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1
PASINI, SILVIA. « Role of activated transcription factor 4 (ATF4) in learning and memory ». Doctoral thesis, Università degli Studi di Milano-Bicocca, 2011. http://hdl.handle.net/10281/27132.
Texte intégralRésumé :
The aim of this study is to understand the role of Activated Transcription Factor 4 (ATF4) in the processes of learning and memory. The topic of learning and memory has always aroused great interest from time immemorial and although a lot of researches have been focused on this subject for a long time, many mechanisms have not yet been fully understood.
Identifying the players and the mechanisms involved in learning and memory is of utmost importance because deficits in these cognitive functions are symptoms of common neurological diseases like stoke, depression, dementia and Alzheimer’s disease, one of the most wide spread neurodegenerative disease.
It has already been established that new gene expression and protein synthesis are required for long term memory, providing the basis to think that transcription factors may play a key role in these processes. Several studies have demonstrated the involvement of different transcription factors in memory formation such as cAMP response element binding protein (CREB), CCAAT enhancer binding protein (C/EBP), activated protein 1 (AP1), early growth response factor (Egr) and Rel/nuclear factor kB (Rel/NFkB).
Very little is known about the involvement of another transcription factor, Activated Transcription Factor 4. ATF4 is a member of the activated transcription factor (ATF)/cyclic AMP response element binding protein (CREB) family. It was originally described as a repressor of CRE-dependent gene transcription but recent studies have shown it to be a transcriptional activator. It is also a stress responsive gene, regulating the adaptation of cells to stress stimuli such as anoxia, endoplasmic reticulum stress and oxidative stress. ATF4 plays an essential role in development, and is particularly required for proper skeletal and eye development and is also involved in tumor progression and metastasis.
ATF4 has always been reported as a memory repressor that blocks new gene expression required for memory formation but no study has ever investigated it in a specific and direct way. The aim of this thesis is to study, in a specific and direct manner, the role of ATF4 in the processes of learning and memory. To reach this goal, ATF4 expression was modified in mouse hippocampi, the brain region mainly involved in learning and memory, with the injection of lentivirus carrying ATF4 gene, for the gain-of-function analysis, and lentivirus carrying shATF4, for the loss-of-function studies.
Before starting the experiments of ATF4 overexpression and downregulation, preliminary experiments were conducted to set up the injection coordinates to target the mouse hippocampi, to verify the lentiviral tropism and most importantly to evaluate the lentiviral spread, within the hippocampus, after the injection.
The consequence of ATF4 gain- and loss-of-function was then studied in the performance of standard behavioral tests such as Water Maze tests and Fear Conditioning, widely used to assess spatial and associative memory respectively. The behavioral test results showed that ATF4 protein overexpression enhances spatial memory, under the weak training paradigm in the Morris Water Maze test, and associative memory while ATF4 downregulation impairs spatial memory under the standard training condition.
After completing the behavioral tests, ATF4 overexpressed and downregulated mice were subjected to electrophysiological and neuronal spine analysis to verify if the alteration in cognitive functions, as a result of ATF4 modification, is supported by changes in synaptic potentiation and spine density and morphology.
Long Term Potentiation (LTP) is a long lasting enhancement in neuronal transmission and is widely considered as a cellular model of learning and memory in the central nervous system. The long-term memory impairment of ATF4 downregulated mice is supported by electrophysiological analysis, in which ATF4 downregulated slices showed an impairment in LTP. Unexpectedly, LTP impairment was also found in ATF4 overexpressed slices, maybe due to the difference in the time between the injection and the behavioral tests or the electrophysiological recordings.
Most of the intracellular pathways responsible for LTP require new gene expression and protein synthesis. This, in turn, leads to morphological changes required to sustain the enhancement of signal transmission. One of these morphological changes is the modification of the density and the morphology of dendritic spines. ATF4 up- and downregulation in hippocampal neurons does not affect spine density but ATF4 overexpression causes a significant increase in the percentage of mushroom spines as compared to that found after ATF4 downregulation. Mushroom spines with a large head are the most stable neuronal spines and contribute to strong synaptic connections, hence it has been hypothesized that they represent the “memory spines”.
Collectively, these results support the hypothesis that the transcription factor ATF4 plays a positive role in synaptic plasticity and memory formation. Further studies need to be done to understand the molecular mechanisms through which ATF4 acts.
This thesis represents only a step on the road towards understanding the complicate mechanisms of learning and memory, not forgetting that the most important discoveries were the result of small knowledge acquired step by step.
2
Kaikkonen, L. (Leena). « p38 mitogen-activated protein kinase and transcription factor GATA-4 in the regulation of cardiomyocyte function ». Doctoral thesis, Oulun yliopisto, 2014. http://urn.fi/urn:isbn:9789526205038.
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Abstract Cardiovascular diseases are the leading causes of death in the developed countries and their incidence is not expected to decrease in the future. There is a lifetime risk of one in five of developing heart failure, which still has poor prognosis and current treatments only cover part of the pathophysiology behind this syndrome. Pathological processes contributing to heart failure include cardiac hypertrophy and remodeling, which involves neurohumoral activation, reactivation of fetal genes, impaired Ca2+ cycling, increased apoptosis, and increased fibrosis. Intracellular signalling pathways and transcription factors mediating the response to various extracellular stresses have a key role in the regulation of myocardial remodeling and they are investigated in order to develop new approaches for the treatment of heart failure. The aim of this thesis was to elucidate roles of mitogen-activated protein kinases (MAPKs) and transcription factor GATA-4 in the regulation of cardiomyocyte function in cell cultures, and in hearts ex vivo and in vivo. The main findings were that (i) Inhibition of p38α MAPK enhanced function of sarco/endoplasmic reticulum Ca2+ -ATPase and thus cardiac contractility by increasing phosphorylation of protein phosphatase inhibitor-1 and phospholamban, (ii) p38 MAPK isoforms p38α and p38β regulated promoter activity of B-type natriuretic peptide via distinct pathways, (iii) p38α and p38β MAPKs also had different effects on gene expressions related to fibrosis and hypertrophy, and (iv) p38 and ERK1/2 MAPKs mediated stretch-induced activation of GATA-4 by phosphorylation at Ser 105. GATA-4 also seems to be regulated by ubiquitination. This study provides novel data of p38 MAPK and GATA-4 in the regulation of cardiomyocyte function. Inhibition of p38α MAPK could be beneficial in the treatment of heart failure. Also GATA-4 is a potential target for treatment of cardiovascular diseasesTiivistelmä Sydän- ja verisuonisairaudet ovat yleisin kuolinsyy länsimaissa, eikä niiden ilmaantuvuus tule vähenemään lähitulevaisuudessa. Elinikäinen riski sairastua sydämen vajaatoimintaan on 20 %, ja sydämen vajaatoiminnan ennuste on edelleen huono. Nykyisillä hoitomuodoilla voidaan puuttua vain osittain sydämen vajaatoiminnan patofysiologisiin mekanismeihin. Sydämen vajaatoiminnan kehittymiseen liittyvät sydänlihaksen liikakasvu ja uudelleenmuovautumisprosessi, johon liittyy neurohumoraalinen aktivaatio, sikiöaikaisten geenien uudelleenilmentyminen, häiriöt solunsisäisessä Ca2+-viestinnässä sekä lisääntynyt ohjelmoitu solukuolema ja sidekudoksen muodostuminen sydämeen. Solunsisäisillä viestinvälitysketjuilla sekä transkriptiotekijöillä, jotka vastaavat solunulkoisten ärsykkeiden välittämisestä solun sisällä, on keskeinen rooli edellämainittujen prosessien säätelyssä. Uusien lähestymistapojen kehittäminen sydämen vajaatoiminnan hoitoon edellyttää myös solunsisäisen viestinvälityksen ja geenien säätelyn mekanismien selvittämistä. Tämän väitöstyön tavoite oli selvittää p38 mitogeeniaktivoituvan proteiinikinaasin (p38 MAPK) ja transkriptiotekijä GATA-4:n merkitystä sydämen vajaatoiminnan patogeneesissä soluviljelymalleissa. Päälöydöksiä olivat: (i) p38α MAPK -isoformin estäminen paransi kalsiumia solulimakalvostoon pumppaavan SERCA2a:n toimintaa ja sydänlihassolun supistumiskykyä lisäämällä fosfolambaanin ja proteiinifosfataasi-inhibiittori-1:n fosforylaatiota. (ii) p38 MAPK isoformit p38α ja p38β säätelivät B-tyypin natriureettisen peptidin geenin promoottorialuetta erillisten reittien kautta. (iii) p38α ja p38β isoformit vaikuttivat myös eri tavoin sydämen sidekudoksen muodostumiseen ja hypertrofiaan liittyvien geenien ilmentymiseen. (iv) p38 ja ERK1/2 välittävät venytyksen aiheuttaman GATA-4:n aktivaation fosforyloimalla seriini-105 fosforylaatiopaikan. Lisäksi GATA-4:n toimintaa säädellään ubiquitinaation avulla. Tämä tutkimus tuo uutta tietoa p38 MAPK:n ja GATA-4:n rooleista sydämen vajaatoiminnan kehittymisessä. p38α-isoformin toiminnan estäminen voisi olla hyödyllinen hoitomuoto sydämen vajaatoiminnassa. Myös GATA-4 on potentiaalinen lääkehoidon kohde sydänsairauksien hoidossa
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Arpiainen, S. (Satu). « Transcriptional regulation of the hepatic cytochrome P450 2a5 gene ». Doctoral thesis, University of Oulu, 2007. http://urn.fi/urn:isbn:9789514285653.
Texte intégralRésumé :
Abstract
Cytochrome P450 (CYP) enzymes are the major metabolizers of xenobiotics, e.g. drugs, and environmental toxins. Thus, changes in CYP expression have an important impact on drug metabolism and susceptibility to chemical toxicity.
In the present study, the transcriptional mechanisms of both constitutive and inducible regulation of the Cyp2a5 gene in mouse liver were investigated. Mouse primary hepatocyte cultures were used as the main model system together with cell and molecular biology methods.
The key activation regions of the Cyp2a5 5' promoter were determined using reporter gene assays. Two major transcription activation sites of the Cyp2a5 5' promoter, called the proximal and the distal, were found. Transcription factors hepatocyte nuclear factor-4 (HNF-4) and nuclear factor I were shown to bind to the proximal promoter. Aryl hydrocarbon receptor nuclear translocator (ARNT) and upstream stimulatory factor bound to a common palindromic E-box element in the distal promoter region. All three response elements were shown to be essential for constitutive expression of CYP2A5 in murine hepatocytes. ARNT appeared to control Cyp2a5 transcription without a heterodimerization partner suggesting active involvement of the ARNT homodimer in mammalian gene regulation.
Aryl hydrocarbon receptor (AHR) ligands were shown to induce Cyp2a5 transcriptionally by an AHR-dependent mechanism, and established Cyp2a5 as a novel AHR-regulated gene. The AHR response element and the E-box, identified in these studies, were located near to each other and close to a separately defined nuclear factor (erythroid-derived 2)-like 2 binding site in the distal region of the Cyp2a5 promoter, suggesting cooperation between these elements.
Peroxisome proliferator-activated receptor gamma coactivator-1α was shown to up-regulate Cyp2a5 transcription through coactivation of HNF-4α. This indicates that xenobiotic metabolism can be regulated by modification of co-activation.
The present results show that CYP2A5 is regulated by several different cross-regulatory pathways. The regulatory mechanisms involved in the transcription of the Cyp2a5 gene may also control other CYP genes, especially the human ortholog CYP2A6, and may explain some of the individual variations in the metabolism of xenobiotics.
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Houlard, Martin. « Étude de l'adressage et de l'implication nucléaire du proto-oncogène Vav-1 ». Paris 7, 2002. http://www.theses.fr/2002PA077098.
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Mak, Man-chi. « Characterization of transcription factor nuclear factor of activated T-cells 5, in knockout embryos and mice ». Click to view the E-thesis via HKUTO, 2008. http://sunzi.lib.hku.hk/hkuto/record/B41633684.
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Arabanian, Laleh Sadat. « Role of NFAT (Nuclear Factor of Activated T Cells) Transcription Factors in Hematopoiesis ». Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2012. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-99739.
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Understanding the transcriptional mechanisms that control hematopoiesis and the interaction between hematopoietic stem cells and the bone marrow (BM) microenvironment in vivo is of considerable interest. The calcineurin-dependent transcription factor NFAT (Nuclear Factor of Activated T cells) is known as master regulator of cytokine production in T lymphocytes and therefore central for T cell-dependent immune reactions, but has also been shown to regulate a process of differentiation and tissue adaptation in various cell types. The activation of NFAT is dependent on the calcium level within the cell. In resting cells, calcium levels are low and NFAT is cytoplasmic and inactive. A sustained increase in the internal calcium concentration within an external stimuli leads to activation of the calcium-dependent calcineurin, followed by dephosphorylation and nuclear translocation of NFAT. We have previously shown that NFATc2, a member of the NFAT family, is expressed in CD34+ hematopoietic stem cells (HSC). A mouse model harboring NFATc2 deficiency provides the opportunity for in vivo investigation of the role of NFATc2 in hematopoiesis.
