Thèses sur le sujet « Cl11 »
Pour voir les autres types de publications sur ce sujet consultez le lien suivant : Cl11.
Créez une référence correcte selon les styles APA, MLA, Chicago, Harvard et plusieurs autres
Consultez les 50 meilleures thèses pour votre recherche sur le sujet « Cl11 ».
À côté de chaque source dans la liste de références il y a un bouton « Ajouter à la bibliographie ». Cliquez sur ce bouton, et nous générerons automatiquement la référence bibliographique pour la source choisie selon votre style de citation préféré : APA, MLA, Harvard, Vancouver, Chicago, etc.
Vous pouvez aussi télécharger le texte intégral de la publication scolaire au format pdf et consulter son résumé en ligne lorsque ces informations sont inclues dans les métadonnées.
Parcourez les thèses sur diverses disciplines et organisez correctement votre bibliographie.
1
Duncan, Peter I. « Characterisation of the murine CLK1 dual-specificity kinase ». Thesis, University of Ottawa (Canada), 1997. http://hdl.handle.net/10393/10402.
Texte intégralRésumé :
Murine Clk1 (also known as Sty) was identified as a dual-specificity kinase capable of phosphorylating serine, threonine and tyrosine residues when expressed in bacteria. Expression of Clk1 is deveolpmentally regulated at the level of RNA expression. Embryonic cells express two mRNAs of similar size, whereas differentiated cells express two additional transcripts of larger size. Little is known about the bochemical and biological activity of Clk1 in mammalian cells. We demonstrate that the two embryonically expressed mRNAs are derived through alternative splicing of a unique exon (exon EB) within the Clk1 precursor mRNA. These mRNAs encode full-length catalytically active Clk1 and a truncated inactive polypeptide, Clk1$\rm\sp{T}.$ Larger incompletely spliced Clk1 transcripts accumulate in differentiated cells. When expressed in mammalian cells Clk1 possesses dual-specificity kinase activity and is capable of forming complexes with other molecules of Clk1 and Clk1$\rm\sp{T}.$ The regions involved in binding map to the amino-terminal non-catalytic domain of Clk1. Phosphorylation sites map to the amino acids encoded by the alternatively splice exon EB. Clk1$\rm\sp{T}$ and catalytic mutants of Clk1 co-localise with splicing factors in intranuclear speckles, whereas catalytically active Clk1 causes a redistribution of these factors within the nucleus. This activity requires the presence of amino acids encoded by exon EB. These results suggest a role for Clk1 and Clk1$\rm\sp{T}$ in the regulation of RNA splicing. Splicing of a Clk1 mini-gene, encompassing exon EB, in vivo is regulated by Clk1 and Clk1$\rm\sp{T}.$ Catalytically active Clk1 stimulates exclusion of EB leading to the production of Clk1$\rm\sp{T}$ mRNA. In contrast, Clk1$\rm\sp{T}$ promotes EB inclusion leading to production of Clk1 mRNA. Two Clk1-related human kinases, hClk2 and hClk3, also exhibit dual-specificity kinase activity and cause the redistribution of nuclear splicing factors. Similar to Clk1$\rm\sp{T},$ the hClk truncated isoforms, hClk2$\rm\sp{T}$ and hClk3, co-localise with splicing factors in nuclear speckles. hClk2 and hClk3 are able to influence the splicing pattern of a murine Clk1 mini-gene in vivo, indicating that they can also regulate precursor mRNA splicing. Taken together these results imply a role for the Clk family of kinases in the regulation of gene expression at the level of RNA processing.
2
Tibbles, Katherine L. « Regulation of Clb1 during meiosis in Saccharomyces cerevisiae ». Thesis, University of Warwick, 2013. http://wrap.warwick.ac.uk/60444/.
Texte intégralRésumé :
Meiosis is a specialised form of cell division in which diploid cells divide to form four non-identical spores containing half the genetic complement of the parent. During this cell division program, much of the usual machinery regulating cell division is put to alternate use to allow the cells to undergo an extra round of division without an intervening phase of DNA synthesis. In particular, the end of the first division, meiosis I, must be regulated differently than the end of the mitotic division. We used the model organism Saccharomyces cerevisiae to determine some of these differences in regulation. The cell division program is driven by the sequential association of cyclins with the CDK (cyclin dependent kinase), leading to waves of kinase activity. Exit from mitosis requires the downregulation of CDK activity, and is coordinated by two signalling networks, the FEAR (Cdc14 Early Anaphase Release) network and the MEN (Mitotic Exit Network). Both networks initiate the release of the phosphatase Cdc14 from its inhibitor, Net1, to counter CDK activity. Exit from meiosis I similarly relies on Cdc14 activity, but is driven only by the FEAR network. Experimental results showed that the phosphorylation state and subcellular localisation of the meiotic cyclin, C1b1, are altered in meiosis I. We investigated this relationship and aimed to determine the kinase responsible. We used modelling techniques to explore several rationales for the specific regulation of C1b1. We examined the functional significance of C1b1 localisation, using localisation mutants, and made an investigation into Cdc14 release in meiosis I.
3
Lange, Jenny. « Neuron-glia interactions in infantile neuronal ceroid lipofuscinosis (CLN1 disease) ». Thesis, King's College London (University of London), 2016. https://kclpure.kcl.ac.uk/portal/en/theses/neuronglia-interactions-in-infantile-neuronal-ceroid-lipofuscinosis-cln1-disease(ceed9c8f-f40d-4796-be7b-e799fe88b431).html.
Texte intégralRésumé :
The neuronal ceroid lipofuscinoses (NCLs) are the most common cause of childhood dementia and are invariably fatal. Early, localised glial activation occurs in all forms of NCL and has been shown to be an accurate predictor of areas in which neuronal loss is most pronounced. Indeed, in a tissue culture model of juvenile NCL, glial cells have been shown to actively contribute to neuronal dysfunction and death. So far the role of glial cells has not been established in other forms of NCL and in order to assess glial function in infantile NCL (INCL, CLN1 disease), a series of different tissue cultures were generated from Ppt1 deficient mice (Ppt1-/- ). These studies revealed that both Ppt1-/-astrocytes and microglia exhibit a more activated phenotype under basal conditions, as well as alterations to their protein expression profile following pharmacological stimulation. Ppt1-/- astrocytes displayed moderately reduced glutamate uptake, as well as changes in lactate release. Perhaps as a consequence of these changes, Ppt1- /-astrocyte survival was severely impaired. In addition the morphological phenotypes of Ppt1-/-neurons were explored, revealing decreased neurite outgrowth, complexity and a reduction in cell body size. Ppt1-/-neuronal cultures contained significantly fewer inhibitory neurons and displayed decreased cell survival after prolonged time in culture. Most importantly, using a co-culture system, the presence of Ppt1-/- glial cells appeared to increase cell death in both WT and Ppt1-/- cultures. Notably, Ppt1-/- microglia appeared to trigger increased Ppt1-/- neuronal death, whereas Ppt1-/- astrocytes also exhibited increased cell death. Ppt1-/- glial cells also affected both wild type and Ppt1-/- neuronal morphology, including further reduced neurite outgrowth. In contrast, wild type glial cells ameliorated some of the morphological defects observed in Ppt1-/- neurons, and this was most apparent when wild type astrocytes where grown in co-cultures with Ppt1-/- neurons. Taken together, these finding present novel evidence for compromised glial function in Cln1 disease, demonstrating that both Ppt1-/- microglia and astrocytes may potentially contribute to the neurodegeneration observed in CLN1 disease. These data highlight the importance of targeting glial cells in the development of therapeutic interventions for CLN1 disease. Furthermore, although sharing some similarities Ppt1-/-glial phenotypes were broadly different from those observed in the tissue culture model of juvenile NCL, suggesting that the role of glia may differ between forms of NCL.
4
Nelvagal, Hemanth Ramesh. « Spinal cord pathology in CLN1 disease : a novel therapeutic target ». Thesis, King's College London (University of London), 2017. https://kclpure.kcl.ac.uk/portal/en/theses/spinal-cord-pathology-in-cln1-disease(8b1dc3ed-dfd9-442d-a427-43ded0d82a6a).html.
Texte intégralRésumé :
The neuronal ceroid lipofuscinoses (NCLs) are a group of up to 14 inherited progressive neurodegenerative lysosomal storage disorders affecting children and young adults. Together, they are the most common pediatric neurodegenerative storage disorders. Symptoms include loss of vision, epileptic seizures and the loss of cognitive and motor function. A lack of any effective therapies means that all forms are fatal. CLN1 disease or Infantile NCL is one of the most rapidly progressing forms of the disease and is caused by a deficiency of the lysosomal enzyme palmitoyl protein thioesterase – 1 (PPT1). Ppt1 deficient (Ppt1-/-) mice recapitulate features of the human disease and have proved to be a useful tool in characterizing disease progression and pathology in the brain. However, these pathological changes fail to fully explain the sensorimotor deficits seen in this mouse model as well as in human CLN1 disease. Along with the limited success of various brain directed therapies, this led us to analyze the spinal cord. Our analysis revealed unexpectedly profound and rapidly progressing disease pathology in the spinal cords of these mice, which occurs earlier than similar events in the brain. This included regional atrophy, neuroinflammation, and significant neuron loss at all levels of the cord as well as novel phenotypes indicating a postnatal developmental delay and significant white matter pathology. Automated gait analysis also showed novel early phenotypes in these mice including an increased dependence on the forelimbs for locomotion. Similar spinal cord pathology was also demonstrated in human INCL autopsy samples as well as in mouse models of the other major forms of NCL. Targeting the spinal cords of Ppt1-/-mice with enzyme replacement therapy (ERT) and gene therapy significantly improved disease pathology, motor function and lifespan in these mice, demonstrating the clinical significance of spinal cord pathology in these mice. Furthermore, combining intracranial and intrathecal gene therapy showed a synergistic effect, showing the greatest improvements for any CLN1 disease therapy to date. Taken together, these findings highlight the spinal cord as not only being significantly affected in CLN1 disease, but also as a novel and effective therapeutic target.
5
GARSUAULT, SOPHIE. « Degradation et fonction de cln1 identification de sequences en cis responsables de la degradation et de la fonction de cln1, une cycline g1 de saccharomyces cerevisiae ». Paris 6, 1998. http://www.theses.fr/1998PA066496.
Texte intégralRésumé :
Ces travaux ont consiste a identifier des signaux en cis necessaires a la degradation et a la fonction de cln1, une cycline g1 de s. Cerevisiae. Les cyclines g1 sont au coeur de la regulation du cycle cellulaire eucaryote, controlant la transition entre proliferation et differenciation. Chez la levure saccharomyces cerevisiae, cln1 et cln2 s'accumulent dans la cellule en phase g1, declenchent l'engagement irreversible dans un nouveau cycle en activant la kinase cdc28, puis disparaissent brutalement en raison de leur haute instabilite. L'association a cdc28 se fait par l'intermediaire d'une region tres conservee chez toutes les cyclines, la cyclin-box, situee dans la region n-terminale, et correspondant au premier domaine de repliement defini par cristallographie (cyclin-fold). Nous montrons que la derniere helice du second cyclin-fold joue un role determinant dans la fonction de cln1. Peu d'exemples de sequences situees hors de la cyclin-box et contribuant a la fonctionnalite de la cycline ont ete rapportes jusqu'a maintenant. Des deletions carboxy-terminales progressives ont permis de montrer que les determinants de la degradation sont repartis dans la region c-terminale de cln1, et sont similaires a ceux identifies chez cln2 et cln3. Les sites de phosphorylation semblent etre les determinants majeurs, mais la region c-terminale contient d'autres signaux non identifies, necessaires pour une degradation rapide. Les determinants de proteolyse contenus dans la region carboxy-terminale de cln1 ne fonctionnent pas comme des signaux autonomes de degradation, contrairement a ce qui a ete observe chez cln2 et cln3. La mutation ponctuelle dans la cyclin-box d'un residu hautement conserve, l'arginine 72, est suffisante pour abolir la fixation a cdc28, et a stabiliser significativement la cycline. Ainsi, la fixation de la cycline a la kinase est un pre-requis a sa degradation rapide. Nous montrons par ailleurs que les cyclines libres - non fixees a cdc28 - sont reconnues par une voie de degradation operant plus lentement, ne reconnaissant probablement pas les memes signaux que la voie de degradation des cyclines associees a cdc28.
