réponse quant à l'interconnexion de ces stress et leur importance pour la plante.En utilisant le riz et le soja comme modèles, je montre que le HS suit une « signature Ca2+ » en provoquant une entrée de calcium de l'apoplaste vers le cytosol, assurant ainsi une rigidité à la paroi cellulaire et une cascade de signaux. J'identifie aussi une Pectine Methylesterase nécessaire au remodelage de la paroi cellulaire et à l'intégrité de la membrane. J'ai aussi recherché comment la plante perçoit les changements de température et transmet ce signal vers des effecteurs. Par des analyses d'expression de gènes, je montre qu'une CaM bien spécifique coordonne la réponse au HS, qui se traduit par l'expression spécifique de certaines petites Hsp nucléaires et cytosoliques.Je réalise enfin une étude moléculaire de TDX, une TRX suspectée d'agir dans la réponse au HS. Je montre que TDX interagit avec des Hsp70 de type cytosoliques/nucléaires de façon redox dépendante, que les stress HS et OS induisent une relocalisation nucléaire de TDX. Je montre enfin que TDX est essentielle pour la thermotolérance acquise et la transduction du signal oxydant. Ces résultats sont discutés et des modèles de transduction des signaux entre HS et OS sont proposés<br>While being unable to escape their lands, plants are continuously submitted to the modifications of their environment, and need to adjust proper physiological processes in response to various stimuli. During this work, I devoted my studies on two major stresses affecting plant development, heat shock (HS) and oxidative stresses (OS), focusing on key elements in these pathways (HS chaperons and HS-related thioredoxins) in order to bring news elements of knowledge and interconnexion of these pathways.Using rice and soybean as mono- and dicotyledonous plant systems, I show how HS leads to calcium release from plant cell apoplast to the cytosol in a typical calcium signature, conferring cell wall rigidity and enhancing HS signaling pathway. I also identify Pectin Methylesterase as required in this pathway for cell wall remodeling and plasma membrane integrity. I further investigate how plant sense temperature increases and how they transmit the HS signal to downstream elements. Using systematic analyses of Calmodulin (CaM) and small heat shock protein (sHsp) gene expression, I identify one CaM as a coordinator of HS response, which I characterize as involving specific cytosolic/nuclear isoforms of the sHsp family.I latter perform the molecular analysis of TDX, a Thioredoxin suspected to be involved in heat shock response. I show that TDX interacts with cytosolic/nuclear members of the Hsp70 family in a redox dependent manner, both HS and OS inducing its nuclear relocation, and that TDX is required for both acquired thermotolerance and OS signaling.I finally discuss the data brought by this work and propose models with cross-talks between HS and oxidative stress signaling
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Gok, Abdulkerim. "Degradation Pathway Models of Poly(ethylene-terephthalate) Under Accelerated Weathering Exposures." Case Western Reserve University School of Graduate Studies / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=case1449244610.
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Almazroue, Hanadi Abdulaali. "Identification, Cloning, and Expression of Tobacco Responsive to Dehydration like Protein (RD22), SBIP-355 and Its Role in SABP2 Mediated SA Pathway in Plant Defense." Digital Commons @ East Tennessee State University, 2014. https://dc.etsu.edu/etd/2456.
Full textAbstract:
Abscisic acid (ABA) induces RD22, responsive to dehydration stress gene. Salicylic acid (SA) has been the focus of research due to its role against pathogens and abiotic stress. Interaction between ABA and SA signaling pathways is still poorly understood. SA-Binding Protein 2 (SABP2) converts methyl salicylate to SA. An attempt was made to identify proteins that interact with SABP2 using a yeast 2-hybrid screening. Several interactors were identified. One of them, SA-Binding Protein 2 Interacting Protein-355 (SBIP-355), showed high homology to RD22. Bioinformatic approaches showed that SBIP-355 contains a BURP domain. Phylogenetic analysis reveals that SBIP-355 clustered into the clade of RD22-like proteins. Thus, SBIP-355 gene might be a stress-inducible gene and encodes a dehydration-responsive protein, which is important for the stress tolerance of tobacco. The complementary DNA (cDNA) of tobacco SBIP-355 was cloned into pDEST-17 vector and then expressed in E. coli to detect the expression of SBIP-355 protein.
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Amoedo, Machi Hugo [Verfasser], Gerhard [Akademischer Betreuer] Braus, Gerhard [Gutachter] Braus, Stefanie [Gutachter] Pöggeler, and Ralph [Gutachter] Kehlenbach. "The Aspergillus fumigatus Vap-Vip methyltransferase pathway modulates stress response, secondary metabolism and azole resistance / Hugo Amoedo Machi ; Gutachter: Gerhard Braus, Stefanie Pöggeler, Ralph Kehlenbach ; Betreuer: Gerhard Braus." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2019. http://d-nb.info/1190353644/34.
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