Our recent observations showed that aged mice lacking the transcription factor NFATc2 develop peripheral blood anemia and thrombocytopenia, BM hypoplasia and extramedullary hematopoiesis in spleen and liver. The proliferation and differentiation of NFATc2-deficient hematopoietic stem cells ex vivo, however, was found to be intact. It remained therefore unclear whether the disturbed hematopoiesis in NFATc2-deficient mice was caused by the hematopoietic or the stroma component of the BM hematopoietic niche. In the current study we dissected the relative contribution of hematopoietic and stroma cells to the phenotype of the NFATc2-deficent mice by transplanting immuno-magnetically purified NFATc2-deficient (KO) HSCs to lethally irradiated wild type (WT) mice, and vice versa. After a post-transplantation period of 6-8 months, peripheral blood, BM as well as spleen and liver of the transplanted animals were analyzed and compared to WT and KO mice transplanted with control cells.
Transplantation of NFATc2-deficient HSCs into WT recipients (KO WT) induced similar hematological abnormalities as those occurring in non-transplanted KO mice or in KO mice transplanted with KO cells (KO KO). Compared to WT mice transplanted with WT cells (WT WT), KO WT mice showed evidence of anemia, thrombocytopenia and a significantly reduced number of hematopoietic cells in their BM. Likewise, KO WT mice developed clear signs of extramedullary hematopoiesis in spleen and liver, which was not the case in WT WT control animals. In addition to the hematopoietic abnormalities, transplantation of NFATc2-deficient HSC also induced osteogenic abnormalities such as BM sclerosis and fibrosis in WT mice. This phenomenon was rather subtle and of incomplete penetrance, but never seen in mice transplanted with WT cells. These data demonstrate for the first time, that the NFATc2 transcription factor directly regulates the intrinsic function of hematopoietic stem cells in vivo. However, the transcriptional targets for NFAT in these cells are yet unknown. In addition to hematopoietic stem cells, NFATc2 has been shown to be expressed in a lineage-specific manner during myeloid differentiation and, notably, is maintained during megakaryopoiesis while it is suppressed during the differentiation of neutrophils. Bone marrow megakaryocytes are the precursors of peripheral blood platelets and therefore constitute an integral part of primary hemostasis, thrombosis and wound healing. The biological role of NFAT in megakaryocytes is unknown. We have recently shown that NFATc2 is not necessary for megakaryocytic differentiation. On the other hand, recent evidence suggests that NFATc2 is required for the transcription of specific megakaryocytic genes. In this study, we showed that activation of the calcineurin/NFAT pathway in either primary megakaryocytes or CMK megakaryocytic cells forces the cells to go into apoptosis. Cell death in megakaryocytes is induced by treating the cells with the calcium ionophore ionomycin and suppressed by either the pan-caspase inhibitor zVAD or the calcineurin inhibitor cyclosporin A (CsA). Ionomycin stimulation of megakaryocytes leads to the expression of Fas Ligand (FASLG), a pro-apoptotic member of the tumor necrosis factor superfamily. Expression of FASLG was detectable as early as four hours after stimulation on the membrane of ionomycin-treated megakaryocytes, was augmented in cells stably overexpressing NFATc2, and was suppressed in cells either pretreated with CsA or expressing the specific peptide inhibitor of NFAT, VIVIT.
To investigate the physiological relevance of FASLG expression on megakaryocytes, we performed co-cultures of megakaryocytes with Fas-expressing T-lymphocytes, in which CMK cells were left either unstimulated or pre-stimulated with ionomycin and then added to Jurkat cells. The presence of ionomycin-stimulated CMK cells, but not of unstimulated cells or cells stimulated in the presence of CsA, significantly induced apoptosis in Jurkat cells. Overexpression of NFATc2 in CMK cells enhanced their potency to induce apoptosis in Jurkat cells, while cells expressing VIVIT were less effective. Apoptosis induction of Jurkat cells by stimulated CMK cells was partially blocked by the presence of either a neutralizing antibody against FASLG or an antagonistic antibody to Fas during the co-culture period, indicating involvement of the FASLG/Fas apoptosis pathway. These results represent the first clear evidence for a biological function of the calcineurin/NFAT pathway in megakaryocytes, namely the regulation of Fas/FASLG-dependent apoptosis. Second, they underline that the biological role of megakaryocytes is not restricted to the production of proteins and other cellular structures for platelet assembly, but that this population of cells fulfills an independent regulatory function in the context of the surrounding tissue.
Finally, we have identified by RNA sequencing analysis of NFATc2-expressing and -deficient cells, the entire set of genes which is induced by NFATc2 in stimulated megakaryocytes. Functional pathway analysis suggests an involvement of NFATc2 in pro-inflammatory pathways in these cells. The significance of these findings has to be addressed in further studies.
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Ulrich, Jason Daniel. « The regulaton and function of nuclear factor of activated T-cells in neurons ». Diss., University of Iowa, 2011. https://ir.uiowa.edu/etd/2782.
Texte intégralRésumé :
Ca2+-dependent transcription is a fundamental process by which neurons translate activation experience into cellular level adaptations. The nuclear factor of activated T-cells (NFAT) family of proteins comprise four Ca2+/CaN-dependent transcription factors that are widely expressed throughout virtually all tissues. Within neurons, NFAT dependent signaling is critical for axonal development, regulation of synapse number and efficacy, and survival. Furthermore, NFAT is implicated in activity dependent regulation of genes involved in synaptic transmission, learning and memory, mood, and pain sensation. NFAT is activated upon elevations in intracellular Ca2+, which results in CaN -dependent dephosphorylation of multiple serine residues within an N-terminal regulatory region. NFAT dephosphorylation permits NFAT translocation to the nucleus, where it can regulate gene expression, frequently co-operatively with other transcription factors, including AP-1 and MEF2. NFAT activation is opposed or terminated by several kinases, including CK1 and GSK3. Despite the importance of NFAT proteins as regulators of Ca2+-dependent transcription, little is known about the regulation and function of specific NFAT isoforms within neurons.
In Aim 1 of this thesis I characterized the differential activation of NFATc3 and NFATc4 in DRG neurons. While NFATc3 rapidly translocates the nucleus upon Ca2+-influx through voltage-gated calcium channels, NFATc4 remained remarkably intransient. Modular substitution of NFATc3 regulatory elements increased the rate or retention of NFATc4, whereas converse substitutions of NFATc4 regulatory elements into NFATc3 decreased NFATc3 nuclear translocation. The activation of NFATc4 appears to be inhibited by preferential phosphorylation by kinases, such as GSK3, which counteract CaN-dependent dephosphorylation. In Aim 2 I investigated the role of NFATc3 in hippocampal neurons. While the majority of NFAT reports in neurons have focused on NFATc4, my data suggest that NFATc3 is the predominantly expressed isoform in hippocampal neurons and is critical for depolarization-induced NFAT target gene expression. I further characterized NFATc3 KO mice in a battery of behavioral assays to test whether loss of NFATc3 expression would affect the baseline anxiety/depression state of the animal, or if NFATc3 was critical for learning and memory. Taken together, my data suggest that NFATc3 is important for NFAT-dependent gene expression in central and peripheral neurons and that distinct regulation of NFAT isoforms within neurons may underlie isoform-specific effects on gene expression.
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Rosso, Michele <1984>. « Role of the Transcription Factor Sox2 in the Osteogenic Lineage ». Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6489/4/rosso_michele_tesi.pdf.
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The Sox2 transcription factor is modified by sumoylation at the K247 position although the addition of SUMO1 and Pias1 promotes the sumoylation of Sox2 at the additional K123 site. The role of sumoylation on Sox2 biological functions was analyzed by comparing the activity of WT and sumoylation mutants on the transcription of the FGF4 gene in HeLa cells and on the downregulation of the Wnt pathwayvin 293T cells. When SUMO1 and PIAS1 promote the sumoylation of WT Sox2, the transcriptional activity of the FGF4 promoter is inhibited showing that Sox2 sumoylation is necessary for the repression function. However, there is no effect of Sox2 sumoylation on β-Catenin activity.
Since we were interested in osteoblast differentiation we set up an inducible system for Sox2 in primary osteoblasts. Following Sox2 doxycycline induction, 158 genes were differentially expressed: 120 up-regulated and 38 down-regulated. We annotated as direct Sox2 targets a number of genes involved in osteoblast biology and we further analyzed 3 of them involved in the BMP pathway. The results show that Sox2 regulates the BMP pathway without affecting SMAD phosphorylation, and that Sox2 sumoylation is not necessary for this function.
We also found that genes involved in the Hippo pathway were direct Sox2 targets. As the Hippo pathway is activated by Sox2 and Sox2 interacts with the NF2 promoter, we checked the effect of Sox2 on the expression of NF2. We showed that Sox2 down-regulates the transcriptional activity of the NF2 promoter, allowing the transcription of the YAP/TEAD genes in osteoblasts, thus acting as an upstream regulator of the Hippo pathway. We conclude that Sox2 induction in osteoblasts triggers FGF dependent inhibition of the BMP, Wnt and Hippo pathways.
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Brehm, Alexander Jorg Georg. « Octamer-dependent transcriptional activation by the embryonal transcription factor Oct-4 ». Thesis, Open University, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.338344.
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Hilton, Traci Leigh. « Dual function of TAF1 in basal and activated cyclin D1 transcription / ». Thesis, Connect to this title online ; UW restricted, 2003. http://hdl.handle.net/1773/6275.
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Tong, Ho-kwan. « Functional regulation of the forkhead box M1 transcription factor by Raf/MEK/MAPK signaling ». Click to view the E-thesis via HKUTO, 2006. http://sunzi.lib.hku.hk/hkuto/record/B37654597.
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Tong, Ho-kwan, et 湯皓鈞. « Functional regulation of the forkhead box M1 transcription factor by Raf/MEK/MAPK signaling ». Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B37654597.
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Zazzi, Henric. « Human insulin-like growth factor binding protein -4 and -6 : gene structure and transcription regulation / ». Stockholm, 2000. http://diss.kib.ki.se/2000/91-628-3873-3/.
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Traboulsi, Hussein. « Relationship between the transcription/DNA repair factor TFIIH and the Peroxisome Proliferator-Activated Receptor coactivator 1α PGC-1α ». Strasbourg, 2011. http://www.theses.fr/2011STRA6063.
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Des mutations dans la sous unité XPD du facteur de transcription IIH (TFIIH) conduit à des maladies génétiques rares incluant trichothiodystrophy TTD. Connaissant que la sous unité MAT1 de TFIIH est nécessaire pour une activité complète de l’activateur proliferator activated receptor g coactivator 1a (PGC-1a), on a décidé de regarder l’influence de la mutation R722W dans XPD sur l’activité de PGC-1a dans les hépatocytes TTD. Nos résultats montrent que le facteur transcriptionnel TFIIH est nécessaire pour l’activité hépatique du cofacteur PGC-1α dans la voie de la néoglucogenèse. En plus on montre que la mutation R722W dans la sous unité XPD du TFIIH perturbe l’interaction du PGC-1α avec SIRT1, dérégulant par ca l’activité du PGC-1α. Etant PGC-1α un régulateur important du métabolisme du glucose et des lipides, nos résultats permet donc de suggérer un lien entre les symptômes liés à un défaut du métabolisme observés chez les patients TTD et le dérèglement du comportement du PGC-1α. Des études ultérieures seront nécessaires pour mieux comprendre le mécanisme d’activation du PGC-1α par TFIIH, surtout voir si ca implique un lien directe entre SIRT1 et TFIIH. Il est ainsi important d’analyser si d’autres voies du PGC-1α sont aussi affectéesMutations in the subunit XPD of the transcription factor IIH (TFIIH) lead to genetic disorders including trichothiodystrophy TTD. Knowing that subunit MAT1 of TFIIH is required for the full function of the proliferator activated receptor γ coactivator 1 α (PGC-1α), we decided to study the influence of the mutation R722W in XPD on PGC-1α activity. Using immortalized hepatocytes isolated from TTD mutant mice, we investigated the expression of PGC-1α target genes involved in gluconeogenesis. We observed that these genes are downregulated in TTD hepatocytes. Moreover, we found that XPD mutation disrupted the interaction between PGC-1α and the deacetylase Sirtuin1 (SIRT1) leading to reduced activation of PGC-1α. We also showed that PGC-1α and SIRT1 both interact with TFIIH. We thus established that TFIIH is required for full PGC-1α activity in the gluconeogenesis pathway, and more likely through modulation of SIRT1 activity. Therefore, our results suggest that the deregulation in TTD likely results from the inability of the mutated TFIIH to fully participate in recruitment of PGC-1α and/or SIRT1 to the DNA
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Chan, Jonathan Ka Lok. « Functional characterization of the regulation of transcription factor MEF2C by histone acetyltransferase p300 and histone deacetylase 4 / ». View abstract or full-text, 2004. http://library.ust.hk/cgi/db/thesis.pl?BICH%202004%20CHANJ.
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Thesis (Ph. D.)--Hong Kong University of Science and Technology, 2004.Includes bibliographical references (leaves 135-159). Also available in electronic version. Access restricted to campus users.