6
Zou, Meilin. « Haplotypes of cleistogamous flowering gene cly1 and association with temperature stress ». Thesis, Zou, Meilin (2017) Haplotypes of cleistogamous flowering gene cly1 and association with temperature stress. Honours thesis, Murdoch University, 2017. https://researchrepository.murdoch.edu.au/id/eprint/37942/.
Texte intégralRésumé :
Cleistogamy refers to a type of sexual breeding system, a floret type with closed flowers. Cleistogamous flower sheds its pollen before flower opening, so this behavior leads to a great mass of autogamy. Furthermore, the cleistogamy has been found and presented widely in the angiosperm. In recent years, the cleistogamy has been regarded as an important reproductive system in various plant taxa and already attracted widespread attention in the world. However, the understanding of the molecular mechanism of the cleistogamous trait was very limited. It is valuable to explore the cleistogamy gene cly1 in barley.
Cly1 has been cloned, and two SNPs within the open reading frame region were identified to be associated with floral types. New markers were developed to genotype 672 barley accessions. Among them, the floral types of 436 lines were investigated. A total of 45 novel lines were identified as the genotype of the two markers could not explain their phenotypes. Five novel lines which showed non-cleistogamous types in the field but had cleistogamous genotype were sequenced. Thirteen SNPs were detected among the gene region. But no SNPs were associated with non-cleistogamy. Promoter region, methylation, and miR172 gene sequence need to be investigated in the future.
The purposes of this article are to investigate the occurrence mechanism of cleistogamy, particularly on genetic aspects in barley. Temperature stress including frost stress and heat stress is one of the biggest obstacles to crop production. So the impacts of cleistogamy against temperature stress were discussed in the present study. Grain fertility rate was investigated in the four field trials. Katanning experienced frost stress in 2016 with the lowest temperature of -3.4°C. Grain fertility percentage was as low as 85% in Katanning, while in other three trials, the grain fertility percentage ranged from 92% to 96%. The correlation coefficient of grain fertility between cleistogamous type and non-cleistogamous type was analyzed. No significant difference was detected in the four trials between the two floral types.
7
Aigner, Christian [Verfasser], et Franz [Akademischer Betreuer] Bracher. « Entwicklung neuer CLK1-Inhibitoren mit antiviraler Aktivität / Christian Aigner ; Betreuer : Franz Bracher ». München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2017. http://d-nb.info/1138566160/34.
Texte intégral
8
Durkan, Anne Maria. « The expression of CX₃CL1 (fractalkine) in renal tubular epithelial cells and the regulation of CX₃CL1 by stimulation of the thromboxane prostanoid receptor ». Thesis, University of Edinburgh, 2012. http://hdl.handle.net/1842/24546.
Texte intégralRésumé :
Most renal diseases have a common end inflammatory pathway, which is associated with a leukocytic infiltrate. Chemokines are small proteins that are responsible for the chemoattraction of leukocytes into areas of injury or insult. CX3CL1, also known as fractalkine, exists as a transmembrane protein as well as a soluble protein. It acts as a cell adhesion molecule in addition to its chemoattractant properties. This thesis firstly examines the distribution of CX3CL1 in renal tubular epithelial cells (RTEC) and in the second part of the thesis the regulation of CX3CL1 by stimulation of the thromboxane prostanoid (TP) receptor is examined. The localisation of CX3CL1 was initially demonstrated primarily on the apical surface of tubular epithelial cells in human renal biopsy specimens with histological diagnoses of acute tubular necrosis and acute allograft rejection. A cell model was then developed in MDCK cells to examine the distribution more closely. There are a limited number of mechanisms potentially responsible for the trafficking of CX3CL1 to the apical membrane and it was established that N-glycosylation of CX3CL1 is required for its presence on the apical membrane of RTEC. The mobility of CX3CL1 within the cell membrane was next assessed and it was shown to be relatively immobile. We hypothesized that this would promote cell adhesion and indeed further experiments confirmed that CX3CL1 in RTEC does promote adhesion of cells bearing the cognate receptor. Given that CX3CL1 and thromboxane A2 are both found in similar inflammatory conditions, are both present early in the inflammatory process and that stimulation of the TP receptor has been shown to regulate other chemokines, we next evaluated the effect of stimulation of TP on CX3CL1. We found that both total and surface cellular levels of CX3CL1 were reduced following stimulation of TP. A maximal nadir was present after 30-60 minutes and the levels returned to baseline by 4 hours. The mechanism for the loss of CX3CL1 was then assessed. CX3CL1 is known to recycle between the cell surface and an internal compartment. No effect of TP stimulation was seen on the endocytosis or exocytosis of CX3CL1. Stimulation of TP was however, shown to stimulate tumour necrosis factor-a converting enzyme (TACE) via ERK phosphorylation. TACE inducibly cleaves CX3CLI, releasing the soluble chemokine. TACE siRNA was used to knock down TACE gene expression and this prevented the loss of cellular CX3CL1, confirming that TP stimulation induces TACE cleavage of CX3CL1. T he results of further experiments are discussed in the discussion chapter.
9
Hedden, J. J. « The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing ». Thesis, University College London (University of London), 2012. http://discovery.ucl.ac.uk/1369193/.
Texte intégralRésumé :
Eukaryotic transcripts require a number of complex cotranscriptional modifications and processing events before translation to protein. Clp1 and Pcf11 are subunits of cleavage factor IA (CFIA), an essential component of the Saccharomyces cerevisiae pre-mRNA 3’-end processing machinery. The crystal structure of a Clp1-Pcf11 complex was determined previously and revealed the binding of ATP to a highly-conserved P-loop motif and a tight Pcf11-Clp1 interaction facilitated by a number of highly-conserved Pcf11 residues. Nonetheless, the biological function of both Clp1-ATP binding and the Pcf11-Clp1 interaction was not well understood. The work in this thesis combines an in vitro and in vivo investigation of the Clp-ATP and Clp-Pcf11 interactions in an effort to understand the function of these factors in transcription and pre-mRNA 3’-end processing. It is demonstrated that the interaction of ATP and Pcf11 with Clp1 are linked events: Loss of Clp1-ATP binding results in the abrogation of the Pcf11-Clp1 interaction and leads to Clp1 instability in vitro, and similarly, mutations that directly uncouple the Pcf11-Clp1 interaction also disrupt Clp1-ATP binding and cause Clp1 instability in vitro. An in vivo mutational analysis in S. cerevisiae revealed that both Clp1-ATP binding and the Pcf11-Clp1 interaction are essential for yeast survival. Further cell and immunoprecipitation studies demonstrated that one essential function of Clp1 is as a chaperone of Pcf11, and RT-qPCR analysis of mRNA from a sample set of yeast genes points to a role for these proteins in transcription and transcription termination rather than in poly(A) site selection.
10
Isosomppi, Juha. « Molecular and cell biology of infantile (CLN1) and varaint late infantile (CLN5) neuronal ceroid lipofuscinoses ». Helsinki : University of Helsinki, 2003. http://ethesis.helsinki.fi/julkaisut/laa/haart/vk/isosomppi/.
Texte intégral
11
Pescheteau, Clémentine. « Conception et synthèse d’inhibiteurs duaux de DYRK1A et CLK1, kinases impliquées dans la maladie d’Alzheimer ». Electronic Thesis or Diss., Orléans, 2021. http://www.theses.fr/2021ORLE3203.
Texte intégralRésumé :
La maladie d’Alzheimer est la forme de démence la plus répandue dans le monde et à ce jour, aucun traitement efficace n’existe malgré de nombreuses recherches en cours. De nouvelles stratégies thérapeutiques ont émergé, ciblant des protéines kinases impliquées dans les processus de neurodégénération et de neuroinflammation. DYRK1A et CLK1 ont notamment été identifiées comme des cibles intéressantes pour lutter contre des pathologies du système nerveux central, et sont particulièrement impliquées dans la maladie d’Alzheimer. Afin de concevoir des inhibiteurs puissants et sélectifs de ces deux kinases, nous avons synthétisé des molécules hétérocycliques originales constituées de noyaux aromatiques de type [5-5] et [6-5]. Nous avons d’abord poursuivi les études du laboratoire sur les imidazo[2,1-b][1,3,4]thiadiazoles, puis créé une librairie de composés originaux qui a révélé des activités intéressantes contre DYRK1A et CLK1. Nous avons optimisé les voies d’accès aux inhibiteurs duaux les plus prometteurs, et leurs évaluations biologiques menées par nos collaborateurs ont permis d’identifier un hit moléculaire qui sera engagé dans des études in vivo. Nous avons ensuite exploré d’autres séries moléculaires grâce à la modulation du scaffold de nos composés. L’accès au bicycle [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole a été étudié pour concevoir des analogues de nos structures, et une méthodologie de synthèse efficace d’imidazo[1,2-d][1,2,4]thiadiazoles a été développée pour accéder à une diversité de molécules à haut potentiel de valorisation. D’autre part, le scaffold [1,2,4]triazolo[4,3-b]pyridazine de type [6-5] a été employé, et les pharmacomodulations de ses substituants nous ont permis d’affiner les relations de structure-activité des inhibiteurs obtenus. Des structures innovantes ont également été développées, comme des macrocycles inhibant sélectivement nos deux kinases d’intérêtAlzheimer's disease is the most common form of dementia worldwide. To date, no effective treatment exists despite many ongoing research projects. New therapeutic strategies have emerged, targeting protein kinases involved in neurodegeneration and neuroinflammation, processes leading to diseases of the central nervous system. DYRK1A and CLK1 have been identified as interesting targets against some neuropathologies, and are particularly involved in Alzheimer's disease. In order to design powerful and selective inhibitors of these two kinases, we have synthesized original heterocyclic molecules with aromatic [5-5] and [6-5] core rings. We first continued our team’s studies on imidazo[2,1-b][1,3,4]thiadiazoles and we created a library of original compounds which revealed interesting activities against DYRK1A and CLK1. We optimized the access routes to the most promising dual inhibitors, and their biological evaluations carried out by our collaborators allowed us to identify a molecular hit that will be studied in vivo. We then explored other molecular series by modulating the scaffold of our compounds. Access to the bicycle [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole has been studied to design analogs of our structures. An efficient synthetic methodology of imidazo[1,2-d][1,2,4]thiadiazoles was then developed to reach a variety of molecules with high value potential. Furthermore, the [1,2,4]triazolo[4,3-b]pyridazine scaffold was used, and pharmacomodulations of its substituents allowed us to refine the structure-activity relationships of the designed inhibitors. Finally, innovative structures were developed, such as selective inhibitory macrocycles of our two kinases of interest
12
Sako, Yukiya. « Development of an orally available inhibitor of CLK1 for skipping a mutated dystrophin exon in Duchenne muscular dystrophy ». Kyoto University, 2017. http://hdl.handle.net/2433/226771.
Texte intégral
13
Lagrange, Thierry. « Evolution, structure et expression du gène nucléaire rpl21 codant pour la protéine ribosomique plastidiale cL21 d'épinard ». Grenoble 1, 1993. http://www.theses.fr/1993GRE10152.