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De, Silva Jasenthu L. P. « The role of the transcription factor FOXP1 in the immune response to breast cancer ». Doctoral thesis, Universite Libre de Bruxelles, 2018. https://dipot.ulb.ac.be/dspace/bitstream/2013/264152/4/Thesis.pdf.
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Breast cancer (BC) was not initially considered an immunogenic tumor; however, recent data show that immune-related factors are associated with patient prognosis and the response to treatment. Several large adjuvant clinical trials have shown that tumor infiltrating lymphocytes (TIL) are significantly associated with a better prognosis and can also predict responsiveness to pre-operative chemotherapy, particularly in the triple negative (TN) & HER2+ BC subtypes (Carsten Denkert et al. 2010; Loi et al. 2013a). Recently, the presence of ectopic lymph node-like structures characterized by distinct T and B cell zones, called tertiary lymphoid structures (TLS), were identified adjacent to the tumor (Gu-Trantien et al. 2013) in 60% of BC (Buisseret et al. 2017b) and linked with a good prognosis (Gu-Trantien et al. 2013). The mechanisms involved in TLS formation and activities and their impact on tumor immunity is relatively unknown. TIL infiltration and TLS formation are likely regulated, in part, by transcription factors (TF) that control cytokine/chemokine production within the tumor microenvironment (TME) (Pimenta and Barnes, 2014). One such TF, the forkhead box protein 1 (FOXP1) is abnormally expressed in various human tumors and has a known role in regulating immune cell functions. Contradictory data on FOXP1 expression together with a lack of information on its immune regulation led us to explore its role in this tumor type. The first part of this thesis research focused on FOXP1-mediated regulation in BC. Gene/protein analysis was examined in the four BC molecular subtypes, revealing its enriched expression in estrogen receptor positive (ER+) tumors (Luminal A/B). Luminal BC is generally less infiltrated compared with frequently high TIL infiltration in ER negative (ER-) tumors (i.e. HER2+ and TN) [reviewed in (Solinas et al. 2017a) and (Loi et al. 2014)]. We found that high FOXP1 expression in a cohort of untreated primary BC was significantly associated with a lower TIL and fewer TLS compared to FOXP1 low (FOXP1lo) tumors. This observation led us to investigate the effect of FOXP1 on cytokines and chemokines potentially involved in TIL recruitment and/or TLS formation. BC cancer cell lines were used to silence [MCF7; FOXP1hi] or overexpress [MDA-MB-231; FOXP1lo] FOXP1 expression. FOXP1 repression upregulated a number of cytokines and chemokines involved in T and B cell migration and function, while FOXP1 overexpression repressed a majority of the same factors. Expression analysis of the major T and B cell cytokine and chemokine genes was performed for FOXP1lo and FOXP1hi primary BC. These data reveal that FOXP1hi BCs have significant decreases in CXCL9, CXCL10, CXCL11, CXCL13, CX3CL1, CCL20, IL2, IL21, granzyme B and IFNγ and high levels of the immunosuppressive cytokines, IL10 and TGFβ. We next performed a lymphocyte migration assay using primary tumor supernatants prepared from FOXP1lo and FOXP1hi BC finding significantly decreased migration of total CD45+ lymphocytes, B cells, helper (CD4+) and cytotoxic (CD8+) T cells using FOXP1hi compared to FOXP1lo SN. Overall, our data suggest that FOXP1 plays an important role in repressing anti-tumor immune responses by negatively regulating TIL migration directed by specific cytokines and chemokines.The second part of this thesis research focused on the role FOXP1 plays in BC TLS. FOXP1 expression in T and B cell TIL and TLS was evaluated using RT-qPCR, multicolor flow cytometry, immunofluorescence (IF) and immunohistochemistry (IHC) and fresh, fixed and frozen breast tissues. Based on the FOXP1 expression two types of TLS were identified in BC: 1) TLS containing a germinal center (GC-TLS) and 2) TLS lacking a GC (non-GC-TLS). Examination of proteins specifically associated with active humoral immune responses allowed us to identify GC-TLS but not non-GC-TLS as functional. Gene expression analysis of micro-dissected tissues revealed distinct immune profiles that characterize B cell follicles in tonsils and spleen as well as aggregates, non-GC-TLS and GC-TLS in BC. This analysis further demonstrates that ongoing cell-mediated immune responses are associated with GC-TLS. The findings from this thesis research add important information to our understanding of how immune responses are initiated and maintained in BC and provide further insight into the identification and organization of functional immune responses at the tumor site.Doctorat en Sciences biomédicales et pharmaceutiques (Médecine)
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Kőszegi, Dávid. « RKD genes : a novel transcription factor family involved in the female gametophyte development of Arabidopsis and wheat ». kostenfrei, 2008. http://nbn-resolving.de/urn:nbn:de:gbv:3:4-823.
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Malenfant, Daniel. « Étude des fonctions développementales et métaboliques du récepteur nucléaire fetoprotein transcription factor (FTF) ». Thesis, Université Laval, 2012. http://www.theses.ulaval.ca/2012/28755/28755.pdf.
Texte intégralRésumé :
Le récepteur nucléaire Fetoprotein Transcription Factor (FTF) identifié par notre laboratoire et exprimé principalement dans le système digestif est un régulateur important du métabolisme des lipides et des stéroïdes, de la prolifération cellulaire et du développement embryonnaire. Plusieurs groupes ont constaté que l’influence du récepteur FTF sur la synthèse de stéroïdes et la régulation du cycle cellulaire stimule la prolifération tumorale de cellules d’origine tissulaire diverse. Mes études de doctorat ont porté sur l’expression tissulaire de FTF, sur la caractérisation d’un nouvel élément régulateur de son promoteur et sur l’identification par immunoprécipitation de chromatine (ChIP-chip) des cibles transcriptionnelles de FTF dans le foie de souris fœtale et adulte et dans les cellules d’hépatome humain. Ces études ont permis de mieux définir le rôle métabolique de FTF ainsi que son rôle développemental et son implication potentielle dans la carcinogenèse hépatique. L’expression de FTF par les organes du système digestif et par certaines structures nerveuses, sa régulation par des récepteurs nucléaires métaboliques et sa liaison aux promoteurs de multiples enzymes et transporteurs impliqués dans le métabolisme énergétique placent FTF dans une position clé dans l’homéostasie métabolique et énergétique de l’organisme. Le facteur de transcription C/EBPpartenaire de FTF au promoteur de l’AFP et impliqué lui aussi dans le développement hépatique et le métabolisme énergétique, est lié au promoteur de 20% des cibles transcriptionnelles de FTF. De plus, C/EBP lie le promoteur de FTF formant ainsi une autre boucle activatrice s’ajoutant au réseau transcriptionnel hépatique. Dans les cellules d’hépatome, FTF lie les promoteurs de plusieurs gènes impliqués dans la prolifération et le maintien des cellules tumorales, soit des régulateurs de la réplication, de la croissance et de l’apoptose cellulaire. FTF fait donc partie intégrante du réseau transcriptionnel hépatique régissant le développement et la différenciation hépatique et le maintien du métabolisme énergétique chez l’adulte et est vraisemblablement impliqué dans la promotion de la cancérogenèse hépatique.FTF is a nuclear receptor principally expressed in adult digestive organs that has been shown to act as a major regulator of lipids and steroids metabolism, cellular proliferation and embryonic development. FTF involvement in steroid synthesis and cell cycle regulation tends toward the stimulation of tumor proliferation in neoplasic tissues in which FTF is expressed. However, more studies of FTF function in normal and disease states and on its regulation are needed to draw a complete picture of FTF activity in cell physiology. Within the context of my studies, I delineated the FTF adult and fetal tissular expression, characterized a novel Ftf promoter element and identified FTF direct hepatic transcriptional targets in fetal, adult and tumor cell lines by using chromatin immunoprecipitation (ChIP-on-chip). These studies defined new FTF functions in metabolism, fetal development and hepatic carcinogenesis. FTF expression in digestive system and in neural structures controlling eating behavior, its transcriptional regulation by metabolic nuclear receptors and its binding to enzyme and transporter gene promoters driving energy metabolism, puts FTF in a key location for governing cellular and organismal energy metabolism. C/EBP, a transcriptional FTF partner on the Afp gene promoter and also involved in energy metabolism, is bound to 20% of the FTF targets including FTF itself thus adding branches to the complex hepatic transcriptional network. In hepatoma cells, FTF binds to proliferation and tumor cell maintenance genes like replication, growth and apoptosis regulators. Therefore, FTF belongs to the hepatic transcription network that governs hepatic development, differentiation and adult energy metabolism and is likely to be involved in promoting hepatic tumorogenesis.
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Wong, Andrew Hoi-Tao 1974. « Interdependence of the double-stranded RNA-activated protein kinase, PKR, and the transcription factor, STAT1, in intgerferon signalin and translatioinal control ». Thesis, McGill University, 2000. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=36850.
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Interferons (IFNs) are polypeptides that protect the body from microbial infection by the activation of genes that inhibit the replication of such pathogens. STAT1 is a transcription factor that is activated by IFNs and virus infection, and propagates signals to the nucleus by binding DNA enhancer elements to induce the expression of IFN-stimulated genes (ISGs). One such gene product is the double-stranded RNA (dsRNA)dependent protein kinase, PKR. PKR is an important determinant of host resistance since it has the ability to bind dsRNA, an intermediate produced during viral replication, and to phosphorylate the eukaryotic translation initiation factor-2alpha (eIF-2alpha), a modification that inhibits of protein synthesis. Additionally, recent findings have shown that PKR can modulate various signaling pathways. The objective of this research was to understand the involvement of PKR in the IFN signaling pathways. We observed that the catalytic domain of PKR interacts with the DNA-binding region of STAT1 in vitro and in vivo. Overexpression of catalytic mutants of PKR blocked STAT1 DNA-binding and ISG expression upon IFN or dsRNA treatment. The capacity of catalytic mutants of PKR to negatively regulate STAT1 DNA-binding is attributed to its ability to interact with STAT1 since in cells lacking PKR or expressing an RNA-binding mutant of PKR, unable to associate with STAT1, STAT1 displays augmented activity. A mutant of STAT1 (TM) unable to interact with PKR displayed elevated biochemical and biological activities; findings consistent with the role of PKR in regulating STAT1 function. Alternatively, the interaction between PKR and STAT1 has the ability to inhibit PKR activity in vitro and in vivo; a phenomenon not observed with STAT1 TM.In parallel, PKR was also found to interact with TYK2, the upstream activator of STAT1. IFN treatment promoted the dissociation of PKR from TYK2 in a manner dependent on TYK2 kinase activity. Furthermore, we observed that TYK2 phosphorylates the amino terminus of PKR in vitro and in vivo on tyrosine. Taken together, PKR appears to function as an important scaffolding protein for the IFN signaling pathways. Alternatively, PKR activity itself is also regulated at multiple steps along this pathway. Thus, it appears that there is an intimate cross-talk between components of the JAK/STAT pathway and translational machinery.
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Malabanan, Kristine Paz Centre for Vascular Research Faculty of Medicine UNSW. « Roles of activation transcription factor 4 (ATF4) and YrdC in the response of vascular smooth muscle cells to injury ». Publisher:University of New South Wales. Centre for Vascular Research, 2008. http://handle.unsw.edu.au/1959.4/41338.
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Neointimal proliferation is a key process underlying many cardiovascular diseases such as atherosclerosis and angioplasty-induced restenosis. Vascular smooth muscle cells (SMC) are significant contributors to the development and stability of the neointimal lesion. This is due, in part, to their capacity to be phenotypically modulated, facilitating SMC proliferation in response to mechanical injury, their subsequent migration, and deposition of extracellular matrix. The aim of this thesis was to characterize the function of two genes identified in our laboratory to be upregulated shortly after mechanical injury of vascular SMC and their exposure to fibroblast growth factor (FGF)-2, an injury-induced cytokine. The first is activation transcription factor (ATF) 4, which is upregulated by FGF-2 and mechanical injury in vascular SMC in vitro, and by balloon-injury in the artery wall. The induction of ATF4 by FGF-2 was shown to be mediated through the PI3K pathway, and preceded by phoshorylation of eIF2alpha, a known upstream effector of ATF4 activation. Knock-down of ATF4 expression inhibited balloon-injury induced neointimal hyperplasia, suggesting that ATF4 is a key player in the SMC response to injury. Furthermore, microarray analysis identified several genes whose transcription in response to FGF-2 may be regulated by ATF4. In particular, this work demonstrates that ATF4 is necessary for VEGF-A upregulation in SMC in response to FGF-2 and mechanical injury in vitro and in the artery wall following balloon-injury. The second is a translation factor, YrdC203. Using confocal fluorescence microscopy, YrdC203 was found to localize partially to the ER, and with RPL12, a component of the 60S ribosomal subunit. Immunoprecipitation studies demonstrate that YrdC203 also interacts with an initiation factor, eIF5B. Mutation of an initiation factors signature on the exterior of YrdC203 perturbed its interaction with RPL12 and eIF5B, and inhibited the increase in protein synthesis observed with overexpression of YrdC203. This implicates YrdC203 as a translation factor responsible for ensuring protein synthesis in vascular SMC in response to injury. The present work provides evidence for new molecular mechanisms, transcriptional and translational, regulating the response of vascular SMC to injury. This would provide leads for future therapeutic targets.