Texte intégralRésumé :
Le ribosome plastidial presente une synthese accrue dans les cellules photosynthetiques, et une expression basale dans les autres types cellulaires. C'est donc afin de comprendre par quels mecanismes, les genes nucleaires qui codent pour des proteines ribosomiques (p. R. ) plastidiales repondent a cette demande traductionnelle tres variable que nous avons isole chez spinacia oleracea un gene nucleaire modele, le gene rpl21. Nous avons tout d'abord caracterise un adnc codant pour une p. R. Plastidiale. La sequence primaire deduite de cette proteine presente une homologie avec la sequence de la p. R. L21 d'e. Coli. Cette proteine a ete nommee cl21. Les implications evolutives sur l'origine de la proteine cl21 d'epinard, que ces resultats suggerent, sont discutees. Dans la suite de ce travail, nous avons montre que la proteine cl21 est codee par un unique gene nucleaire, nomme rpl21. Nous avons ensuite clone et sequence la totalite de la region codante ainsi que la region 5 regulatrice du gene rpl21. L'analyse de la repartition des messagers rpl21 par la technique de northern nous a permis d'observer une accumulation preferentielle de ces messagers dans les feuilles par rapport aux racines. Cette expression variable suivant les organes de la plante a pu etre associee a la presence de transcrits alternatifs, nommes p1 et p2. Des experiences d'expression transitoire suggerent que ces deux transcrits proviennent chacun d'une initiation de transcription. L'adaptation de l'expression des genes nucleaires de proteines ribosomiques plastidiales aux contraintes fonctionnelles du systeme vegetal est discutee. Dans la derniere partie de ce travail, l'analyse systematique de l'organisation du promoteur du gene rpl21 nous a permis de mettre en evidence plusieurs elements cis qui sont impliques dans l'activation transcriptionnelle de ce gene. Les roles potentiels de ces elements dans l'expression de ces genes et dans le developpement du chloroplaste sont discutes
14
Hanemian, Mathieu. « Rôle de la protéine CLV1 dans la sensibilité d'Arabidopsis thaliana à la bactérie phytopathogène Ralstonia solanacearum ». Toulouse 3, 2012. http://thesesups.ups-tlse.fr/2754/.
Texte intégralRésumé :
Les mécanismes moléculaires associés au développement de la maladie causée par la bactérie phytopathogène R. Solanacearum sont relativement peu connus à ce jour. La recherche de mutants d'Arabidopsis incapables de développer des symptômes de flétrissement associée à la maladie a mené à l'identification de gènes dit de sensibilité. Les produits de ces gènes peuvent être des cibles de facteurs de virulence aussi bien que des composantes végétales requises pour la " fitness " de la bactérie. Le gène CLAVATA1 (CLV1), très étudié dans le contexte du développement, code pour une protéine appartenant à la famille des récepteurs kinase possédant un domaine extracellulaire riche en leucine. Cette protéine joue un rôle crucial dans le maintien d'une population de cellules souches au niveau du méristème caulinaire donnant naissance à toute la partie aérienne de la plante. La mutation clv1 entraîne une résistance accrue à R. Solanacearum associée à une réduction de la croissance bactérienne in planta. Mes travaux de thèse ont consisté à élucider les mécanismes sous-tendant la résistance accrue conférée par la mutation du gène CLV1 en utilisant différents types d'approche (génétique, moléculaire et transcriptomique). Nous avons été capables de démontrer l'implication de facteurs de transcription de la famille de gènes NF-YA, eux-mêmes contrôlés par les microARNs miR169, dans la résistance accrue des mutants clv1. Ces résultats démontrent que la protéine CLV1 est une composante requise pour l'établissement de la maladie causée par R. Solanacearum. Mes travaux de thèse mettent en lumière une nouvelle fonction de cette protéine et illustrent la grande diversité des rôles biologiques de protéines de type récepteur-kinaseThe molecular mechanisms associated to disease development caused by the phytopathogenic bacteria Ralstonia solanacearum are poorly understood. Search for mutants altered in their response to the pathogen led to the identification of some susceptibility genes including targets of virulence factors as well as plant components required for pathogen fitness. The CLAVATA1 (CLV1) gene, extensively studied for its role in plant development, encodes a receptor-like kinase with a leucin-rich repeat extracellular domain. This protein plays indeed a key role in maintaining a pool of stem cell within the shoot apical meristem. The clv1 mutation leads to an increased resistance to R. Solanacearum, associated with a decrease of in planta bacterial growth. The aim of my PhD work was the understanding of the mechanisms underlying the increased resistance conferred by the clv1 mutation in using different approach (molecular, genetic and transcriptomic). We have been able to demonstrate the implication of the NF-YA transcription factor family, controlled by microRNA miR169, in the increased resistance of these mutants. These results demonstrate that the CLV1 protein is a required component for the establishment of the disease caused by R. Solanacearum. And illustrate the wide diversity of functions fulfilled by receptors kinases
15
Esvan, Yannick. « Conception et synthèse de nouveaux composés hétéroaromatiques inhibiteurs potentiels de kinases ». Thesis, Clermont-Ferrand 2, 2016. http://www.theses.fr/2016CLF22743.
Texte intégralRésumé :
Depuis la mise en évidence de l’existence des protéines kinases vers la fin des années 1950 cette famille d’enzymes s’est vu attribuer d’importants rôles dans divers mécanismes pathologiques notamment dans des processus de cancérisations. Plus récemment ces enzymes ont été identifiées comme potentiellement impliquées dans d’autres types de maladies telles que les maladies neurodégénératives.Deux projets de recherche seront présentés. Le premier projet expose la conception et la synthèse de nouveaux composés tricycliques de la famille des pyrido[3,4-g]quinazolines. Les propriétés inhibitrices de kinases des premiers dérivés ont été évaluées sur un panel de cinq kinases (CDK5, CK1, GSK3, CLK1 and DYRK1A) connues pour leurs implications dans la maladie d’Alzheimer. L’intérêt de ces nouveaux squelettes tricycliques comme inhibiteurs de kinases a été validé par des activités inhibitrices nanomolaire à l’encontre des kinases DYRK1A et CLK1. D’autre part l’obtention de structures co-crystallographiques d’interaction de deux dérivés avec le site ATP de la kinase CLK1 a permis de rationnaliser la substitution du motif pyrido[3,4-g]quinazoline. Le second projet présente le développement d’un nouveau dérivé de la staurosporine aglycone (K252c) dans lequel la partie lactame a été remplacée par un noyau pyrazole. Une étude préliminaire des propriétés biologiques de l’indolopyrazolocarbazole obtenu met en avant une cytotoxicité, du même ordre de grandeur que K252c, contre les lignées cellulaires K562 (leucémie humaine) et HCT116 (carcinome du colon). En revanche, le composé chef de file s’est révélé être un faible inhibiteur de cibles connues de K252c, les isoformes α and γ de la protéine kinase C et présente un bon potentiel inhibiteur des kinases Pim 1-3. Ce nouveau chemotype pourrait être un inhibiteur de kinases prometteurIn 1950’s protein kinases were found to play a critical role in cell signaling, rising strong research potential for this enzyme family. Initially investigated for their implications in cancerogenesis they were more recently found to be involved in a wide variety of diseases including neurodegenerative pathologies. Herein will be presented two research projects that offer bright new perspectives for the inhibition of kinases involved whether in neurodegenerative diseases or cancers.First, the design and synthesis of new pyrido[3,4-g]quinazoline derivatives will be described as well as their protein kinase inhibitory potencies toward five CMGC family members (CDK5, CK1, GSK3, CLK1 and DYRK1A) that are known to play a potential role in Alzheimer’s disease. The interest for this original tricyclic heteroaromatic scaffold as modulators of CLK1/ DYRK1A activity was validated by nanomolar potencies. CLK1 co-crystal structures with two inhibitors revealed the binding mode of these compounds within the ATP-binding pocket and led to the synthesis of new diversely substituted pyrido[3,4-g]quinazolines.Then the synthesis of a new derivative of the staurosporine aglycon (K252c), in which the lactam ring was replaced by a pyrazole moiety, will be depicted. The resulting indolopyrazolocarbazole inhibited Pim isoforms 1–3 whereas it did not impair the activity of two known targets of K252c, protein kinase C isoforms α and γ . The lead compound exhibited same cytotoxic activity as K252c toward both human leukemia and colon carcinoma cell lines (K562 and HCT116), strongly suggesting that this new scaffold deserves further investigations for treatment of malignancies associated with kinases activities
16
Dikfidan, Aytac [Verfasser], et Ilme [Akademischer Betreuer] Schlichting. « Structural and functional characterization of Clp1, a eukaryotic RNA‐specific polynucleotide kinase / Aytac Dikfidan ; Betreuer : Ilme Schlichting ». Heidelberg : Universitätsbibliothek Heidelberg, 2013. http://d-nb.info/1177809524/34.
Texte intégral
17
Place, Matthieu. « Méthodologies de synthèses d'hétérocycles bicycliques (6-5) et (5-5). Application à la conception d'inhibiteurs de kinases impliquées en oncologie et dans les maladies du système nerveux central ». Thesis, Orléans, 2017. http://www.theses.fr/2017ORLE2056.
Texte intégralRésumé :
Depuis le début des années 2000, la connaissance précise du kinome a entraîné l’émergence de nouvelles stratégies thérapeutiques ciblant des protéines kinases impliquées dans de nombreuses pathologies en oncologie et dans les maladies du système nerveux central. Afin de cibler les kinases d’intérêts identifiées au sein de ces travaux, nous avons effectué, dans une démarche orientée vers la diversité moléculaire, la synthèse de nouveaux hétérocycliques a fort potentiel de valorisation. Nous nous sommes appuyé sur la création et la fonctionnalisation de bicycles à 5 ou 6chaînons de type [6-5] ou [5-5], ces espèces chimiques représentant la voie d’accès à des inhibiteurs compétitifs du substrat naturel des kinases, l’ATP. Nous avons dans un premier temps travaillé autour des imidazo[1,2-b]pyridazines puis des[1,2,4] triazolo[4,3-b]pyridazines, scaffold plus original, pour concevoir des inhibiteurs plus actifs et spécifiques de la kinase HASPIN, nouvelle cible prometteuse en oncologie.Nous avons ensuite poursuivi les études précédentes du laboratoire sur les imidazo[2,1-b][1,3,4]thiadiazoles. Nous basant sur une méthodologie de synthèse bien développée, nous avons créé une librairie de composés dirigés contre les kinases DYRK1A et CLK1 impliquées dans les processus de neuro dégénération, notamment dans la maladie d’Alzheimer. Ainsi, au travers d’analogues des imidazothiadiazoles originaux, nous avons proposé des méthodologies de synthèses de ces nouveaux hétérocycles permettant des pharmaco modulations aisées.Ces divers projets de chimie médicinale ont pu être entrepris de façon à améliorer les connaissances des relations structure-activité, et concevoir de nouveaux inhibiteurs puissants des kinases HASPIN, DYRK1A etCLK1Since the early 2000s, precise knowledge of kinome has induced the emergence of novel therapies targeting kinases involved in several kinds of pathologies in oncology and nervous central systems disorders.In order to target original kinases of interest identified in this work, we have developed diversity-oriented synthesis to create new high-valuable heterocycles. We have focused our efforts on the design and functionalization of [6-5] or [5-5] fused ring bicycles. Those chemical species representing a great pathway tocreate competitive inhibitors of ATP; the natural substrate of kinases.First-of-all, we have worked on imidazo[1,2-b]pyridazines and then on [1,2,4]triazolo[4,3-b]pyridazinesscaffolds, to create more active and selective HASPIN kinase inhibitors, a new hot-target in oncology.Then, we have pursued previous laboratory studies on imidazo[2,1-b][1,3,4]thiadiazoles. Based on a well-built methodology, we have synthesized severals DYRK1A and CLK1 kinases inhibitors involved in neurodegenerative disorders, as Alzheimer’s disease. Thus, through original imidazothiadiazoles analogues,we have proposed synthetic methodologies to design these novel heterocycles allowding esay pharmacomodulations.These medicinal chemistry projects have been undertaken to improved knowledge of structure-activityrelashionship, and providing novel strong inhibitors of HASPIN, DYRK1A or CLK1 kinases
18
Faist, Benjamin [Verfasser], et J. [Akademischer Betreuer] Kämper. « Mechanismus der Clp1-vermittelten Inhibition der bW- und Rbf1-Funktion in Ustilago maydis / Benjamin Faist ; Betreuer : J. Kämper ». Karlsruhe : KIT-Bibliothek, 2017. http://d-nb.info/1139360442/34.