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GRIGOLATO, Jessica. « ACTIVATING TRANSCRIPTION FACTOR 4 (ATF4) IS UPREGULATED BY HUMAN HERPESVIRUS 8 INFECTION, INCREASES VIRUS REPLICATION AND PROMOTES VIRUS PROANGIOGENIC PROPERTIES ». Doctoral thesis, Università degli studi di Ferrara, 2013. http://hdl.handle.net/11392/2388888.
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Previous results from our group demonstrate that human herpesvirus 8 (HHV-8) triggers proangiogenic behavior in endothelial cells by promoting transcriptional activation and secretion of monocyte chemoattractant protein 1 (MCP-1), through activation of Nuclear Factor κB (NF-κB). However, inhibition of NF-κB still results in partial MCP-1 induction and consequent capillary-like structure formation, suggesting the involvement of another transcriptional pathway. Here we describe that HHV-8 infection upregulates cellular activating transcription factor 4 (ATF4), a member of the CREB family involved in the cell response to ER stress. Upregulation of ATF4 promotes HHV-8 infection, whereas its silencing decreases virus replication, transcription and antigen expression. Furthermore, ATF4 silencing decreases virus-induced MCP-1 production, as well as viral induction of tube-like structures in endothelial cells. We also show that ATF4 per se activates the MCP-1 promoter and induces proangiogenic properties in transfected endothelial cells. The elucidation of molecular mechanism involved in this process will result in a better understanding of the angiogenic process, its involvement in cancer and will help in designing novel therapies to reduce growth and vascularisation of Kaposi’s sarcoma.
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Gao, Juehua. « Biological functions and molecular mechanisms of the interleukin-4 signaling pathways in autoimmune exocrinopathy using the nod.b10.h2b mouse model of sjogren's syndrome ». [Gainesville, Fla.] : University of Florida, 2004. http://purl.fcla.edu/fcla/etd/UFE0006639.
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Thesis (Ph. D.)--University of Florida, 2004.Typescript. Title from title page of source document. Document formatted into pages; contains 152 pages. Includes Vita. Includes bibliographical references.
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GENTILI, Valentina. « HUMAN HERPESVIRUS 8 UPREGULATES ACTIVATING TRANSCRIPTION FACTOR 4 (ATF4) and REAL TIME PCR TO ASSESS TOTAL BACTERIAL LOAD IN CHRONIC WOUNDS ». Doctoral thesis, Università degli studi di Ferrara, 2010. http://hdl.handle.net/11392/2389363.
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I. Human Herpesvirus 8 Upregulates Activator Transcription Factor 4 BACKGROUND: Human herpesvirus 8 (HHV-8) is the primary etiologic agent of Kaposi’s sarcoma, a highly vascularised neoplasm of endothelial origin characterized by inflammation, neoangiogenesis, and by the presence of characteristic spindle cells. HHV-8 angiogenic activity is due to the activation of NF-kB and the subsequent induction of MCP-1 synthesis. MCP-1 is a chemokine produced by macrophages and endothelial cells in response to different stimuli, and is a direct mediator of angiogenesis. HHV-8-induced angiogenesis is MCP-1 dependent. However, HHV-8 activation of MCP-1 is not completely dependent on NF-kB induction and another cellular factor is involved. A potential candidate is the cellular activating transcription factor 4 (ATF4), a stress responsive gene. ATF4 is upregulated in several condition, including ER-stress, viral infection (e.g. CMV) and in tumours.
AIM: The aim of the research is to study the interaction between HHV-8 and ATF4, and verify whether ATF4 is involved in angiogenesis characteristic of HHV-8 infection.
METHODS: To demonstrate the effect of HHV-8 on ATF4 expression, Jurkat cells were infected with a cell-free viral inoculum, obtained in our laboratory by stimulating reactivation of latent HHV-8 in chronically infected cells (PEL-derived). HHV-8 reactivation from latency was assessed by transfection of BC-3 and BCBL-1 cells (PEL-derived) with pCG-ATF4 recombinant plasmid. DNA or RNA were analysed by PCR, rtPCR or quantitative real time PCR. Promoters activation was assessed by luciferase assays.
RESULTS: HHV-8 upregulates the expression of ATF4 gene, and overexpression of ATF4 is able to increase replication and transcription of HHV-8, but not to reactivate HHV-8 from latency. Preliminary results show that ATF4 activates the MCP-1 promoter in absence of the NF-kB binding sites.
CONCLUSION: HHV-8 induces ATF4 expression during the productive infection, probably to obtain advantage for its replication and neoplastic development. ATF4 might be implicated in HHV-8 induced tumorigenesis, being involved in several aspects of viral replication, and could represent a potential therapeutic target for HHV-8 induced transformation.
II. Real Time PCR To Assess Total Bacterial Load in Chronic Wounds
BACKGROUND: Wounds and wound healing are important issues in chronic patients (i.e. diabetes, pressure). Critical colonization and infection are strictly linked to a delay in wound healing. Analysis of pathogens present in chronic wound is an essential aspect for the wound care. Classic microbiologic methods have several limits to ensure the correct analysis of ulcer environment.
AIM: To analyse total bacterial load by a single quantitative real time PCR reaction in swabs and biopsies obtained from infected chronic wounds treated with an innovative hydrophobic dressing.
METHODS: Biopsies were collected at the beginning and after 4 weeks of treatment, and swabs were collected once a week for 4 weeks. Real time PCR was carried out on DNA extracted from biopsies and swabs amplifying a region of the 16S rRNA gene, highly conserved among bacteria. Moreover, DNA extracted from biopsies was also analysed for the detection of 2 anaerobic bacteria (B. Fragilis, F. necrophorum, frequently associated with delayed healing) by real time PCR amplifying specific unique regions of their genome. In parallel, classical culturing methods were performed on biopsies searching for Staphylococcus and Pseudomonas species.
RESULTS: We evaluated the correlation between the molecular data obtained by real time PCR and the clinical data, in particular considering the area of the wound. We observed a mean 253-fold decrease of the total bacterial load in 10/20 wounds those also showed an average 58% decrease of their area. This 10 wounds showed a positive correlation between clinical and molecular data. In 5/20 wound, we found a non significant 5,2-fold decrease of the total bacterial load, correlate with a 27% increase of the wound’s area. Thus, 75% of molecular results (15/20 wounds) were correlate to the clinical data. In contrast, classical culturing method did not correlate with the clinical data, confirming that classical methods have several limits and disadvantages. B. Fragilis was present in 10/20 wounds, and F. necrophorum in 2/20 wounds.
CONCLUSION: The molecular approach can be considered a reliable and rapid test to assess infection levels in chronic wounds, being more sensitive than the classic cultural techniques. The research of specific pathogens is not sufficient to assess the outcome of the wounds, whereas total bacterial load can give a prognostic value to the wound care.
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Oh, Seung Wook. « Regulation of Life Span by DAF-16/Forkhead Transcription Factor in Caenorhabditis elegans : A Dissertation ». eScholarship@UMMS, 2005. https://escholarship.umassmed.edu/gsbs_diss/22.
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The insulin/IGF-1 signaling pathway plays a pivotal role in life span regulation in diverse organisms. In Caenorhabditis elegans, a PI 3-kinase signaling cascade downstream of DAF-2, an ortholog of the mammalian insulin and insulin-like growth factor-1 (IGF-1) receptor, negatively regulates DAF-16/forkhead transcription factor. DAF-16 then regulates a wide variety of genes involved in longevity, stress response, metabolism and development. DAF-16 also receives signals from other pathways regulating life span and development. However, the precise mechanism by which DAF-16 directs multiple functions is poorly understood.
First, in Chapter II, we demonstrate that JNK is a novel positive regulator of DAF-16 in both life span regulation and stress resistance. Our genetic analysis suggests that the JNK pathway acts in parallel with the insulin-like signaling pathway to regulate life span and both pathways converge onto DAF-16. We also show that JNK-1 directly interacts with and phosphorylates DAF-16. Moreover, in response to heat stress, JNK-1 promotes the translocation of DAF-16 into thc nucleus. Our findings define a novel interaction between the stress response pathway (JNK) and the master regulator of life span (DAF-16), and provide a mechanism by which JNK regulates longevity and stress resistance.
Next, in Chapter III, we focus on the downstream targets of DAF-16. Here, we used a modified chromatin immunoprecipitation (ChIP) method to identify direct target promoters of DAF-16. We cloned 103 target sequences containing consensus DAF-16 binding sites and randomly selected 33 targets for further analysis. The expression of majority of these genes is regulated in a DAF-16-dependent manner. Moreover, inactivation of more than 50% of these genes significantly altered DAF-16-dependent functions such as longevity, fat storage and dauer diapause. Our results show that the ChIP-based cloning strategy leads to greater enrichment of DAF-16 target genes, compared to previous studies using DNA micro array or bioinformatics. We also demonstrate that DAF-16 is recruited to multiple promoters to coordinate regulation of its downstream target genes.
In summary, we identified the JNK signaling pathway as a novel input into DAF-16 to adapt animals to the environmental stresses. We also revealed a large number of novel outputs of DAF-16. Taken together, these studies provide insight into the complex regulation by DAF-16 to control diverse biological functions and eventually broaden our understanding of aging.
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Arabanian, Laleh Sadat [Verfasser], Gerhard [Akademischer Betreuer] Rödel, Alexander [Akademischer Betreuer] Kiani et Gerhard [Akademischer Betreuer] Ehninger. « Role of NFAT (Nuclear Factor of Activated T Cells) Transcription Factors in Hematopoiesis / Laleh Sadat Arabanian. Gutachter : Gerhard Rödel ; Alexander Kiani ; Gerhard Ehninger. Betreuer : Gerhard Rödel ». Dresden : Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2012. http://d-nb.info/1068148918/34.
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Junker, Astrid [Verfasser], Karin Akademischer Betreuer] Breunig, Gerd [Akademischer Betreuer] Jürgens et Ulrich [Akademischer Betreuer] [Wobus. « Characterization of the LEAFY COTYLEDON1 regulon : transcription factor controlled hormone cross-talk during somatic and zygotic embryogenesis / Astrid Junker. Betreuer : Karin Breunig ; Gerd Jürgens ; Ulrich Wobus ». Halle, Saale : Universitäts- und Landesbibliothek Sachsen-Anhalt, 2009. http://nbn-resolving.de/urn:nbn:de:gbv:3:4-2177.
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Andrade, João Nilton Barreto. « O SP1 (transcription factor Sp1) participa da regulação transcricional do Slc2a4 mediada pelo receptor de estrógeno ER-alfa em adipócitos 3T3-L1 ». Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/5/5160/tde-17082018-095039/.
Texte intégralRésumé :
O diabetes mellitus tipo 2 (DM2) é caracterizado pela presença de resistência à insulina, a qual pode ser modulada pelo estrógeno, tanto em fêmeas como em machos. Nesse processo, o transportador de glicose GLUT4 (gene Slc2a4, solute carrier family 2 member 4) desempenha papel importante, pois aumento da expressão do GLUT4 melhora o controle glicêmico. Estradiol (E2) regula a expressão do Slc2a4 por meio do balanço dos efeitos contrários de seus receptores (ERs): ER-alfa estimula e ER-beta inibe a expressão. Efeitos transcricionais dos ERs envolvem a participação de co-reguladores, destacadamente o SP1 (transcription factor Sp1), potente estimulador do Slc2a4. Entretanto, o papel do SP1 na regulação do Slc2a4 mediada pelos ERs é desconhecido; e este foi o objetivo do presente estudo. Investigou-se adipócitos maduros 3T3-L1, tratados por 24 horas com E2, agonista de ER-alfa (PPT) ou agonista de ER-beta (DPN). Avaliou-se: a expressão gênica (RT-qPCR) de Slc2a4 e Sp1; o conteúdo (Western blotting) total de GLUT4 e o nuclear de ER-alfa/beta e SP1; a atividade de ligação do SP1 no Slc2a4 (ensaio de mobilidade eletroforética); e a formação de complexos SP1/ER-alfa (imunoprecipitação). Os resultados confirmaram que E2 aumenta a expressão de Slc2a4/GLUT4 pela ação preponderante do ER-alfa. O agonista PPT aumentou: o conteúdo nuclear de SP1, a interação SP1/ER-alfa e a atividade de ligação do SP1 no Slc2a4. O agonista DPN indicou que a ação repressora do ER-beta não envolve o SP1. Conclui-se que o efeito estimulador do ER-alfa na expressão do Slc2a4 envolve mecanismo de transativação gênica via SP1. Essas observações colocam a cooperação ER-alfa/SP1 como um novo alvo para o desenvolvimento de medidas terapêuticas para resistência à insulina e diabetes mellitus tipo 2Type 2 diabetes mellitus (T2DM) is characterized by insulin resistance, which can be modulated by estrogen in both females and males. In this process, the glucose transporter GLUT4 (solute carrier family 2 member 4 gene - Slc2a4) plays an important role, since increasing GLUT4 expression improves glycemic control. Estradiol (E2) regulates the expression of Slc2a4, by a mechanism in which estrogen receptors (ERs) play opposite effects: ER-alpha stimulates, whereas ER-beta inhibits the expression. Transcriptional effects of ERs involve co-regulators, notably the transcription factor SP1, a powerful enhancer of Slc2a4. However, the role of SP1 in the ERs-mediated regulation of Slc2a4 is unknown; and that was the aim of the present study. Differentiated adipocytes 3T3-L1 were treated (24 hours) with E2, ER-alpha agonist (PPT) or ER-beta agonist (DPN). It was analyzed: gene expression (RT-qPCR) of Slc2a4 and Sp1; total content o GLUT4 and nuclear content of ER-alpha/beta and SP1 (Western blotting); binding activity of SP1 into Slc2a4 promoter (electrophoretic mobility shift assay); and content of nuclear SP1/ER-alpha complexes (immunoprecipitation). Results confirmed that E2 increases the expression of Slc2a4/GLUT4, by the dominant effect of ER-alpha. The ER-alpha agonist PPT increased the nuclear content of SP1, the interaction of SP1/ER-alpha, and the binding activity of SP1 into the Slc2a4. The agonist DPN evinced that ER-beta activity does not involve the SP1. In conclusion, the enhancer effect of ER-alpha upon Slc2a4 gene expression involves a transactivation mechanism via SP1. This observation point outs the cooperation of ER-alpha/SP1 as a new target for the development of approaches to treat insulin resistance and T2DM
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Humphrey, Peter Saah. « Signal transduction mechanisms for stem cell differentation into cardiomyocytes ». Thesis, University of Hertfordshire, 2009. http://hdl.handle.net/2299/3760.