Texte intégral
19
White, Gemma. « The role of fractalkine (CX₃CL1) and its receptor (CX₃CR1) in vascular biology ». Thesis, University of Oxford, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.670108.
Texte intégral
20
Pinter, Niko [Verfasser], Kai [Akademischer Betreuer] Heimel, Kai [Gutachter] Heimel et Gerhard [Gutachter] Braus. « Analysis of Clp1-dependent UPR modulation in Ustilago maydis / Niko Pinter ; Gutachter : Kai Heimel, Gerhard Braus ; Betreuer : Kai Heimel ». Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2019. http://d-nb.info/1188464884/34.
Texte intégral
21
Chen, Chun-Ti. « Regulation of the Cdc14-like Phosphatase CLP1 in Schizosaccharomyces pombe and Identification of SID2 Kinase Substrates : A Dissertation ». eScholarship@UMMS, 2009. https://escholarship.umassmed.edu/gsbs_diss/449.
Texte intégralRésumé :
Coordination of mitosis and cytokinesis is crucial to generate healthy daughter cells with equal amounts of genetic and cytoplasmic materials. In the fission yeast Schizosaccharomyces pombe, an evolutionarily conserved Cdc14-like phosphatase (Clp1) functions to couple mitosis and cytokinesis by antagonizing CDK activity. The activity of Clp1 is thought to be regulated in part by its subcellular localization. It is sequestered in the nucleolus and the spindle pole body (SPB) during interphase. Upon mitotic entry, it is released into the cytoplasm and localized to the kinetochores, the actomyosin ring, and the mitotic spindle to carry out distinct functions. It is not clear how Clp1 is released from the nucleolus, however, once released, a conserved signaling pathway termed Septation Initiation Network (SIN) functions to retain Clp1 in the cytoplasm until completion of cytokinesis. The SIN and Clp1 function together in a positive feedback loop to promote each other’s activity. That is, the SIN promotes cytoplasmic retention of Clp1, and cytoplasmic Clp1 antagonizes CDK activity and reverses CDK inhibition on the SIN pathway to promote its function and activity. However, at the start of this thesis, the mechanism by which the SIN regulated Clp1 was unknown. The SIN pathway is also required to promote constriction of the actomyosin ring, and the septum formation. However, its downstream targets were still uncharacterized. In two separate studies, we studied how Clp1 is released from the nucleolus at mitotic entry and how the SIN kinase Sid2 acts to retain Clp1 in the cytoplasm. We identified several Sid2 candidate substrates, and revealed other functions of the SIN pathway in coordinating mitotic events.
22
Fortenbacher, Julia [Verfasser], et J. [Akademischer Betreuer] Kämper. « Charakterisierung des Proteinkomplexes Clp1 mit bE/bW und seine Auswirkung auf die pathogene Entwicklung in Ustilago maydis / Julia Fortenbacher ; Betreuer : J. Kämper ». Karlsruhe : KIT-Bibliothek, 2019. http://d-nb.info/1202076750/34.
Texte intégral
23
Dahlmann, Cordula [Verfasser]. « Retinaler Phänotyp dreier Mausmodelle für die neuronale Ceroidlipofuszinose : (CLN1-knockout Mausmodell, CLN3Δex7/8-knock-in Mausmodell und CLN6-knockout Mausmodell) / Cordula Dahlmann ». Berlin : Medizinische Fakultät Charité - Universitätsmedizin Berlin, 2013. http://d-nb.info/1031097074/34.
Texte intégral
24
Guyon, Clotilde. « Contrôle post transcriptionnel des transcrits des auto-antigènes induits par AIRE dans le thymus ». Thesis, Sorbonne Paris Cité, 2016. http://www.theses.fr/2016USPCB138/document.
Texte intégralRésumé :
La tolérance immunologique assure le maintien de l’intégrité des organismes contre les pathogènes tout en respectant les constituants du soi. La dérégulation de ce mécanisme entraîne la survenue de maladies autoimmunes qui touchent de 5 à 10% de la population générale. Un mécanisme clé de la tolérance immunologique est la délétion clonale des lymphocytes T auto-réactifs après qu’ils ont reconnu leur antigène spécifique au cours de leur maturation dans le thymus. Il a été montré qu’un vaste répertoire d’autoantigènes est exprimé par les cellules épithéliales médullaires du thymus (mTECs) sous l’action de la protéine AIRE (AutoImmune Regulator). Les études présentées dans ce travail participent à améliorer la compréhension du fonctionnement de AIRE. Au-delà de la fonction de transactivation de AIRE, nous avons montré, avec le reséquençage à haut débit (RNAseq) des mTECs, que les transcrits des autoantigènes induits par AIRE ont des extrémités 3’UTRs courtes associées à l’utilisation des sites de polyadénylation (pA) alternatifs. Nous avons identifié par analyse de données de CLIPseq une fixation préférentielle du complexe de terminaison de la transcription au niveau des pAs alternatifs des gènes sensibles à AIRE. Nous avons également mis en évidence l’interaction de AIRE au complexe de terminaison de la transcription. Parmi plusieurs partenaires de AIRE associés à ce complexe, nous avons montré par interférence d’ARN et RNAseq le rôle de CLP1 dans le choix des pAs alternatifs. De plus nous montrons que CLP1 est le seul membre du complexe de terminaison à être préférentiellement exprimé dans les mTECs matures. Fonctionnellement, nous avons mis en évidence une stabilité plus importante pour les transcrits des autoantigènes induits par AIRE en bloquant la transcription des mTECs ex-vivo par traitement à l’Actinomycine D. Nous montrons également l’existence d’un raccourcissement 3’UTR général dans les mTECs matures par rapport aux mTECs immatures et autres tissus de la souris, auquel se combine le raccourcissement spécifique des gènes dépendant de AIRE. Après avoir identifié par des analyses de Gene Ontology une activité cellulaire exacerbée dans les mTECs matures vs immatures, nous confirmons l’activité transcriptionnelle exacerbée des mTECs matures in-vivo grâce à l’incorporation de 5Ethynyl Uridine (EU) dans les ARN néosynthétisés après injection intrathymique. Le raccourcissement des transcrits des auto-antigènes associé à leur stabilité accrue suggère qu’ils échappent à la répression transcriptionnelle médiée par les microARNs. Ce travail a permis d’identifier les bases moléculaires de la régulation post-transcriptionnelle des autoantigènes dans le thymus. Dans l’étude faite en collaboration avec l’équipe de Jakub Abramson du laboratoire Weizmann, démontre que Sirt1, une désactylase ADN dépendante, est exprimé de façon abondante dans les mTEC AIRE+ et ce grâce à l’utilisation de profil d’expression génétique, de cytométrie en flux et d’analyses d’immunoblot de différents types cellulaires thymiques. De plus lorsque Sirt1 est inactivé, dans les lignées germinales et des lignées TEC, l’expression des gènes AIRE dépendants diminuent et donc avec elle la tolérance immune induite par AIRE. La capacité désacétylase de Sirt1 est nécessaire pour l’expression des gènes induits par AIRE dans les mTECs. Sirt1 cible surement d’autres molécules nucléaires, impliquées dans la voie de AIRE. Elle pourrait avoir un rôle plus étendu dans la régulation du système immunitaire et être présent à la périphérie. Cette étude a mis en évidence un rôle important de Sirt1 dans la tolérance centrale dans le thymus à travers la régulation des gènes induits par AIRE. (...)No abstract
25
Ziegler, Yvonne [Verfasser], Volker [Akademischer Betreuer] Lipka et Thomas [Akademischer Betreuer] Teichmann. « The role of the putative receptor-like cytoplasmic kinase CLR1 in chitin signalling / Yvonne Ziegler. Betreuer : Volker Lipka. Gutachter : Volker Lipka ; Thomas Teichmann ». Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2016. http://d-nb.info/1081543574/34.
Texte intégral
26
Hieber, Marie-Lisa [Verfasser]. « Die prognostische Bedeutung der TP53-Mutation für Ansprechen und Überleben von CLL-Patienten : Analyse im Rahmen der prospektiven CLL11-Studie (Clb vs. RClb vs. GClb) / Marie-Lisa Hieber ». Ulm : Universität Ulm, 2018. http://d-nb.info/1166757110/34.
Texte intégral
27
Kwag, Doo Young [Verfasser], et Michael [Akademischer Betreuer] Hallek. « Disease-specific complications of chronic lymphocytic leukemia in binet stage a patients : analysis of immunodeficiency, autoimmune constellations and infections in the CLL1-protocol / Doo Young Kwag. Betreuer : Michael Hallek ». München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2012. http://d-nb.info/1029040273/34.
Texte intégral
28
Schmitt, Christian [Verfasser], et Rolf W. [Akademischer Betreuer] Hartmann. « Development of new lead-like dual inhibitors of the cdc2-like kinase 1 (Clk1) and dual specificity Y-phosphorylation regulated kinases 1A and 1B (Dyrk1A and Dyrk1B) / Christian Schmitt. Betreuer : Rolf W. Hartmann ». Saarbrücken : Saarländische Universitäts- und Landesbibliothek, 2014. http://d-nb.info/1056906855/34.
Texte intégral
29
Da, Silva Sophie. « Identification des gènes CiSTM et CiCLV1p chez Cichorium intybus : implicatio potentielle dans les modifications d'identité cellulaire au cours des phases précoces de l'embryogenèse somatique et du développement des embryons zygotiques et de la graine ». Lille 1, 2004. https://ori-nuxeo.univ-lille1.fr/nuxeo/site/esupversions/fb7d2815-c338-4560-bd04-b5e881028bde.
Texte intégralRésumé :
L'embryogenèse somatique est un processus au cours duquel des cellules différenciées se dédifférencient pour ensuite réorienter leur programme génétique vers une voie embryogène. Dans le cadre de l'étude du déterminisme de la réactivation cellulaire à l'origine de la fonnation des embryons somatiques, notre objectif était d'identifier chez la chicorée des gènes impliqués dans l'identité des cellules méristématiques, tels que les gènes AtSTM et AtCLV1 d'Arabidopsis, et de détenniner leur profil d'expression au cours des stades précoces de l'embryogenèse zygotique et de l'embryogenèse somatique. La première partie de ce travail a été consacrée au clonage des ADNc CiCLV1p et CiSTM chez la chicorée. Des analyses de séquences et des études de phylogénie ont montré que CiSTM était potentiellement un gène orthologue de AtSTM alors que CiCLV1p, bien que membre de la sous'-famille des RLK-LRR XI, était plutôt rattaché à un autre groupe de séquences,proche de AtCLV1. La seconde partie de cette étude a regroupé la caractérisation cytologique des phases principales de l'embryogenèse zygotique de la chicorée et la détennination des profils de réactivation de nouveaux génotypes embryogènes et non embryogènes sélectionnés au laboratoire. Au cours de l'embryogenèse zygotique de la chicorée, nous avons identifié des fenêtres de développement caractérisant les stades classiquement observés chez Arabidopsis et d'autres espècesLes génotypes embryogène K59 et non embryogène C15 ont été choisis comme génotypes à réponse extrême, selon leur capacité à produire les différentes figures cytologiques observées. La dernière partie de la thèse a été consacrée à l'étude des profils d'expression des gènes CiSTM et CiCLV1p au cours de l'embryogenèse zygotique et au cours de l'embryogenèse somatique par RT PCR en temps réel. Ces résultats ont été comparés aux fenêtres de développement de l'embryon zygotique et aux profils de réactivation des génotypes sélectionnés. Au cours de l'embryogenèse zygotique, une induction importante de l'expression des gènes CiSTM et CiCLVlp a été corrélée à la mise en place d'un dôme méristématique entre les cotylédons. Au cours de l'embryogenèse somatique, les transcrits des gènes CiSTM et CiCLV1p sont accumulés de façon différentielle lors du passage de certaines cellules en cours de réactivation vers l'état réactivé. Ces résultats suggèrent une implication des gènes CiSTM et CiCLV1p dans les mécanismes de changement d'identité cellulaire chez la chicorée,que ce soit au cours de l'embryogenèse zygotique ou au cours de l'embryogenèse somatique
30
Monaghan, Richard. « A novel nuclear role for the mitochondrial hydroxylase Clk-1 ». Thesis, University of Manchester, 2012. https://www.research.manchester.ac.uk/portal/en/theses/a-novel-nuclear-role-for-the-mitochondrial-hydroxylase-clk1(8de582a1-bfe3-4421-bfbf-75f9229a13b1).html.