Texte intégralRésumé :
Cardiovascular diseases are among the leading causes of death worldwide and particularly in the developed World. The search for new therapeutic approaches for improving the functions of the damaged heart is therefore a critical endeavour. Myocardial infarction, which can lead to heart failure, is associated with irreversible loss of functional cardiomyocytes. The loss of cardiomyocytes poses a major difficulty for treating the damaged heart since terminally differentiated cardiomyocytes have very limited regeneration potential. Currently, the only effective treatment for severe heart failure is heart transplantation but this option is limited by the acute shortage of donor hearts. The high incidence of heart diseases and the scarcity donor hearts underline the urgent need to find alternative therapeutic approaches for treating cardiovascular diseases. Pluripotent embryonic stem (ES) cells can differentiate into functional cardiomyocytes. Therefore the engraftment of ES cell-derived functional cardiomyocytes or cardiac progenitor cells into the damaged heart to regenerate healthy myocardial tissues may be used to treat damaged hearts. Stem cell-based therapy therefore holds a great potential as a very attractive alternative to heart transplant for treating heart failure and other cardiovascular diseases. A major obstacle to the realisation of stem cell-based therapy is the lack of donor cells and this in turn is due to the fact that, currently, the molecular mechanisms or the regulatory signal transduction mechanisms that are responsible for mediating ES cell differentiation into cardiomyocytes are not well understood. Overcoming this huge scientific challenge is absolutely necessary before the use of stem cell-derived cardiomyocytes to treat the damaged heart can become a reality. Therefore the aim of this thesis was to investigate the signal transduction pathways that are involved in the differentiation of stem cells into cardiomyocytes. The first objective was the establishment and use of cardiomyocyte differentiation models using H9c2 cells and P19 stem cells to accomplish the specific objectives of the thesis. The specific objectives of the thesis were, the investigation of the roles of (i) nitric oxide (ii) protein kinase C (PKC), (iii) p38 mitogen-activated protein kinase (p38 MAPK) (vi) phosphoinositide 3-kinase (PI3K) and (vi) nuclear factor-kappa B (NF-kB) signalling pathways in the differentiation of stem cells to cardiomyocytes and, more importantly, to identify where possible any points of convergence and potential cross-talk between pathways that may be critical for differentiation to occur. P19 cells were routinely cultured in alpha minimal essential medium (α-MEM) supplemented with 100 units/ml penicillin /100 μg/ml streptomycin and 10% foetal bovine serum (FBS). P19 cell differentiation was initiated by culturing the cells in microbiological plates in medium containing 0.8 % DMSO to form embryoid bodies (EB). This was followed by transfer of EBs to cell culture grade dishes after four days. H9c2 cells were cultured in Dulbecco’s Modified Eagle’s medium (DMEM) supplemented with 10% FBS. Differentiation was initiated by incubating the cells in medium containing 1% FBS. In both models, when drugs were employed, they were added to cells for one hour prior to initiating differentiation. Cell monolayers were monitored daily over a period of 12 or 14 days. H9c2 cells were monitored for morphological changes and P19 cells were monitored for beating cardiomyocytes. Lysates were generated in parallel for western blot analysis of changes in cardiac myosin heavy chain (MHC), ventricular myosin chain light chain 1(MLC-1v) or troponin I (cTnI) using specific monoclonal antibodies. H9c2 cells cultured in 1% serum underwent differentiation as shown by the timedependent formation of myotubes, accompanied by a parallel increase in expression of both MHC and MLC-1v. These changes were however not apparent until 4 to 6 days after growth arrest and increased with time, reaching a peak at day 12 to 14. P19 stem cells cultured in DMSO containing medium differentiated as shown by the timedependent appearance of beating cardiomyocytes and this was accompanied by the expression of cTnI. The differentiation of both P19 stem cells and H9c2 into cardiomyocytes was blocked by the PI3K inhibitor LY294002, PKC inhibitor BIM-I and the p38 MAPK inhibitor SB2035800. However when LY294002, BIM-I or SB2035800 were added after the initiation of DMSO-induced P19 stem cell differentiation, each inhibitor failed to block the cell differentiation into beating cardiomyocytes. The NF-kB activation inhibitor, CAPE, blocked H9c2 cell differentiation into cardiomyocytes. Fast nitric oxide releasing donors (SIN-1 and NOC-5) markedly delayed the onset of differentiation of H9c2 cells into cardiomyocytes while slow nitric oxide releasing donors (SNAP and NOC-18) were less effective in delaying the onset of differentiation or long term differentiation of H9c2 cells into cardiomyocytes. Akt (protein kinase B) is the key downstream target of PI3K. Our cross-talk data also showed that PKC inhibition and p38 MAPK inhibition respectively enhanced and reduced the activation of Akt, as determined by the phosphorylation of Akt at serine residue 473. In conclusion, PKC, PI3K, p38 MAPK and NF-kB are relevant for the differentiation of stem cells into cardiomyocytes. Our data also show that the PKC, PI3K and p38 MAPK signalling pathways are activated as very early events during the differentiation of stem cells into cardiomyocytes. Our data also suggest that PKC may negatively regulate Akt activation while p38 MAPK inhibition inhibits Akt activation. Our fast NO releasing donor data suggest that nitric oxide may negatively regulate H9c2 cell differentiation.
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Lobner, Elena Verfasser], Dirk [Akademischer Betreuer] [Haller, Dirk [Gutachter] Haller et Mathias [Gutachter] Heikenwälder. « Epithelial Cell-Specific Induction of Activating Transcription Factor 6 Signaling Promotes Intestinal Dysbiosis and Colonic Tumorigenesis / Elena Lobner ; Gutachter : Dirk Haller, Mathias Heikenwälder ; Betreuer : Dirk Haller ». München : Universitätsbibliothek der TU München, 2018. http://nbn-resolving.de/urn:nbn:de:bvb:91-diss-20180724-1402311-1-4.
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Goldstein, Evan Zachary. « TLR4-activated microglia have divergent effects on oligodendrocyte lineage cells ». The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1468967532.
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Lavaur, Jérémie. « Rôle de la phosphorylation du facteur de transcription Elk-1 dans son trafic intracellulaire et ses fonctions neuronales ». Paris 6, 2007. http://www.theses.fr/2007PA066108.
Texte intégralRésumé :
La phosphorylation du facteur de transcription Elk-1 par les MAPK/ERK est primordiale pour l’activité transcriptionnelle du complexe ternaire formé de Elk-1 et du dimère SRF (Serum Response Factor) sur le site SRE (Serum Response Element). En plus de sa localisation nucléaire Elk-1 est exprimé dans les dendrites des neurones. Nous avons montré in vivo et in vitro une relocalisation nucléaire de Elk-1 dépendante de l’activation de ERK. La phosphorylation des Sérines 383 et 389 de Elk-1 contrôle à la fois sa translocation nucléaire et l’expression de gènes porteurs de sites SRE. La mutation de ces Sérines en résidus non phosphorylables ou l’utilisation d’un peptide pénétrant (TAT-DEF-Elk-1), qui interfère spécifiquement avec le site d’ancrage, domaine DEF de Elk-1, bloque à la fois sa translocation nucléaire et la régulation de gènes SRE-dépendants. Enfin, le peptide TAT-DEF-Elk-1 diminue le niveau d’expression de SRF et d’Actine, et est corrélé à un blocage de la croissance des dendritesThe transcription factor Elk-1 is phosphorylated by MAPK/ERK, a critical post-translational event for the transcriptional activity of the ternary complex composed of Elk-1 and a dimer of Serum Response Factor Serum (SRF) at the Serum Response Element (SRE) regulatory site of transcription. In addition to its nuclear localization, Elk-1 is found in the dendrites and soma of neuronal cells. We showed both in vitro and in vivo a nuclear relocalization of Elk-1 in ERK-dependent activation. Phosphorylation on Ser383/389 triggers both Elk-1 nuclear translocation and SRE-dependent gene transcription. Mutating these sites into inactive residues or using a synthetic penetrating peptide (TAT-DEF-Elk-1), which specifically interferes with the DEF docking domain of Elk-1, blocks both Elk-1 nuclear translocation and SRE-dependent gene regulation. Finally, the TAT-DEF-Elk-1 decreases both SRF and actin expression levels, which is associated with an impairment of dendritic extension
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Carpenter, Oliver L. « Ultraviolet Light-Induced Regulation of Transcription and Translation, COX-2 Expression and Noncanonical NF-κB Activation ». Ohio University / OhioLINK, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1382624015.
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Reardon, Sara Noraen. « Characterizing the Role of Mammalian DEAF-1 in Reproduction, Neural Tube Closure, and Gene Expression in the Developing Embryo ». Available to subscribers only, 2008. http://proquest.umi.com/pqdweb?did=1674100501&sid=4&Fmt=2&clientId=1509&RQT=309&VName=PQD.
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Thesis (M.S.)--Southern Illinois University Carbondale, 2008."Department of Molecular, Cellular, and Systemic Physiology." Keywords: Deaf-1, Knockout mouse, Neural tube defects, Testis, Transcription factor. Includes bibliographical references (p. 61-75). Also available online.
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Osafo, Joy Kwakyewaa. « Regulation of the V1 mRNA variant of the human growth hormone receptor gene by Gfi-1, Gfi-1b, a GAGA element and the liver enriched transcription factor, HNF-4[alpha] ». Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=102688.
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GH acts through its specific receptor, GHR. In the human liver, more than twelve hGHR mRNAs are transcribed from unique TUTR exons, seven of which are found in two small clusters: module A (V2,V9,V3) and module B (V7,V1,V4,V8). While module A mRNAs are ubiquitously expressed, module B transcripts are restricted to normal postnatal liver, suggesting developmental- and liver-specific regulation of the hGHR gene. To begin characterising the elements regulating module B mRNA expression, I studied the promoter region of the V1 exon, the most abundantly expressing variant in liver. A 1.8 kb region upstream of the V1 transcriptional start site (TSS) was actively repressed in transient transfection assays (TTAs). However, T or 3' deletions relieved the suppression, suggesting the presence of multiple negative and positive regulatory elements. Two sites for growth factor independence-1 (Gfi-1) and Gfi-1b and a GAGA element were identified in the most 3' 300 by regulatory region. Gfi-1 was detected by western blot in human foetal and postnatal liver. Gfi-1 and Gfi-lb strongly repressed while the Drosophila GAGA factor (GAF) stimulated V1 transcription through their specific sites, as determined by TTAs and site-directed mutagenesis (SDM). Six putative sites for hepatic nuclear factor-4 (HNF-4) were also identified in the 1.8 kb region. HNF-4alpha is developmentally regulated in human liver: HNF-4alpha,2 and alpha8 proteins are expressed in foetal hepatocytes but only HNF-4alpha2 is detected postnatally. TTAs and SDM demonstrated that HNF-4alpha2 and HNF-4alpha8 have a similar dual effect on V1 transcription: activation via site #1 in the proximal promoter and repression through site #6, ~1.7 kb upstream of the TSS. Results from EMSA/EMSSA/ChIP analyses suggest these sites are bound by HNF-4alpha.Thus, V1 transcriptional activity is repressed by Gfi-1/Gfi-1b, stimulated through a GAGA element by GAF, and repressed or stimulated by HNF-4alpha. (2+8) depending on the site. Similar sites are present in homologous regions of the bovine, ovine and mouse GHR genes suggesting that their regulatory roles are conserved. However, none of these factors individually appear to be responsible for the postnatal hepatic-specific expression of V1 mRNA. To define the specific regulatory mechanisms, future studies should examine their interactions with additional liverenriched factors (e.g. C/EBPalpha).