Texte intégralRésumé :
The lipid hydroxylase Clk-1 catalyses an essential step in the mitochondria localised ubiquinone biosynthetic pathway that is conserved throughout eukarya. Like many canonical mitochondrial proteins, murine Clk-1 is targeted and imported into mitochondria by virtue of a ~4kDa N-terminal mitochondrial targeting domain that is cleaved following translocation into the mitochondrial matrix. Clk-1 mutants in C. elegans and heterozygous Clk-1+/- mice exhibit increased longevity and delayed development rates compared to wild type individuals, with associated changes in oxidative stress signalling pathways. Previous work in the laboratory identified human Clk-1 as a potential interactor of Sin1, a component of the mammalian target of rapamycin signaling pathway, which has also been associated with modulations that affect lifespan. Here the interaction between Clk-1 and Sin1 is further characterised and Clk-1 is identified as a potential substrate of the Sin1-associated kinases cAMP dependent kinase, protein kinase C and unc-51-like kinase 1. Interestingly, a fraction of Clk-1 was observed residing in the nucleus, in addition to its mitochondrial localisation. The sequence determinants for Clk-1 nuclear localisation were found to be in the same N-terminal region required for mitochondrial localisation and a single point mutant was identified that translocated to the mitochondria but not the nucleus. Oxidative stress treatment was shown to increase the level of uncleaved Clk-1 and this form was enriched in nuclear and chromatin fractions. Clk-1 was found to be associated with over 1000 genomic loci following Clk-1 chromatin immunoprecipitation followed by promoter microarray analysis. Cells stably expressing the Clk-1 non-nuclear point mutant displayed decreased resistance to oxidative stress-induced cell death and increased levels of oxidative species following treatment with exogenous stress. In addition, c-Jun N-terminal kinase signaling was enhanced in these cells in response to tumour necrosis factor-α stimulation. Microarray analysis of these cells showed both positive and negative transcript changes compared to wild type Clk-1 expressing cells which was significant for over 2000 genes. Functional clustering analysis identified enrichment for gene groups associated with glycolytic and tricarboxylic acid cycle metabolism, Wnt signaling, and several specific differentiation and oncogenic pathways. Many of the genes identified are reported to be regulated by promoter methylation, and this was confirmed for glutathione-S-transferase P1 that had significantly decreased expression in mutant Clk-1 expressing cells. Loss of Clk-1 nuclearisation or Clk-1 activity within the nucleus could therefore be acting as part of specific differentiation pathways either during early development or following differentiation of specific cell lineages. Nuclear Clk-1’s ties to respiration and cell survival pathways, and sensitivity to oxidative stress, could also implicate it in oncogenic progression and/or the Clk-1 ageing phenotype.
31
Andreou, Tereza. « Nuclear localisation of Clk-1 : a novel regulator of mitochondrial to nuclear signalling ». Thesis, University of Manchester, 2015. https://www.research.manchester.ac.uk/portal/en/theses/nuclear-localisation-of-clk1-a-novel-regulator-of-mitochondrial-to-nuclear-signalling(11ce8f0a-0e9e-40ae-840f-7805f2abede6).html.
Texte intégralRésumé :
Mitochondria generate cellular energy through oxidative phosphorylation and perturbation of this process can lead to increased levels of reactive oxygen species (ROS). It is critical that mitochondria communicate with the nucleus to regulate gene expression and maintain ROS homeostasis. This occurs via retrograde signalling pathways. Recently, a novel nuclear role was uncovered for Clk-1, a mitochondrial enzyme with a well-established role in oxidative phosphorylation. Upon conditions of increased ROS, a pool of Clk-1 translocates to the nucleus where it associates with chromatin and regulates gene expression. Experiments described in this thesis were aimed at characterising the regulation of Clk-1 localisation and dissecting the biological function of nuclear Clk-1.A non-classical nuclear localisation signal in the N-terminal region of Clk-1 was identified, containing three conserved Arginine residues, the most important of which was Arginine28. It was confirmed that the positive charge of Arginine is important for determining Clk-1 nuclear localisation. In addition, Tyrosine26 was also identified as having a regulatory role in Clk-1 localisation, possibly through a phosphorylation event. Increased ROS levels block the import of Clk-1 into the mitochondrial matrix and its cleavage by the mitochondrial processing peptidase. However, uncleaved Clk-1 was detected in the mitochondrial fraction, in addition to the nucleus, upon oxidative stress treatment. These results suggest that a pool of Clk-1 pre-protein may first be targeted to mitochondria prior to translocating to the nucleus. Interestingly it was found that a potentially misfolded Clk-1 mutant (E178K) is not ROS-responsive and that uncleaved Clk-1(E178K) predominantly associates with mitochondria. Similarly, a pharmacological inducer of Clk-1 misfolding promoted the association of uncleaved Clk-1 with the mitochondrial fraction. Collectively, these results suggest that Clk-1 import efficiency and the kinetics of Clk-1 folding determine its subcellular localisation. To specifically investigate the function of nuclear Clk-1, a conditional knock-in mouse model featuring a mutant version of Clk-1 (R28A) with impaired nuclear localisation is being generated. The cloning strategy employed to generate the targeting vector for the R28A conditional knock-in mouse and the strategy used to screen for positive mouse embryonic stem cell clones harbouring the R28A mutation in their genome, is presented in this thesis. The identification of Clk-1 as a novel regulator of mitochondrial to nuclear signalling has significant implications for understanding the cellular mechanisms that regulate mitochondrial homeostasis and could contribute to our understanding of diseases in which mitochondrial homeostasis has been disrupted.
32
Barnes, Robert. « A nuclear role for the respiratory enzyme CLK-1 in regulating reactive oxygen species, the mitochondrial unfolded protein response and longevity ». Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/a-nuclear-role-for-the-respiratory-enzyme-clk1-in-regulating-reactive-oxygen-species-the-mitochondrial-unfolded-protein-response-and-longevity(9a41f8d6-d11d-4102-b250-7b20091f4ded).html.
Texte intégralRésumé :
As the major source of energy in the cell, mitochondria need to be able to effectively communicate their status to the nucleus. Defects in this process can have profound effects on the health of an organism and may affect its lifespan. The mitochondrial enzyme CLK-1 is required to produce ubiquinone for respiration and CLK-1 loss of function mutants have been shown to increase lifespan in both Caenorhabditis elegans and mammals. In this thesis, it is demonstrated that in addition to its mitochondrial role, CLK-1 also localises to the nucleus in C. elegans. This nuclear localisation is mediated by reactive oxygen species (ROS) and can be blocked using anti-oxidant treatment. In the nucleus CLK-1 regulates gene expression to suppress ROS production, suppresses the mitochondrial unfolded protein response and regulates lifespan. The importance of CLK-1 enzymatic activity for its nuclear role was also tested and it appears that enzymatic function is required to regulate ROS homeostasis but not for lifespan regulation. Interestingly, in fertile adult hermaphrodites nuclear CLK-1::GFP was only detected during the reproductive period. This suggests that there is a second mechanism regulating its localisation. This research indicates that CLK-1 may be part of a homeostatic regulatory mechanism that acts to suppress activation of stress responses in response to minor fluctuations in ROS levels.
33
Pinter, Niko. « Analysis of Clp1-dependent UPR modulation in Ustilago maydis ». Doctoral thesis, 2019. http://hdl.handle.net/21.11130/00-1735-0000-0003-C12F-F.
Texte intégral
34
Yeh, I.-Hsin, et 葉怡欣. « Characterization of putative metastasis-related proteins enriched from comparative analysis of lung adenocarcinoma cell lines CL1-0 and CL1-5 secretomes ». Thesis, 2013. http://ndltd.ncl.edu.tw/handle/72804496264841341863.
Texte intégralRésumé :
碩士國立陽明大學
醫學生物技術暨檢驗學系
101
Adenocarcinoma, the most frequent type of lung cancer, is one of the most common diseases around the world and a major health problem in Taiwan. Based on microscopic morphologies, lung cancers can be classified as non-small cell lung cancer (NSCLC) (85%) and small cell lung cancer (SCLC) account (15%). The major therapeutic methods for lung cancer include surgical resection, radiation therapy, chemotherapy, and target therapy. Lung cancer has often spread to other areas of the body when it is diagnosed. Metastasis-related proteins may serve as potential diagnostic markers or targets for cancer intervention. In this study, we used two NSCLC cell lines, a low-metastatic CL1-0 and the highly metastatic CL1-5 for the identification of such proteins. Of the 82 candidates identified in our previous proteomic analyses, I performed further Ingenuity Pathways Analysis (IPA) and literature and selected fifteen candidate genes to validate their expression in these two cell lines by semi-quantitative RT-PCR. We found eight candidates had higher mRNA levels in CL1-5 than in CL1-0. To examine whether the proteins are associated with NSCLC metastasis, we have knocked down the expression eight candidate genes with shRNA, and perform wound healing assay to examine the effect of silencing these genes on the cell migration abilities. In addition, immunofluorescence staining indicated the alteration of beta-actin and cell morphology after knocking down Candidate #19, Candidate #7, Candidate #18 and Candidate #42 of the eight candidates. The role of these genes in metastasis of lung cancer deserve further investigation. Keywords: Non-small cell lung cancer, Proteomics, Cancer metastasis, Secretome
35
Wang, Chung-yao, et 王重堯. « Phosphoproteome Analysis of Lung Cancer Cell Line CL1-5 ». Thesis, 2008. http://ndltd.ncl.edu.tw/handle/30877637547436967369.
Texte intégralRésumé :
碩士國立成功大學
環境醫學研究所
97
Lung cancer is one of the most common causes of cancer death in the world. Although the total annual number of cases has declined, probably due to the decreased trend in cigarette consumption, the incidence and mortality rate of lung cancer have increased at an alarming rate in the female population and in developing countries. Cancer metastasis and invasion are the most crucial steps in the cancer progress, occurring via a series of biologic activities including the interaction of tumor cells with the surrounding environment. These steps are the major causes of treatment failure and cancer deaths. Therefore, understanding the mechanisms that regulate progress of tumor cells is important for development of novel therapies to control cancer. Based on recent researches, evidences indicate that the changed protein phosphorylation result in altered invasive and metastatic properties of tumor cells. For example, Phosphorylated FAK was seen in invasive ovarian carcinoma, but not in the normal cells. Here, we used metal affinity based enrichment coupled with LC-MS/MS analysis, qualitatively but comprehensively defined the phosphoproteome of this lung cancer cell lines CL1-5. Cancer cell lysates were digested and phosphopeptides were enriched by self-packing titanium dioxide (TiO2) microcolumn. The enrichment protocol was based on a recently published method. After enriched by TiO2 microcolumn, eluted peptides were subjected to LC-MS/MS analysis and the raw data were searched against Swiss-Prot database. Although phosphopeptide enrichment has been set up, a process to reduce the false-positive phosphopeptides identification by current database search tools is needed. Commonly accepted MS/MS data filtering criteria are not suitable for data analysis with post-translational modifications such as phosphorylation. Thus, we manual validated all the MS/MS spectra to exclude low confidence identities. A total of 719 phosphorylaed peptides, assigned to 274 proteins, were identified in this study. Those identified phosphorylation events may provide novel information for not only mechanistic aspects of lung cancer but also potential drug targets for hampering the cancer metastatic processes.
36
Ziegler, Yvonne. « The role of the putative receptor-like cytoplasmic kinase CLR1 in chitin signalling ». Doctoral thesis, 2015. http://hdl.handle.net/11858/00-1735-0000-0028-8698-6.
Texte intégralRésumé :
Pflanzen erkennen potentielle Pathogene anhand von konservierten Mikroben-assoziierten molekularen Mustern (MAMPs) welche sie über membranlokalisierte Rezeptoren wahrnehmen. Der durch diese Rezeptoren aktivierte Signalweg spielt eine wesentliche Rolle in der pflanzlichen angeborenen Immunität. Das Binden eines MAMPs an die oberflächenexponierten Ektodomänen der Rezeptoren führt typischerweise dazu, dass diese homo- oder heteromere Komplexe bilden. Diese Komplexe können aus rezeptorartigen Kinasen (RLKs), rezeptorartigen Proteinen (RLPs) sowie aus rezeptorartigen zytoplasmatischen Kinasen (RLCKs), welche keine extrazelluläre Domäne zur Ligandenbindung besitzen, bestehen.
Der Fokus dieser Arbeit liegt auf einem möglichen heteromeren Signalkomplex der unteranderem aus der lysinhaltigen-Motiv (LysM) RLK CERK1 besteht. CERK1 spielt eine Rolle in der durch Chitin induzierten Signaltransduktion und Abwehrantwort in Arabidopsis. In einer vorangegangenen Hefe-Zwei-Hybrid-Analyse wurde die RLCK CLR1 als möglicher Interaktor der CERK1 Kinasedomäne identifiziert. Vergleichende Sequenzanalysen zeigen, dass die Aminosäuresequenz von CLR1 eine hohe Homologie zu den Sequenzen der Kinasedomänen anderer Arabidopsis LysM-RLKs aufweist. Dies könnte möglicherweise für die Funktion des Proteins eine Rolle spielen. Die auf TAIR10 annotierte CLR1 Sequenz scheint falsch annotiert worden zu sein, da das eigentliche Protein laut Analysen in dieser Arbeit wahrscheinlich erst 23 Aminosäuren Richtung C-Terminus beginnt, wodurch dann ein mögliches N-Myristoylierungsmotiv exponiert wird.
In vitro wird CLR1 direkt von der CERK1 Kinasedomäne phosphoryliert. CLR1 Fusionsproteine wurden in stabil transgenen Arabidopsis-Pflanzen CERK1-abhängig durch Chitin phosphoryliert. Unabhängig von der möglichen N-terminalen Myristoylierung scheint CLR1 sowohl in vitro also auch in vivo ein Phosphorylierungssubstrat von CERK1 darzustellen. Mikrosomale Fraktionierungen und Analysen zur subzellulären Lokalisation in transgenen Pflanzen zeigten dass die Mehrheit der CLR1 Proteine löslich ist, wobei auch eine kleine Subpopulation von CLR1 membrangebunden in Pflanzenzellen vorliegt. Drei unabhängige T DNA Insertionslinien wurden isoliert und im Hinblick auf die Weiterleitung Chitin-induzierter Signale und Immunität gegen pilzliche und bakterielle Schädlinge getestet. Die clr1 T-DNA Linien wiesen eine verringerte ROS Produktion, MAPK Aktivierung und Expression von Abwehrgenen auf, was eine Rolle für CLR1 im Chitin-induzierten Signalweg bestätigt. Dabei hing die Ausprägung des Phänotyps von der Position der T-DNA ab. clr1 Pflanzen waren nicht in der Resistenz gegen pilzliche Schädlinge beeinträchtigt, wohingegen sie eine leicht erhöhte Anfälligkeit gegenüber bakterieller Infektionen zeigten. Da der CLR1 Promotor erhöhte Aktivität in Hydathoden zeigt, könnte CLR1 darin involviert sein selektiv das Eintreten von Pathogenen über diese konstitutiv geöffneten Öffnungen einzugrenzen.
37
Liu, Yu-Lin, et 劉玉琳. « The Role of NAD(P)H:Quinone Oxidoreductase 1 (NQO1) in β-Lapachone-Mediated Cell Death on Non-Invasive and Invasive Human Lung Adenocarcinoma cells (CL1-1 and CL1-5) ». Thesis, 2005. http://ndltd.ncl.edu.tw/handle/06503226907624730615.
Texte intégralRésumé :
碩士國立陽明大學
解剖暨細胞生物學研究所
93
β-Lapachone, the product of a tree (Tabebuia avellanedae) from South America, has been known to exhibit anti-parasitic, -tumor and -virus effects. The aim of the present study was to elucidate the mechanisms by which β-lapachone exerts its cytotoxicity to non-invasive (CL1-1) and invasive human lung adenocarcinoma cells (CL1-5). Exposure of CL1-1 and CL1-5 cells to β-lapachone resulted in survival inhibition and apoptosis. This increase in apoptosis was associated with a decrease of mitochondrial membrane potential suggesting that β-lapachone treatment destroyed mitochondrial function. Furthermore, the induction of calcium influx in β-lapachone-treated cells following 1h treatment might induce ER stress for cytotoxicity. On the other hand, inhibition of PI3K/Akt pathway was also found. However, all of these events were dependent upon NAD(P)H: quinone oxidoreductase (NQO1) activity. Counteracting NQO1 activity with dicumoral prevented cell death induced by β-lapachone. Therefore, our results implied that β-lapachone-mediated cytotoxicity against non-invasive and invasive human lung adenocarcinoma cells (CL1-1 and CL1-5) might occur through the activation of NQO1 that might induce the intracellular calcium imbalance and mitochondrial dysfunction and inhibit PI3K/Akt signaling pathway.
38
Chang, Kung-Yan, et 張工彥. « Studies of ALA Photodynamic Effects on Mitochondria and Related Biological Responses in CL1-5 Cells ». Thesis, 2003. http://ndltd.ncl.edu.tw/handle/74242193490507297284.
Texte intégralRésumé :
碩士國立臺灣大學
口腔生物科學研究所
92
Photodynamic therapy (PDT) is based on the selective retention of a photosensitizer by tumor tissue and the subsequent irradiation of this tissue with light. 5-aminolevulinic acid (5-ALA), a pro-drug of photosensitizer, is metabolized in mitochondria to protoporphyrin IX (PpIX) which selectively accumulates to greater extent in cancer cells. Using CL1-5 Human lung adenocarcinoma, we demonstrated that the production of PpIX occurs in mitochondria under fluorescence microscopy. After ALA-PDT, we observed morphological changes of cell and breakdown of mitochondrial membrane potential. This was followed by chromosome condensation and caspase-3 activation which result in 10% cell death. Besides, there was significant number of cells which could not be detached from the dish by trypsin compared to control cells without ALA-PDT. This reduced trypsin detachment accompanies F-actin reorganization. N-acetyl-cysteine (NAC), free radical scanvanger, inhibited PDT-induced resistance to trypsinization and F-actin reorganization. After 5 times ALA-PDT (5-PDT) growth rate and the ability of invasion and migration in CL1-5 were significantly decreased. These data indicate that the PDT-induced mitochondrial photo-damage are critical in decreasing mitochondrial membrane potential, which leads to morphological changing, F-actin reorganization, apoptotic death, and decreased growth rate and invasion during PDT with ALA.
39
Li, Rung-Chi, et 李容奇. « Mechanism of COX-2-mediated anti-apoptotic effects in human lung adenocarcinoma cell line CL1 ». Thesis, 2000. http://ndltd.ncl.edu.tw/handle/85080070357411740503.
Texte intégralRésumé :
碩士國立臺灣大學
毒理學研究所
88
Apoptosis profoundly influences a wide variety of physiological processes. Thus, dysregulation of apoptosis has been implicated in several human diseases, ranging from cancer to immunity, AIDS and neurological disorders. Recently, emerging evidence show that resistance to apoptosis becomes an important mechanism in regulating the invasive ability of malignant tumors. Here we demonstrated that a human lung cancer CL1-5 cells with higher invasive ability exhibited remarked resistance to apoptosis; in contrast, another lower invasive subline CL1-0 was susceptible to apoptosis. Immunoblot analysis showed that cyclooxygenase(Cox)-2 and Mcl-1 proteins were consistently elevated in CL1-5 cells. This prompted us to hypothesize a closely functional interaction between Cox-2 and Mcl-1. To address this, we treated CL1-0 cells with PGE2, a product of Cox-2, and examined the cellular response to apoptosis. PGE2 treatment rendered CL1-0 cells became resistance to UV-elicited apoptosis and induced the increased expression of Mcl-1. We further transiently transfected Cox-2-expressing plasmid along with an indicator vector, pCMV-bgal, into CL1-0 cells. After 48-hr transfection, Cox-2 transfected CL1-0 cells exhibited more viable cells following exposure to b-lapachone or UV when compared to vector control cells. Furthermore, we co-transfected the anti-sense Mcl-1 and COX-2 plasmids into CL1-0 and demonstrate that anti-sense Mcl-1 could decrease the Cox-2-caused viable blue cells. Again we found that overexpression of Cox-2 in CL1-0 cells led to an up-regulation of Mcl-1. Pretreatment with phosphatidylinositol 3-Kinase (PI 3-K) inhibitors, LY294002 and Wortmannin, effectively inhibited PGE2-mediated up-regulation of Mcl-1 in CL1-0 cells. In line with the observation, when Cox-2-transfected cells were further transfected a dominant-negative mutant of PI 3-K, DN-p85, the Cox-2-mediated Mcl-1 expression was blocked. In contrast to mock cells we found COX-2 overexpressing cells exhibited highly Akt phosphorylation, one major target of PI 3-K, which was abolished by pretreatment of LY294002 and Wortmannin, too. To investigate these observations completely, we established CL1-0 cells which stably expressed cox-2 gene. Pretreatment of LY294002 and Wortmannin disrupted COX-2 transfectants-caused Mcl-1 protein level induction and Akt phosphorylation. And CL1-0/COX-2 pure clone had higher PI 3-K phosphorylation than CL1-0 cell line. We further examined the invasive ability of CL1-0/COX-2 using matrix gel in vitro invasion assay and Metalloproteinase zymography assay. In conclusion, the present study demonstrates that Cox-2 plays a key role in regulating the invasive ability of human adenocarcinoma cells by suppressing cellular apoptosis. The mechanism underlying Cox-2 prevents apoptosis which is mediated by up-regulation of Mcl-1 with a PI 3-K-dependent pathway.
40
Ma, Haou-Tzong, et 馬豪聰. « Inosine Monophosphate Dehydrogenase Inhibitor Regulates Expression of Fucosyltransferase 8 in lung cancer cell line CL1-5 ». Thesis, 2015. http://ndltd.ncl.edu.tw/handle/23357738560152345903.
Texte intégral
41
Liu, Pei-Yi, et 劉佩宜. « Study on the mechanisms of Pipoxolan inhibiting the migration in CL1-5 human lung adenocarcinoma cells ». Thesis, 2012. http://ndltd.ncl.edu.tw/handle/n75z24.