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Oswal, Dhawal P. « Peroxisome proliferator-activated receptor alpha : Insight into the structure, function and energy homeostasis ». Wright State University / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=wright1401279322.
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Glosse, Philipp [Verfasser], Michael [Gutachter] Föller, Gabriele I. [Gutachter] Stangl et Lars-Oliver [Gutachter] Klotz. « Identification of novel regulators of fibroblast growth factor 23 (FGF23) production : the role of high-fat diet and AMP-activated protein kinase (AMPK) [kumulative Dissertation] / Philipp Glosse ; Gutachter : Michael Marc Uwe Föller, Gabriele I. Stangl, Lars-Oliver Klotz ». Halle (Saale) : Universitäts- und Landesbibliothek Sachsen-Anhalt, 2020. http://nbn-resolving.de/urn:nbn:de:gbv:3:4-1981185920-353921.
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Koivisto, E. (Elina). « Characterization of signalling pathways in cardiac hypertrophic response ». Doctoral thesis, Oulun yliopisto, 2011. http://urn.fi/urn:isbn:9789514294617.
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Abstract Intracellular signalling cascades regulate cardiomyocyte hypertrophic response. Initially hypertrophy of individual myocytes occurs as an adaptive response to increased demands for cardiac work, e.g. during hypertension or after myocardial infarction, but a prolonged hypertrophic response, accompanied by accelerated fibrosis and apoptosis, predisposes the heart to impaired performance and the syndrome of heart failure. The goal of this work was to elucidate some of the main signalling pathways in experimental models of the cardiac hypertrophic response. Mechanical stretching of cultured neonatal rat cardiomyocytes in vitro activates the B-type natriuretic peptide (BNP) gene, a well-established marker of the hypertrophic response, through intracellular signalling cascades mitogen-activated protein kinases (MAPKs) and protein kinase A (PKA) -pathway. Further, transcription factors transcriptional enhancer factor-1 (TEF-1) and activating transcription factor 3 (ATF3) were induced during stretch, and TEF-1 activation was shown to be regulated by extracellular signal-regulated kinase (ERK), while ATF3 activation was modulated by PKA. The BNP gene was also activated by the adenoviral overexpression of the p38 MAPK isoforms p38α and p38β in vitro. Importantly, p38α–induced activation was mediated through activator protein-1 (AP-1) while p38β mediated BNP transcription through GATA-4, which suggests distinct physiological roles for different p38 isoforms. This was further confirmed by quantitative PCR, which demonstrated pro-fibrotic role for the p38α isoform and a pro-hypertrophic role for the p38β isoform. Finally, adenoviral overexpression of ATF3 in vitro and in vivo resulted in activation of cardiac survival factors nuclear factor-κВ and Nkx-2.5, and attenuation of central pro-inflammatory and pro-fibrotic mediators. Together these data suggest a protective role for ATF3 in the heart. Overall this study provides new insights into the role of several signalling molecules involved in cardiac hypertrophic process and suggests potential therapeutic strategies for the diagnosis and treatment of heart failureTiivistelmä Sydämen kammioiden seinämät paksuuntuvat kuormituksen lisääntyessä mm. verenpainetaudissa tai sydäninfarktin jälkeen. Lisääntynyt kuormitus aiheuttaa sydänlihassolujen koon kasvun (hypertrofioitumisen) ohella sidekudoksen kertymistä (fibroosia) ja solukuolemaa. Nämä solutason muutokset lopulta vioittavat sydämen rakennetta niin, että sen toiminta pettää, ja sydän ajautuu vajaatoimintaan. Tätä taudin etenemistä säätelevät molekyylitasolla lukuisat solunsisäiset signaalinvälitysjärjestelmät, joita tässä väitöskirjatyössä tutkittiin eri koemalleissa. Sydämen täyttöpaineen nousun aiheuttama sydänlihassolujen mekaaninen venytys aktivoi natriureettisten peptidien (eteispeptidi, ANP ja B-tyypin natriureettinen peptidi, BNP) synteesiä ja vapautumista verenkiertoon. BNP geenin säätelyä mekaanisen venytyksen aikana tutkittiin rotan sydänlihassoluviljelmissä. Mitogeeni-aktivoituvat proteiinikinaasit (MAPK) sekä proteiinikinaasi A (PKA) säätelivät mekaanisen ärsykkeen aiheuttamaa BNP geenin ekspressiota. Venytys aktivoi myös transkriptiotekijöitä TEF-1 (transcriptional enhancer factor-1) ja ATF3 (activating transcription factor 3). TEF-1 sääteli venytyksen aiheuttamaa BNP:n aktivaatiota ERK:n (extracellular signal-regulated kinase) välityksellä BNP geenin säätelyalueella olevan sitoutumispaikkansa (M-CAT elementti) kautta. ATF3:n säätelyssä PKA:lla oli keskeinen merkitys. Tutkimus osoitti myös, että p38 MAPK:n alatyypeistä p38α lisäsi fibroosiin liittyvien geenien aktiivisuutta, kun taas p38β aiheutti solujen hypertrofioitumista lisäävien geenien ekspressiota. Molemmat alatyypit aktivoivat BNP geenin ekspressiota, mutta aktivaatio tapahtui eri transkriptiotekijöiden kautta. Tutkimuksessa havaittiin myös, että ATF3:n yliekspressio adenovirusvälitteisellä geeninsiirrolla lisäsi kahden sydäntä suojaavan transkriptiotekijän (nuclear factor-κВ ja Nkx-2.5) aktiivisuutta, sekä vähensi sydämen tulehdusvastetta ja fibroosia lisäävien tekijöiden (interleukiini-6 ja plasminogeeniaktivaattorin inhibiittori-1) ekspressiota. Väitöskirjatutkimus antaa uutta tietoa solunsisäisistä signaalinvälitys-järjestelmistä, jotka säätelevät sydänlihaksen kuormitusvastetta sydän- ja verenkiertoelimistön sairauksissa. Näiden solutason mekanismien tunteminen osaltaan edesauttaa jatkossa uusien menetelmien kehittämistä sydämen vajaatoiminnan ehkäisyyn ja hoitoon
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Thurisch, Boris. « Untersuchung der Funktion des Transkriptionsfaktors GATA-4 durch eine Mausmutante mit einem induzierbaren RNA-Interferenz System ». Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2007. http://dx.doi.org/10.18452/15703.
Texte intégralRésumé :
Hintergrund: Der Transkriptionsfaktor GATA-4 ist für die normale Entwicklung des Endoderms essentiell. Mausmutanten mit einer homozygoten Deletion des gata-4 Gens versterben zwischen den embryonalen Tagen 8.5 - 10.5 aufgrund einer Störung der ventralen Morphogenese und der Ausbildung des Herzschlauches. Zielsetzung und experimentelle Strategie: Um die Bedeutung von GATA-4 auch nach der embryonalen Entwicklung untersuchen zu können, wurden doppelt-transgene Mäuse generiert. Diese Mausmutanten exprimieren einen Tetrazyklin-Repressor und eine gegen GATA-4 gerichtete short hairpin RNA (shGATA-4). Die Expression der shGATA-4 steht dabei unter der Kontrolle eines H1-Promotors, welcher durch ein Tetrazyklin-Operator Element modifiziert wurde. Dadurch ist das System durch Doxyzyklin induzierbar. Ergebnisse: Die Integration der Transgene in dem Genom der Maus wurde durch Southern-Blot Analyse nachgewiesen. Die Expression der shGATA-4 wurde durch die Applikation von Doxyzyklin über das Trinkwasser (20 mg/ml) induziert. Langzeitstudien am Herzen haben dabei eine signifikante Suppression von GATA-4 nach 38 Tagen ergeben (80 %). Diese Reduktion konnte durch Western-Blot Analyse bestätigt werden. Obwohl die Expression verschiedener Zielgene von GATA-4 (ANF, BMP-4) ebenfalls herunterreguliert war, fiel bei den transgenen Mäusen kein kardialer Phänotyp auf. Jedoch wurde die GATA-4 Expression in den Hoden und Ovarien transgener Mäuse supprimiert, nachdem shGATA-4 durch die Applikation von Doxyzyklin induziert wurde. Weiterführende Untersuchungen an adulten Mäusen zeigten eine GATA-4 Reduktion von 20 % auch in nicht mit Doxyzyklin-induzierten Mausmutanten. Diese Reduktion könnte durch einen sog. leaky-Effekt des shGATA-4 Transgens hervorgerufen worden sein, wodurch die stark eingeschränkte Fertilität dieser Mauslinie erklärt werden könnte. Interessanterweise haben ca. 10 % der mit Doxyzyklin behandelten transgenen Weibchen Ovarial-Teratome ausgebildet. Histologisch wiesen diese Teratome überwiegend (neuro-) ektodermale, vereinzelt mesodermale und nahezu keine endodermalen Strukturen auf. Schlussfolgerung: In diesem Modell hat die Suppression von GATA-4 keinen Einfluss auf die Funktion des Herzens der adulten Maus. Jedoch scheint GATA-4 für die Fertilität der Maus von großer Bedeutung zu sein. Weiterhin scheint die Suppression von GATA-4 mit der Ausbildung von Ovarial-Teratomen assoziiert zu sein.Background: The transcription factor GATA-4 is crucial for the normal endodermal development. In mice, homozygous deficiency of GATA-4 causes defects in ventral morphogenesis and heart tube formation, resulting in embryonic death between day e8.5 and e10.5. Aim and experimental strategy: To analyze the implication of GATA-4 beyond embryonic development a double transgenic mouse expressing the tetracycline repressor (TetR) and an inducible small interfering RNA directed against GATA-4 was generated. This expression construct contains a H1 promoter modified with a tetracycline operator upstream of the coding region for the GATA-4 short hairpin RNA (shGATA-4). Results: The integration of the transgenes in FvB mice (H1:G4/TetR) was confirmed by Southern blot. To induce the expression of the shGATA-4 construct, transgenic mice were treated with doxycycline (20 mg/ml drinking water). In longitudinal analysis, most efficient GATA-4 suppression was detected after 38 days. Quantitative PCR revealed a GATA-4 reduction of about 80 % in the heart, if normalized against the wildtype. Reduction of GATA-4 was confirmed by Western Blot. Although GATA-4 target genes (ANP, BMP-4) were down regulated, the animals showed no clinical phenotype. In opposite to wildtype mice, GATA-4 expression was undetectable in the ovaries and testis of transgenic mice with induced shGATA-4. Additional analysis in adult transgenic mice, which were not treated with doxycycline, also showed a reduction of GATA-4 expression of about 20 %, probably caused by a leaky-effect of the transgene. This may explain the significantly reduced fertility of the colony. Importantly, 10 % of transgenic females treated with doxycycline developed ovarian teratomas. Histological examination of teratomas showed predominantly (neuro-) ectodermal and to a lower degree mesodermal, but almost no endodermal compounds. Conclusions: GATA-4 reduction in the adult murine heart is – at least to a certain degree – clinically redundant. GATA-4 seems to be required for normal fertility. In our model GATA-4 deficiency seems to be associated with an increased risk for developing ovarian teratoma.
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Codlin, Sandra Maria Susan. « A novel palindromic element common to the granulocyte macrophage colony stimulating factor gene and the Th2 expressed cytokine genes : interleukin (IL) 4, IL 5 and IL 13 acts as a potent enhancer of transcription ». Thesis, King's College London (University of London), 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.249674.
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Amin, Shahreen. « Regulation of the tyrosine phosphatase SHP-1 expression by C-jun-N-terminal kinase and RFX-1 and AP-4 transcription factors in insulin-like growth factor-1 (IGF-1) stimulated breast adenocarcinoma MCF-7 cells ». Thesis, University of Ottawa (Canada), 2005. http://hdl.handle.net/10393/26837.
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This thesis is devoted to reveal the negative regulators in IGF-1 (Insulin like growth factor 1) stimulated growth of a human breast adenocarcinoma cell line. It is a well established fact that increased circulating levels of IGF-1 correlate with increased risk of breast cancer. IGF-1 activation of its receptor, IGF-1R, is implicated in the progression of breast cancer, where IGF-1 stimulation leads to proliferative and anti-apoptotic responses by stimulating MAPK Erk and PI3K, respectively. In this study, IGF-1 stimulated MCF-7 cells proliferated more in the absence of MAPK JNK, implicating the involvement of MAPK JNK in the negative regulation of IGF-1 stimulated cell growth.
In this research study, we show for the first time that IGF-1 stimulation of breast cancer cells induces SHP-1-expression by activating JNK, which in turn, activates RFX-1 and AP-4 transcription factors to allow them to bind to the high-expression region of the SHP-1 P-1 promoter in breast adenocarcinoma MCF-7 cells. (Abstract shortened by UMI.)
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Lamb, Jennifer A. « Role of the JNK Signal Transduction Pathway in Cell Survival : a Dissertation ». eScholarship@UMMS, 2004. http://escholarship.umassmed.edu/gsbs_diss/232.