Texte intégralRésumé :
碩士中國醫藥大學
藥學系碩士班
100
Pipoxolan is a commonly prescribed smooth muscle relaxant in Taiwan. Clinical use is to alleviate symptoms such as muscle spasms induced disease or pain. Our previous studies have shown that pipoxolan induce apoptosis and arrest the cell cycle at the G0/G1 phase in human leukemia cancer cells. HL-60 cells were arrested in the G0/G1 phase via the induction of p53⁄p21 by pipoxolan. Apoptosis was associated with an increased Bax/Bcl-2 ratio, cytochrome c release, cleavage of procaspases -9, -3 and hydrolysis of poly(ADP-ribose) polymerase (PARP) and high levels of ROS were produced early in the drug treatment, but the effects on the migration and invasion of human lung cancer cells have not been reported. Lung cancer is one of the most common malignancies in most countries including Taiwan and its metastasis is the major cause of death. The major reason of its high mortality rate is cancer metastasis, the hardest part of cancer treatment. The aim of our study is to investigate the effect of pipoxolan inhibiting the migration in highly migratory CL1-5 lung adenocarcinoma cells. The data demonstrated that CL1-5 cells were treated with pipoxolan (5.2, 12.9 and 25.8 μM) for 24h having the inhibitory effects of migration by wound scratch assay and transwell assay. The gelatin zymography assay indicated that pipoxolan inhibited the activity of matrix metalloproteinases 2 (MMP-2) and MMP-9 in a concentration-dependent manner. Western blotting analysis indicated that pipoxolan suppressed the phosphorylation of JNK1/2, p38, Rac-1, MMP-2 and -9 protein expressions, thereby inhibiting CL1-5 cells migration. In conclution, pipoxolan significantly inhibited CL1-5 human lung adenocarcinoma cells migration through inactivation of the JNK and p38 MAPK signaling pathway and decrease the activities of MMP-2 and -9.
42
Tsai, You-Wei, et 蔡有緯. « A dual-functional inhibitor causes apoptosis and inhibits metastasis of lung cancer cell line CL1-5 ». Thesis, 2019. http://ndltd.ncl.edu.tw/handle/6n444z.
Texte intégralRésumé :
碩士國立臺灣大學
生化科學研究所
107
Lung cancer has a high mortality rate and often metastasizes to other organs. The cell line used in the study was CL1-5, which is a non-small cell lung cancer (NSCLC). According to previous studies, cancer cells can be killed through blocking the interaction between CCT-β and β-tubulin by I-Trp. The small molecule compound-845 used in this study was screened among a series of bactericidal compounds synthesized by NHRI and is the most potential to kill CL1-5 cells. In addition, its structure is composed of a functional group which is similar to the reversible inhibitors of CCT-β: β-tubulin complex previously simulated by computer and a side chain of I-Trp. It has been found that compound-845 would induce ER stress in CL1-5 cells, which in turn leads to the release of intracellular calcium ion and apoptosis. Furthermore, analyzed by wound healing assay and transwell assay, it has been found that compound-845 would inhibit the migratory and invasive ability of CL1-5 cells. Moreover, compound-845 would inhibit EMT, AKT/β-catenin and integrin signaling pathway investigated by western blot, thereby inhibiting CL1-5 cell metastasis. In conclusion, this study reveals that this dual-functional inhibitor compound-845 probably has anti-tumor and anti-metastasis ability against human lung cancer.
43
Lin, Yu-Cun, et 林俞村. « Cathepsin S expression and its effects in human lung adenocarcinoma CL1 cells after glutathione reductase (GR) siRNA transfection ». Thesis, 2011. http://ndltd.ncl.edu.tw/handle/62964664938735317371.
Texte intégralRésumé :
碩士國立清華大學
生物科技研究所
100
CL1-X (1~5) cells as a lung-cancer metastasis cell model were established through Matrigel-coated Transwell-membrane in cell culture insert (in each well of 24-well micro-titer dish) after 72-h selection and 5 consecutive procedures from CL1-0 human lung adenocarcinoma in the laboratory of Dr. P. C. Yang from National Taiwan University Hospital. Their invasive abilities through basement membrane matrix showed a 4- to 6- fold increase over that of the parental cells. Nevertheless, not all characters for their genetic instabilities were known for many years. Recently, data from our laboratory indicated that endogenous and exogenous cathepsin S (CTSS) protease activity in CL1-3 increased 5~7-fold and 2~3-fold, respectively, in comparison with those in CL1-0 parental cells. Measurement on several anti-oxidative molecules resulted in about one-half glutathione reductase (GR) activity (P<<0.01), less glutathione (GSH) (P<0.01), more GSH transferase (GST) (P<0.03), slightly more Glutathione peroxidase (GpX) activity (P=0.06-0.09) in CL1-3 cells having relatively more invasive (higher CTSS) abilities. To understand the causes of decreased GR and increased CTSS activity in CL1-3 cells after invasive ability selection from parental CL1-0 cells, gene manipulation and GR-siRNA transfection in CL1-0 cells to produce similar phenomenon as CL1-3 cells was conducted in this study. GR-siRNA (#2, #3 and #5 different sequence, from Academia Sinica) and puromycin gene containing lentivirus vectors were transfected into CL1-0 cells. Clones containing low GR activity and high CTSS activity were selected. In Exp. I, among 68 clones picked from puromycin containing medium, 48 clones (>70%) had GR activity lower than 18.1 U/mg protein (close to background of CL1-3 cells). Among them, 19 clones (28%) showed lower than 0.53-fold of parental GR activity. Ten (relative lower GR, < 0.4-fold) of these 19 clones were chosen to further examine their CTSS activities. Only one clone, #3-16 (GR-siRNA #3 transfected) from all selected clones showed one-fifth GR activity (very low) and 1.64-fold endogenous CTSS activity in comparison with that in CL1-0-mock cells. In Exp. II, vectors containing GR-siRNA#5 and puromycin gene were transfected into CL1-0 cells. Among 77 clones picked from puromycin containing medium, 38 clones (49.4%) had 1.1~2.0-fold of parental CTSS activity. Six clones among these high CTSS clones contained more than 1.5-fold of parental CTSS activity and less than one-half (0.16~0.38-fold) of parental GR activity (close to background of CL1-3 cells). Cell migration ability examination by means of wound healing assays and Boyden chamber assays was determined in clones 3-13, 3-16, 5-04 from Exp. I, clone 5-29 from Exp. II and parental CL1-0 cells, respectively. The results indicated that those clones from GR-knock down manipulation and high-CTSS selection have better cell migration abilities. Clone 3-13 [1.64-fold CTSS, 0.22-fold GR] and clone 5-29 [2.0-fold CTSS, 0.38-fold GR] showed 1.5-fold and 16-fold of parental cell migration ability. Therefore, clones containing higher CTSS activity (1.5~2.0-fold) and cell migration abilities than parental cells had been obtained from genetic manipulation to lower down GR activity in parental CL1-0 cells. In the second part of this study, CTSS inhibitors, L-trans-epoxysuccinyl- L-leucylamido (4-guanidino) butane (E-64), N-Ethylmaleimide (NEM) and phenylmethanesulfonyl fluoride (PMSF) were applied in CL1-3 cells. E-64 reduced 50% CTSS activity at 5 nM in vitro and 10 μM in vivo. In addition, NEM reduced 50% CTSS activity at 800 μM in vitro. However, there was no significant inhibition on CTSS activity after PMSF treatment. Therefore, E-64 was a considerable potent inhibitor. Although clone 3-13 had 0.22-fold of parental GR activity, clone 3-13 showed slightly SRB viability difference from parental CL1-0 cells after H2O2 exposure for 4 h. In contrast, CL1-3 cells, containing 0.5-fold GR activity of parental CL1-0 cells, showed significant different sensitivity to H2O2 (4 h at 25~50 μM dose or 1~4 h at 40 μM), from CL1-0 cells. CL1-3 cells have 6~7-fold of parental CTSS activity. Whether higher CTSS activity in CL1-3 cells further increased H2O2 sensitivity (in comparison with clone 3-13 cells) remained to be further investigated.
44
Shu-Ming et 莊書銘. « Characterize the effect of slit2 protein fragments on cell growth and migration in CL1-5 lung cancer cell line ». Thesis, 2008. http://ndltd.ncl.edu.tw/handle/51905152541360802314.
Texte intégralRésumé :
碩士中山醫學大學
醫學分子毒理學研究所
96
Our laboratory established suppressive subtractive cDNA libraries from tissues of a female lung adnocarcinnoma. After serial databases analysis, we identified slit2 gene whose expression level is highly repressed in lung cancer. Slit2 is a 200 kDa secreted glycoprotein. Its role in axon repellent and cell migration were extensively studied in different systems. Recent studies pointed that slit2 possesses tumor suppressor activity, however other report showed that slit2 has ability to induce angiogenesis. Therefore, the role of slit2 in carcinogenesis remains clarified. The expression of slit2 in lung cancer cell lines is very low except in the CL1 series cell line. The low invasive cell line CL1-0 expresses high level of slit2 while slit2 expression in high invasive cell line CL1-5 is very low. Since slit2 showed distinct biological activity in cell migration, and since slit2 expression is negatively correlated with invasive potential in CL1 series cell line, we thus use CL1 series cell line to study the role of slit2 in lung cancer. Our previous studies showed that overexpression of full length slit2 in CL1-5 reduce cell growth, which can be counted by increased in G1 phase cell population and decreased in S phase cell population. Furthermore, overexpression of slit2 decreased in vitro invasive ability of CL1-5 cell. In this study, we aimed to identify the domain of slit2 which confers inhibition of cell growth and/or invasion. To achieve this, we generated slit2-N terminal and slit2-C terminal fragments based on the reported cleavage site of slit2 in neuron system. Expression of slit2-C terminal fragment in CL1-5 showed similar growth repression activity compared with the full length slit2. In our system, we were unable to detect the cleavage product of overexpressed slit2. Nevertheless, it is interesting to investigate whether the cleavage of slit2 is important for its responsible function in CL1-5 or not. Our result showed that a non-cleavable slit2 showed similar growth inhibitory ability as the cleavable slit2. Therefore, the growth inhibitory ability of slit2 does not require cleavage at the reported cleavage site. Interestingly, overexpression of full length slit2 inhibited CL1-5 migration activity in wound healing assay, and this ability is retained in overexpressed slit2-C terminal fragment, while slit2-N terminal only slightly or did not inhibit cell migration. This result is rather surprising, since the ability to affect cell migration by slit2 in most system was referred to the slit2-N terminal domain. Invasive ability of cancer cell is often used to rank its malignancy. Thus, we are currently investigating the effect of slit2-N terminal and C terminal fragments in cell invasion. Once the function of each slit2 domain was confirmed, we will generate deletion mutations to narrow down the effective domain(s). Ultimately, we are hoping to identify small peptide fragment(s) which may inhibit cell growth or invasion.
45
Hui-Yi et 張彗怡. « Characterize the effect of Osteopontin splice variants on cell growth and invasion in CL1-5 lung cancer cell line ». Thesis, 2009. http://ndltd.ncl.edu.tw/handle/69850174392788003831.
Texte intégralRésumé :
碩士中山醫學大學
醫學分子毒理學研究所
97
Osteopontin (OPN) is a glyco-phosphoprotein independently discovered by investigators from diverse scientific backgrounds and implicated in a broad array of physiological and pathological processes. OPN exists both intra- and extracellularly and in numerous post-translational isoforms. OPN is subject to alternative splicing, which yields 3 messages, osteopontin-a (full-length), osteopontin-b (exon 5-) and osteopontin-c (exon 4-). High level of OPN expression has been found in a number of cancer types including breast, prostate, colon, head and neck, and hepatic cancers and lung cancer. Strong association of OPN expression with tumor metastasis has been established in various transformed cell lines, animal tumor models, and human cancers. These findings suggest that OPN is a key extracellular molecule involved in tumor development and progression. However, OPN splice variants have not been extensively evaluated in lung cancer. This study aimed to explore the roles of various splice forms of osteopontin in cell growth and invasion in lung cancer cell lines. The expression of OPN in lung cancer cell lines is very low except in the CL1-0,H460 and H1355 cell lines. The low invasive cell line CL1-0 expresses high level of OPN while OPN expression in high invasive cell line CL1-5 is very low. In this study, we tested the effect of OPN splice variants on growth and invasion in CL1-5 and A549 lung cancer lines. Our data showed that CL1-5 cells stably expressed OPN with deletion of exon 4 (OPN-exon4- ) or exon 5 (OPN-exon5-) had similar growth activity compared with vector control, while exogenous full-length OPN had the ability to inhibit cell growth. Interestingly, stable expression of exogenous OPN-exon4- enhanced in vitro invasive ability of CL1-5 cell compared to vector control, while OPN-exon5- did not change invasive ability. Unexpectedly, overexpression of full-length OPN inhibited CL1-5 invasion ability, and this ability is retained in contioned medium treatment. This result is rather surprising, since most studies showed that OPN enhanced, but not inhibited cell invasion. In summary, overexpression of full-length OPN reduced lung cancer cell growth and inhibited cell invasion. Overexpression of OPN-exon4- enhanced CL1-5 cell invasive ability, while OPN-exon5- has little effect on lung cancer cell growth and invasion. It is important to resolve which isoform of OPN is/are overexpressed in lung cancer and its microenvironment. Since OPN becomes a noticeable target for cancer therapy, it is important to explore the functional role of OPN splice variants in tumor growth and metastasis, and to dissect signaling pathways that involved in these mechanisms.
46
LIN, YI-WEN, et 林以雯. « The Study of Anti-Cancer Effect of Ligusticum chuanxiong Hort. Extract on Human Lung Cancer Cell Line CL1-5 ». Thesis, 2017. http://ndltd.ncl.edu.tw/handle/83344848217820500484.
Texte intégralRésumé :
碩士大葉大學
分子生物科技學系碩士班
105
Lung cancer mortality rates are the highest in the world at present disease, cancer deaths occur mainly due to cancer. There are currently many studies using natural medicine cancer metastasis and proliferation. Effect is a Traditional Chinese herb, effects have been developed to inhibit liver cancer, hyperplasia of pancreatic cancer, skin cancer, breast cancer, and promote apoptosis, but in the lung cancer study is unknow. We used lung cancer cells with high metastasis CL1-5 as a test template, with different concentrations of extracts of Ligusticum chuanxiong Hort., Ferulic acid, Tetramethylpyrazine cell lines and analyze some of the functional analysis, also used gene chips to compare different concentration effect of ferulic acid on lung cancer cell line CL1-5 gene expression after , and then through the DAVID functional annotation tool analysis downstream signal path.Results showed that Ligusticum chuanxiong Hort. extract and the biological activity of ferulic acid and tetramethylpyrazine in inhibition of lung cancer cell Proliferation, Migration, Mobility, Cell cycle, while also promoting Apoptosis. Ferulic acid can promote FAS signaling to achieve apoptosis effect; promote P21 signal CDK4/6, CDK2 was inhibited up to cell cycle inhibition in G0/G1Phase through inhibition of VEGF signaling that angiogenesis was inhibited; dampen TRAF2/5 and metastasis inhibition. These experiments have confirmed that extracts of Ligusticum chuanxiong Hort. and their biological activity of ferulic acid and tetramethylpyrazine in inhibition of lung cancer cell proliferation and metastasis, can also promote apoptosis. Key words: lung cancer, Ligusticum chuanxiong Hort., cell cycle, apoptosis, metastasis
47
Kuo, Chin-Jung, et 郭瑾融. « A non-covalent small inhibitor blocking β-tubulin:CCT-β complex induces apoptosis and suppresses migration and invasionin CL1-5 cells ». Thesis, 2019. http://ndltd.ncl.edu.tw/handle/yj45b7.
Texte intégralRésumé :
碩士國立臺灣大學
生化科學研究所
107
Previously, we reported the protein-protein interaction (PPI) between β-tubulin and CCT-β complex as a potential anti-cancer chemotherapeutic target. Through virtual screening, a compound 3112210 from Sigma-Aldrich compound bank was identified to be a reversible inhibitor of the PPI by docking into hot spots on this PPI interface of β- tubulin. In this study, 3112210 was tested on a highly metastatic non-small cell lung cancer (NSCLC) cell line, CL1-5. The co-IP experiments showed that, in 3112210-treated cancer cells, β-tubulin and CCT-β complex was disrupted. Furthermore, 3112210 caused CL1-5 cell death through ER stress and apoptosis. In addition to verifying its toxicity toward CL1-5, we performed migration and invasion assays using dosage at about IC20. The results indicated that 3112210 also inhibited cancer cell migration and invasion, and MMP-2, -9 were also inhibited. These anti-metastatic effects were endowed via integrin- related pathways and EMT transcriptional factors, as demonstrated by western blot experiments. To sum, 3112210 is a novel non-covalent inhibitor for β-tubulin:CCT-β complex in CL1-5 lung adenocarcinoma cells to induce cancer cell death and impeded cell metastasis.
48
陳美妤. « The role of integrin β3 and fibroblasts in OPN-a isoform mediated growth regulation in CL1-5 lung cancer cell line ». Thesis, 2011. http://ndltd.ncl.edu.tw/handle/86224130108290881934.
Texte intégralRésumé :
碩士中山醫學大學
醫學分子毒理學研究所
99
Osteopontin (OPN) is a glyco-phosphoprotein which is overexpressed in many cancers. The expressin of OPN is significantly correlated with tumor metastasis. Three OPN splicing variants, OPN-a, OPN-b and OPN-c, were identified and the expression of OPN-a is the highest among them in patients with lung cancer tissue and cancer cell lines. Different splicing variants of OPN have distinct impact on the growth and invasion of lung cancer cell lines. It is surprising that the full-length OPN (OPN-a) inhibited CL1-5 cell growth and invasion, and also inhibited invasion of A549 cells but had no effect on cell growth. We further confirmed that the differences of the effect on cell growth by OPN-a between CL1-5 and A549 was due to the diferential expression level of integrin β3 (ITGβ3). Knock-down the expression of ITGβ3 in CL1-5, blocked the effect of OPN-a mediated growth inhibition. On the other hand, overexpression of ITGβ3 in A549, exhibited OPN-a induced growth inhibition. Interestingly, in the absence of ITGβ3, OPN-a not only lost growth inhibition ability but promoted cell growth through NFκB pathway. Although ITGβ3 is required for OPN-a mediated growth inhibition, knock-down expression of ITG 3 almost completely inhibited growth of CL1-5 in the absence of OPN-a. Our results suggested that ITGβ3 is essential for the growth of CL1-5; however, ITGβ3 is no longer required if cells express OPN-a. Thus, the growth rate of CL1-5 may be subjected to the balance between OPN-a and ITGβ3 levels. The growth and invasion inhibitory roles of OPN-a in lung cancer cells are contradictory to the well recognized oncogenic role of OPN. Therefore, we hypothesized that the expression of OPN may promote cross-talk between cancer cells and its microenvironment. Our results showed that conditioned medium collected from fibroblasts treated with tumor secreted OPN-a greatly enhanced growth of CL1-5 cells, but had no effect on the ability of invasion. Thus, although OPN-a secreted by lung cancer cells would inhibit CL1-5 cell growth through autocrine pathway, the secreted OPN-a can activate fibroblast cells through paracrine pathway to secrete growth factor or cytokines that in turn promotes cancer cell growth. Moreover, cancer cells may reverse OPN-a mediated growth inhibition to growth enhancement by down-regulation the expression of ITGβ3. This study reveals two aspects on OPN-a regulated cancer cell growth: one is OPN-a mediated cross-talk between tumor and microenvironment; the other is regualated by the expression levels of different OPN receptors. Therefore, simutaneouly targeting several different receptors may be required in order to inhibit growth of lung cancer cells with overexperssed OPN-a.
49
Sz-Yu et 劉思妤. « Characterize the effect of different domain of slit2 C terminal on cell growth and invasion in CL1-5 lung cancer cell line ». Thesis, 2010. http://ndltd.ncl.edu.tw/handle/05297408913015078762.
Texte intégralRésumé :
碩士中山醫學大學
醫學分子毒理學研究所
98
Slit2 is a 200 kDa secreted glycoprotein. Its role in axon repellent and cell migration were extensively studied in different systems. Recent studies pointed that slit2 possesses tumor suppressor activity by inhibition of tumor growth and invasion. We discovered different exon15 splicing isoforms during slit2 cDNA cloning. With exon 15, Slit2-WT inhibited cell invasion of CL1-5 lung cancer cells, while without exon15, Slit2-△E15 not only inhibited cell invasion but also inhibited cell growth. Previous studies showed that Slit2 can be cleaved into two fragments in neuron cells: a 140 kDa slit2-N terminal fragment; and a 55-60 kDa slit2-C terminal fragment. Previously, we discovered that slit2-C terminal protein can inhibit cell growth and cell invasion. To identify which domain(s) is/are involved in Slit-△E15-mediated growth and/or invasion inhibition activity, we deleted individual domain on slit2-△E15 cDNA. Our studies showed that Slit2-△E15 lacking EGF6-LamG domains lost the ability to inhibit cell growth and cell invasion. To clarify whether the EGF6-LamG domains play a modulation role in Slit2-△E15-mediated growth and invasion inhibition indirectly or these domains possess direct role in growth and/or invasion inhibition? We constructed and expressed individual domain of slit2-C terminal. We found that the C-terminal region of LamG domain (LamG-C; a.a.(1228-1318)) by itself possessed ability to inhibit cell growth but not cell invasion. Our results also suggested that the N-terminal of LamG domain (LamG-N; a.a. (1182-1227)) has an effect on slit2-△E15-mediated cell invasion, but we were unable to resolve whether LamG-N inhibits cell invasion by itself. Thus, we are currently narrow-down the peptide domain in EGF6 and LamG regions, and identify the minimum peptide domain(s) that has/have ability to inhibit cell growth and invasion. Furthermore, we will investigate the receptor(s) involved in slit2-mediated growth and invasion inhibition.
50
Kuo, Yu-Ting, et 郭育廷. « Disrupting β-tubulin/CCT-β complexes induces apoptosis and suppresses migration and invasion of CL1-5 cells through MMP-2 and AKT/GSK-3β inhibition ». Thesis, 2017. http://ndltd.ncl.edu.tw/handle/4jgwn2.
Texte intégralRésumé :
碩士國立臺灣大學
生化科學研究所
105
We have previously demonstrated that I-Trp with an iodomethyl ketone warhead to alkylate Cys354 of β-tubulin, thereby disrupting the protein-protein interaction (PPI) of -tubulin with chaperonin-containing TCP-1β (CCT-β), causes cancer cell apoptosis [1]. In this study, we found that in CL1-5 cells, I-Trp activates both ER stress related proteins and proteasome activity to eliminate the imbalance proteins loading in ER, thereby mitigating ER stress, at the onset of β-tubulin/CCT-β complexes disruption. In addition, ER stress-associated apoptotic signaling is usually accompanied with intracellular Ca2+ release and the activation of MAPKs. We also observed the elevated intracellular Ca2+ levels, activation of MAPKs and caspase over-activation. Since CL1-5 cells are a highly metastatic lung cancer cell line, we assayed for its migration/invasion in the presence of I-Trp, where there were 80% survived cells to ensure most of the cells were not killed. The experimental results demonstrate the dose-dependent inhibition of CL1-5 cell migration and invasion. Furthermore, the mechanistic studies revealed that I-Trp inhibited phosphorylation AKT, and GSK-3β of CL1-5 cells, thereby downregulating MMP-2 expression and activation. In conclusion, this study reveals a novel therapeutic strategy potential for evoking apoptotic signaling by targeting β-tubulin/CCT-β complexes, and its anti-migration/invasion activity against human lung adenocarcinoma.