Texte intégralRésumé :
The c-Jun NH2-terminal kinases (JNK) are evolutionarily conserved serine/threonine protein kinases that are activated by proinflammatory cytokines, environmental stress, and genotoxic agents. These kinases play key regulatory roles within a cell by coordinating signals from the cell surface to nuclear transcription factors. JNK phosphorylates the amino terminal domain of all three Jun transcription factors (JunB, c-Jun and JunD) all members of the AP-1 family. The activated transcription factors modulate gene expression to generate appropriate biological responses, including cell migration, proliferation, differentiation and cell death.
The role of the JNK signaling pathway in cell death/apoptosis is controversial, both pro-apoptotic and pro-survival roles have been attributed to JNK. The mechanism that enables the JNK signaling pathway to mediate both apoptosis and survival is unclear. The aim of this study is to examine the role of TNF-stimulated JNK activation on cell survival.
The proinflammatory cytokine TNF, is known to activate JNK and induce apoptosis. To test whether the JNK signaling pathway contributes to TNF-induced apoptosis, the response of wild type and Jnk1-/- Jnk2-/- (JNK deficient fibroblasts) fibroblasts to TNF was examined. JNK deficient fibroblasts are more sensitive to TNF-induced apoptosis than wild-type fibroblasts. The TNF-sensitivity cannot be attributed to altered expression of TNF receptors or defects in the NF-кB or AKT pathways, known anti-apoptotic signal transduction pathways. (In fact, TNF stimulated NF-кB activation provides a major mechanism to account for survival in both wild-type and JNK deficient cells.) However this increased TNF-sensitivity can be attributed to JNK deficiency. Apoptosis is suppressed in JNK deficient cells when transduced with JNK1 retrovirus. These data implicate the JNK signaling pathway in cell survival.
The AP-1 family of transcription factors is a target of the JNK signal transduction pathway. In addition JNK is required for the normal expression of the AP-1 family member, JunD. Previous studies have indicated that JunD can mediate survival. Interestingly, JNK deficient and JunD null cells display similar phenotypes: premature senescence and increased sensitivity to TNF induced apoptosis. In fact, the TNF-sensitivity is also suppressed in JNK deficient fibroblasts transduced with JunD retrovirus. Although JunD can replace the survival signaling role of JNK, phosphorylation of JunD is essential to inhibit TNF induced apoptosis. JNK deficient cells transduced with phosphomutant JunD retrovirus maintain TNF-sensitivity.
Activated transcription factors modulate gene expression. It is most likely that JunD functions by regulating the expression of key molecules that act to inhibit TNF-stimulated apoptosis. Microarray analysis comparing wild-type with JNK deficient fibroblasts revealed that the expression of the survival gene, cIAP-2, was induced by TNF in only wild-type fibroblasts. Furthermore, protein expression of cIAP-2 was induced by TNF in only wild-type fibroblasts. Analysis of the cIAP-2 promoter revealed two critical NF-кB binding sites and one AP-1 binding site. Luciferase reporter assays indicated key roles for both NF-кB and the AP-1 component, JunD in TNF-induced cIAP-2 gene expression. These experiments establish that the JNK/JunD pathway collaborates with NF-кB pathway to increase the expression of the anti-apoptotic protein cIAP-2 in TNF treated cells. Without this collaboration, the JNK pathway mediates apoptosis.
The integration of JNK signaling with other signaling pathways represents a mechanism to account for the dual ability of the JNK pathway to mediate either survival or apoptosis. The dynamic coordination of signals within and between pathways is critical. The future challenge will be to fit the details of individual signaling pathways into the context of signaling networks.
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Hua, Guoqiang. « Régulation transcriptionnelle du gène ERBB2 dans le cancer du sein ». Aix-Marseille 2, 2009. http://theses.univ-amu.fr.lama.univ-amu.fr/2009AIX22058.pdf.
Texte intégralRésumé :
Le gène ERBB2 code un récepteur tyrosine kinase, dont l’amplification et la surexpression est un mauvais pronostic observé dans 20-30% de cancer du sein et est directement associé avec la résistance de la chimiothérapie. Deux anticorps monoclonaux humain dirigés contre ERBB2 trastuzumab (Herceptin®) et pertuzumab (Omnitarg®) sont actuellement utilisés dans les traitements cliniques pour les ERBB2+ patients. Ils bloquent les activités du récepteur, cependant, la résistance et certains problèmes secondaires sont fréquents. Le facteur de transcription GATA4 a été identifié dans une liste de 29 gènes co-exprimés avec ERBB2 dans des lignées tumorales et des cellules de cancer du sein. Nous avons mis en évidence dans cette thèse une boucle d'autorégulation négative entre le gène ERBB2 et GATA4 dans des lignées cellulaires épithéliales de cancer du sein. La translocation nucléaire d’ERBB2 a été décrite comme une conséquence de l'endocytose de ce récepteur. Notre étude a montré que l’isoforme ERBB2b dépourvu d’un peptide signal est une forme nucléaire du récepteur ERBB2. L’inactivation par siRNA des deux isoformes a mise en évidence un rôle activateur de la transcription d’ERBB2 pour l'isoforme a, qui correspond à la forme membranaire du récepteur, et un rôle répresseur pour l'isoforme ERBB2b. Des essais luciférase avec le promoteur d'ERBB2 ont confirma le rôle activateur de l'isoforma a. De plus, la fixation d'ERBB2 nucléaire in vivo dans la région promotrice d’ERBB2 contenant un site potentiel pour le recrutement d'ERBB2 a été démontrée par Immunoprécipitation de la chromatine (ChIP). L'ensemble de ces observations ouvre de nouvelles perspectives pour notre compréhension de l'amplification préférentielle d'ERBB2 et la quasi absence de mutation dans les cancers du sein ainsi que du rôle respectifs des deux isoformes majeures du gène ERBB2 et des résistances au traitement par anticorps monoclonaux spécifiques d'ERBB2 des cancers du seinThe ERBB2 gene encodes a tyrosine kinase receptor. Amplification and overexpression of ERBB2 is a poor prognosis indicator observed in 20-30% of breast cancers and is directly associated with resistance to chemotherapy. Two humanized monoclonal antibodies trastuzumab (Herceptin®) and pertuzumab (Omnitarg®) directed against ERBB2 are currently used in clinical treatment for ERBB2+ patients by blocking the activity of the receptor; however resistance and some secondary problems are frequent. Transcription factor GATA4 was originally identified as one of the 29 genes co-expressing with ERBB2 in breast cancer cells and tumors. We reported in this thesis a negative feedback regulatory loop associating both ERBB2 and GATA4 in breast cancer cells. ERBB2 nuclear translocation was reported as a consequence of endocytosis. Our studies showed that the ERBB2b isoform which lacks the peptide signal is a nuclear form of ERBB2 receptor. SiRNA assays targeted either ERBB2 isoform a or b have evidenced that isoform, corresponding to the membrane receptor, plays an activator role whereas isoform b acted as a repressor on ERBB2 transcription. Luciferase assays under the control of ERBB2 promoter have confirmed the activator role of ERBB2a. In addition, in vivo binding of nuclear ERBB2 to the promoter region of ERBB2 was determined by chromatin immunoprecipitation (ChIP) via a putative ERBB2 DNA binding motif. Further experiments are required to determine whether the recruitment of ERBB2 at its own promoter is direct or requires interaction with an unkown sequence-specific transcription factors. Altogether these original observations open new perspectives for our understanding why the ERBB2 gene is amplified and not mutated in breast cancers as well as for the respective role of the two major ERBB2 isoforms and for the resistance to ERBB2-specific monoclonal antibodies treatment of breast cancer
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Bedi, Shimpi. « Identification of Novel Ligands and Structural Requirements for Heterodimerization of the Liver X Receptor Alpha ». Wright State University / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=wright1495630797912988.
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Lusito, E. « A Network-based Approach to Breast Cancer Systems Medicine ». Doctoral thesis, Università degli Studi di Milano, 2015. http://hdl.handle.net/2434/265572.
Texte intégralRésumé :
Breast cancer is the most commonly diagnosed cancer and the second leading cause of cancer death in women. Although recent improvements in the prevention, early detection, and treatment of breast cancer have led to a significant decrease in the mortality rate, the identification of an optimal therapeutic strategy for each patient remains a difficult task because of the heterogeneous nature of the disease. Clinical heterogeneity of breast cancer is in part explained by the vast genetic and molecular heterogeneity of this disease, which is now emerging from large-scale screening studies using “-omics” technologies (e.g. microarray gene expression profiling, next-generation sequencing). This genetic and molecular heterogeneity likely contributes significantly to therapy response and clinical outcome. The recent advances in our understanding of the molecular nature of breast cancer due, in particular, to the explosion of high-throughput technologies, is driving a shift away from the “one-dose-fits-all” paradigm in healthcare, to the new “Personalized Cancer Care” paradigm. The aim of “Personalized Cancer Care” is to select the optimal course of clinical intervention for individual patients, maximizing the likelihood of effective treatment and reducing the probability of adverse drug reactions, according to the molecular features of the patient. In light to this medical scenario, the aim of this project is to identify novel molecular mechanisms that are altered in breast cancer through the development of a computational pipeline, in order to propose putative biomarkers and druggable target genes for the personalized management of patients. Through the application of a Systems Biology approach to reverse engineer Gene Regulatory Networks (GRNs) from gene expression data, we built GRNs around “hub” genes transcriptionally correlating with clinical-pathological features associated with breast tumor expression profiles. The relevance of the GRNs as putative cancer-related mechanisms was reinforced by the occurrence of mutational events related to breast cancer in the “hub” genes, as well as in the neighbor genes. Moreover, for some networks, we observed mutually exclusive mutational patterns in the neighbors genes, thus supporting their predicted role as oncogenic mechanisms. Strikingly, a substantial fraction of GRNs were overexpressed in Triple Negative Breast Cancer patients who acquired resistance to therapy, suggesting the involvement of these networks in mechanisms of chemoresistance. In conclusion, our approach allowed us to identify cancer molecular mechanisms frequently altered in breast cancer and in chemorefractory tumors, which may suggest novel cancer biomarkers and potential drug targets for the development of more effective therapeutic strategies in metastatic breast cancer patients.
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Catherinet, Claire. « Etude des effecteurs de la voie Ca2+/Calmoduline dans les leucémies aiguës lymphoblastiques T ». Thesis, Sorbonne Paris Cité, 2017. http://www.theses.fr/2017USPCC293/document.
Texte intégralRésumé :
Les leucémies aigües lymphobastiques (LAL) représentent un tiers des leucémies et constituent le cancer pédiatrique le plus fréquent chez l’enfant. Les LAL de type T (LAL-T)sont caractérisées par l’expansion anormale de progéniteurs de lymphocytes T. Aujourd’hui,la réponse curative aux traitements est proche de 80% chez l’enfant et 50% chez l’adulte. La rechute reste donc fréquente et souvent de mauvais pronostic. Pour ces raisons,l’identification de nouvelles voies de signalisation en vue de développer de nouvelles stratégies thérapeutiques est cruciale afin d’améliorer le traitement des LAL-T.Les résultats précédents du laboratoire ont révélé l’activation soutenue de la voie calcineurine (Cn)/NFAT dans des échantillons humains de lymphomes et de LAL, ainsi que dans des modèles murins de ces pathologies. Le laboratoire a ensuite montré que Cn est intrinsèquement requise pour la capacité des cellules leucémiques de LAL-T à propager la maladie (activité LIC « Leukemia Initiating Cells ») dans un modèle murin de LAL-T induit parun allèle activé de NOTCH1 (ICN1). Puisque l’inhibition pharmacologique de Cn induit de nombreux effets secondaires, la recherche de cibles thérapeutiques en aval de Cn constitue un axe de recherche important. J’ai participé à une étude du laboratoire montrant que l’expression à la surface cellulaire de CXCR4 est régulée par Cn et requise pour la migration des cellules de LAL-T, mais non suffisante pour rétablir le potentiel de ré-initiation suggérant que d’autres effecteurs doivent être impliqués dans cette activité.Les facteurs de transcription NFAT (NFAT1, NFAT2 et NFAT4) sont des effecteurs importants de Cn en réponse à la signalisation calcique lors du développement des thymocytes, mais également dans les lymphocytes T. L’essentiel de ce travail de thèse a utilisé des LAL-T induites par ICN1 dans lesquelles l’inactivation génique des trois facteurs NFAT par recombinaison homologue. Nous avons ainsi montré que (i) les facteurs NFAT sont requis en aval de Cn pour le potentiel LIC des LAL-T-ICN1 in vivo, (ii) leur inactivation altère la survie, la prolifération et la migration des cellules de LAL-T in vitro, (iii) NFAT1,NFAT2 et NFAT4 ont une fonction largement redondante dans les LAL-T. Nous avons également par une approche transcriptomique identifié deux gènes dont l’expression estsous contrôle des facteurs NFAT et impliqués dans la régulation de la survie et de la prolifération des LAL-T in vitro : CDKN1A et MAFB.Tout comme la voie Cn/NFAT, les CaMKs sont des protéines kinases activées en aval de la signalisation calcique dans les lymphocytes T. Nous avons montré par une approche pharmacologique que l’inhibition des CaMKs dans les LAL-T-ICN1 in vitro altère la survie etla prolifération des cellules leucémiques. L’inhibition spécifique par une approche d’ARN interférence de deux isoenzymes CaMKIIγ et CaMKIIδ suggèrent que ces protéines jouent dans le maintien des cellules leucémiques in vitroT cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of T cell progenitors. Despite initial response to chemotherapy, relapses remain frequent in children and adults. Previous results identify sustained activation of Calcineurin (Cn)/NFAT signaling pathway in human T-ALL and murine T-ALL models. Importantly, they also demonstrated Cn is essential for T-ALL Leukemia Initiating Cells (LIC) activity in a murine model of T-ALL induced by an activated allele of NOTCH1 (ICN1). Since pharmacologic inhibition of Cn induces side effects, we aim to identify downstream effectors involved in T-ALL. NFAT (Nuclear Factor of Activated T cells) factors play crucial roles downstream Cn during development and activation of T cells. To address their role in T-ALL, we generated mouse ICN1-induced T-ALL in which NFAT genes can be inactivated either single or in combination following Cre-mediated gene deletion. We demonstrated that (i) NFAT factors are required downstream Cn for LIC activity in T-ALL in vivo (ii) ex vivo NFAT factors deletion alters survival, proliferation and migration of T-ALL (iii) NFAT1, 2 and 4 have a largely redundant function in T-ALL. Moreover, the NFAT-dependant transcriptome allowed to identify important targets (CDKN1A, MAFB) involved in T-ALL survival and proliferation in vitro. Calmodulin-dependant kinases (CaMK) are kinases activated by calcium signaling in T cells. We showed that pharmacologic inhibition of CaMKs in ICN1-induced T-ALL alters survival and proliferation of T-ALL in vitro. Beside, specific inhibition by RNA interference of CaMKIIg and CaMKIId suggests a putative role of these kinases in T-ALL maintenance
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Tesz, Gregory J. « Role of MAP4K4 Signaling in Adipocyte and Macrophage Derived Inflammation : A Dissertation ». eScholarship@UMMS, 2008. https://escholarship.umassmed.edu/gsbs_diss/380.
Texte intégralRésumé :
Human obesity is increasing globally at an impressive rate. The rise in obesity has led to an increase in diseases associated with obesity, such as type 2 diabetes. A major prerequisite for this disease is the development of insulin resistance in the muscle and adipose tissues. Interestingly, experiments in rodent models suggest that adipocytes and macrophages can profoundly influence the development of insulin resistance. Accordingly, the number of adipose tissue macrophages increases substantially during the development of obesity. Numerous research models have demonstrated that macrophages promote insulin resistance by secreting cytokines, like TNFα, which impair whole body insulin sensitivity and adipose tissue function. Additionally, enhancements of murine adipose function, particularly glucose disposal, prevent the development of insulin resistance in mice on a high fat diet. Thus, mechanisms which enhance adipose function or attenuate macrophage inflammation are of interest.
Our lab previously identified mitogen activated protein kinase kinase kinase kinase 4 (MAP4K4) as a potent negative regulator of adipocyte function. In these studies, TNFα treatment increased the expression of adipocyte MAP4K4. Furthermore, the use of small interfering RNAs (siRNA) to block the increase in MAP4K4 expression protected adipocytes from some of the adverse effects of TNFα. Because MAP4K4 is a potent negative regulator of adipocyte function, an understanding of the mechanisms by which TNFα regulates MAP4K4 expression is of interest. Thus, for the first part of this thesis, I characterized the signaling pathways utilized by TNFα to regulate MAP4K4 expression in cultured adipocytes. Here I show that TNFα increases MAP4K4 expression through a pathway requiring the transcription factors activating transcription factor 2 (ATF2) and the JUN oncogene (cJUN). Through TNFα receptor 1 (TNFR1), but not TNFR2, TNFα increases MAP4K4 expression. This increase is highly specific to TNFα, as the inflammatory agents IL-1β, IL-6 and LPS did not affect MAP4K4 expression. In agreement, the activation of cJUN and ATF2 by TNFα is sustained over a longer period of time than by IL-1β in adipocytes. Finally, MAP4K4 is unique as the expression of other MAP kinases tested fails to change substantially with TNFα treatment.
For the second part of this thesis, I assessed the role of MAP4K4 in macrophage inflammation in vitro and in vivo. To accomplish this task, pure β1,3-D-glucan shells were used to encapsulate siRNA. Glucan shells were utilized because they are effectively taken up by macrophages which express the dectin-1 receptor and they survive oral delivery. I demonstrate that these β1,3-D-glucan encapsulated RNAi particles (GeRPs) are efficiently phagocytosed and capable of mediating the silencing of multiple macrophage genes in vitro and in vivo. Importantly, oral treatment of mice with GeRPs fails to increase plasma IFNγ and TNFα or alter serum AST and ALT levels. Orally administered GeRPs are found in macrophages isolated from the spleen, liver, lung and peritoneal cavity and mediate macrophage gene silencing in these tissues.
Utilizing this technology, I reveal that MAP4K4 augments the expression of TNFα in macrophages following LPS treatment. Oral delivery of MAP4K4 siRNA in GeRPs silences MAP4K4 expression by 70% and reduces basal TNFα and IL-1β expression significantly. The depletion of MAP4K4 in macrophages protects 40% of mice from death in the LPS/D- galactosamine (D-GalN) model of septicemia, compared to less than 10% in the control groups. This protection associates with significant decreases in serum TNFα concentrations following LPS/D-GalN challenge. Consistent with reduced macrophage inflammation, hepatocytes from mice treated orally with GeRPs targeting MAP4K4 present less apoptosis following LPS/D-GalN treatment. Thus, MAP4K4 is an important regulator of macrophage TNFα production in response to LPS.
The results presented here add to the knowledge of MAP4K4 action in adipocyte and macrophage inflammation substantially. Prior to these studies, the mechanism by which TNFα controlled MAP4K4 expression in adipocytes remained unknown. Considering that MAP4K4 is a negative regulator of adipocyte function, identifying the mechanisms that control MAP4K4 expression was of interest. Furthermore, the role of macrophage MAP4K4 in LPS stimulated TNFα production was also unknown. To address this question in vivo, new technology specifically targeting macrophages was needed. Thus, we developed a technology for non toxic and highly specific macrophage gene silencing in vivo. Considering that macrophages mediate numerous diseases, the application of GeRPs to these disease models is an exciting new possibility.
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Thomas, Amandine. « Hypoxie intermittente et homéostasie glucidique : étude des mécanismes d'action cellulaire A hybrid model to study pathological mutations of the human ADP/ATP carriers Visceral white fat remodeling contributes to intermittent hypoxia-induced atherogenesis The insulin sensitizing effect of topiramate involves KATP channel activation in the central nervous system The Impact of Sleep Disorders on Glucose Metabolism : Endocrine and Molecular Mechanisms Endoplasmic reticulum stress as a novel inducer of hypoxia inducible factor-1 activity : its role in the susceptibility to myocardial ischemia-reperfusion induced by chronic intermittent hypoxia Chronic intermittent hypoxia improves whole-body glucose tolerance by activating skeletal muscle AMP-activated protein kinase in mice Prolyl-4-hydroxylase 1 (PHD1) deficiency impairs whole-body glucose tolerance and insulin sensitivity in mice but does not worsen high-fat diet-induced metabolic dysfunctions Specific transcriptomic signature in response to intermittent hypoxia exposure in liver and fat tissue ». Thesis, Université Grenoble Alpes (ComUE), 2015. http://www.theses.fr/2015GREAV044.
Texte intégralRésumé :
L'hypoxie intermittente (HI), induite par les apnées du sommeil, conduit à des altérations de la sensibilité à l'insuline et de l'homéostasie glucidique mais les mécanismes impliqués restent mal connus. L'objectif de ce travail était d'étudier les effets et les mécanismes sous jacents d'une exposition chronique à l'HI sur l'homéostasie glucidique. L'HI induit une résistance à l'insuline à la fois systémique et tissulaire, ainsi qu'une amélioration de la tolérance au glucose associée à une activation de l'AMPK musculaire. L'HI cause également des altérations du foie et du tissu adipeux associées à un changement du pattern d'expression des gènes dans ces tissus et à un risque accru de développement de pathologies vasculaires comme l'athérosclérose. Enfin, la délétion de PHD1, une des protéines régulatrices de HIF-1, entraîne une résistance à l'insuline associée une stéatose hépatique, faisant de HIF-1 une cible potentielle impliquée dans les altérations metaboliques induites par l'HIIntermittent hypoxia (IH), induced by sleep apnea, leads to alterations in insulin sensitivity and glucose homeostasis but the mechanisms involved remains poorly understood. The objective of this work was to study the effects and the underlying mechanisms of chronic exposure to IH on glucose homeostasis. IH induces both systemic and tissue-specific insulin resistance , as well as improved glucose tolerance associated with an activation of muscle AMPK. IH also causes a change in the pattern of gene expression in liver and adipose tissue and an increased risk of vascular pathologies such as atherosclerosis development. Finally, the deletion of PHD1, a regulatory protein of HIF-1, leads to insulin resistance associated with hepatic steatosis, making HIF-1 a possible target involved in the metabolic changes induced by IH
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Hsieh, Cheng-Chih, et 謝丞智. « The Functional Study of Activator Transcription Factor 4 (ATF4) Leads to Adipose Differentiation in Zebrafish ». Thesis, 2016. http://ndltd.ncl.edu.tw/handle/51769232884431671089.
Texte intégralRésumé :
碩士國立臺灣海洋大學
生命科學暨生物科技學系
104
ATF4 (activator transcription factor4) is a activator transcription factor. It is a member of cyclic adenosine monophosphate responsive element-binding protein family. When endoplasmic reticulum stress exerts an influence to cells, the expression of ATF4 will be upregulated in the cytoplasm. Then the transcription of ATF4 downstream genes such as CHOP (CCAAT-enhancer-binding protein homologous protein), Atg12 (Autophagy-related protein 12), E-selection are promoted. The unfolded protein response is carried out after their transcription are promoted and supports cells to tide over endoplasmic reticulum stress. Previous several studies demonstrated that overexpression of ATF4 in the embryo fibroblast of the mouse increased adipocyte differentiation. In addition to, silenced ATF4 by siRNA blocked conversion of preadipocytes to adipocytes and a lot of expression of genes associated with adipocyte differentiation such as PPARγ (peroxisome proliferator-activated receptor gamma), C/ EBPβ (CCAT/enhancer-binding protein beta), C/EBPα (CCAT/ enhancer- binding protein alpha), SCD1 (stearoyl CoA desaturase1) were significantly downregulated. Previous studied indicated that ATF4 directly targeted many genes related to adipogenesis and lipid metabolism such as C/EBPβ, PPARγ, FGF19 (Fibroblast growth factor 19), RUNX2a (Runt-related transcription factor 2). The transparent embryos of the zebrafish provide new selection to research adipose biology. We could observe embryos whether they are overexpressed ATF4 with fluorescence microscopy. Therefore, we expect to set up a ATF4 overexpression transgene zebrafish model in our study. We transfected ATF4 gene into constructed transgene vector included tet-off system and microinject this construct into embryos. They would express whole body transgene by β-actin promoter. The whole body β-actin expresses exogenous ATF4 and carry red fluorescent protein. We could turn off expression of ATF4 in this model fish by adding doxycycline. Continuously, we used oil red O staining to assay adipose differentiation and lipid accumulation and detect the expression of ATF4 targeted genes in embryos in7dpf(day-post fertilization) and 21dpf. It provided a platform to study adipogensis in zebrafish early stage.
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Wottawa, Marieke Claudia. « Oxygen-dependent regulation of the activating transcription factor-4 (ATF-4) ». Doctoral thesis, 2009. http://hdl.handle.net/11858/00-1735-0000-0006-AD8D-0.
Texte intégral
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Hsieh, Yi-Hsuan, et 謝易軒. « Analysis of nuclear localization signal of human transcription factor AP-4 ». Thesis, 2004. http://ndltd.ncl.edu.tw/handle/47042057670357414680.
Texte intégralRésumé :
碩士國立嘉義大學
生物科技研究所
92
AP-4 is a transcription factor that activates both cellular and viral genes by binding to the DNA sequence of CAGCTG. According to its amino acid sequence, it reveals that AP-4 is a helix-loop-helix (HLH) protein. The AP-4 HLH motif and its adjacent basic region are required for specific DNA binding. This motif is found in a variety of enhancer binding proteins and putative transcription factors involved in the control of cellular proliferation and differentiation. Thus, what is the mechanism by which the AP-4 is transported into the nucleus? We applied fluorescence protein to localize AP-4 and found that its destination is nucleus. By sequential deletions of AP-4, region containing the NLS sequence has been mapped. NLS is a potential tool for delivery genes and nucleus targeting proteins; therefore, in the future we expect that the NLS of AP-4 will be useful in this field. In the future, we will try to make a variety of mutations on DNA sequences of NLS fragment of AP-4, to realize the significances of each amino acid. In addition we will make advanced researches to study what kinds of importins will bring AP-4 into or out of nucleus.