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Regulation of the Anti-apoptotic Protein Ku70 and the Implications for Bax-Mediated ApoptosisGama, Vivian 23 September 2008 (has links)
No description available.
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Functional analysis on the interactions of the human immunodeficiency virus type 1 integrase with its cofactors that regulate viral replicationZheng, Yingfeng 03 1900 (has links)
Like all viruses, the replication of HIV-1 relies heavily on host proteins due to its limited genome products. HIV-1 integrase (IN) catalyzes the integration of viral DNA into host genome and also impacts other steps of viral replication cycle, all of which are assisted by various cellular proteins. Among them, LEDGF/p75 acts as the IN-to-chromatin tethering factor. However, whether other cellular cofactors also participate in this process still remains elusive. To gain insight into the mechanism of action of HIV-1 IN during viral integration, we used a previously described IN/yeast lethality system and our results revealed that the HIV-1 IN-induced yeast lethality absolutely required its chromatin binding ability. Since there is no yeast homolog of LEDGF/p75, it raises the possibility that IN may recruit other cellular cofactors for its chromatin targeting. Consistently, further analysis in mammalian cells indicated that HIV-1 IN was able to mediate chromatin binding independent of IN-LEDGF/p75 interaction and that HIV-1 fitness relied more on chromatin binding than LEDGF/p75 binding of IN. These data greatly enrich our current knowledge on the dynamic interplay within the ternary complex IN/LEDGF/chromatin.
HIV-1 exploits multiple cellular cofactors not only to facilitate viral replication, but also to evade the host defense system in favor of the virus. IN is known to be an unstable protein, degraded by the host ubiquitin-proteasome pathway. To investigate how IN avoids the host degradation machinery in the context of viral infection, we showed that IN interacted with host protein Ku70 and protected itself from the Lys48-linked polyubiquitination proteasomal pathway. More importantly, Ku70 was shown to be incorporated into the progeny virus in an IN-dependent manner, and both cell- and virus- associated Ku70 were essential for HIV-1 replication. Finally, the data demonstrated that the interactions between HIV-1 IN and host cofactors can be regulated through its SUMO-interacting motifs (SIMs). Three putative SIMs (72VILV75; 200IVDI203 and 257IKII260) in IN were examined and shown to be essential for IN-LEDGF/p75 but not IN-Ku70 interaction.
In summary, this study advances our knowledge of the interaction network between IN and its cofactors, which would have important implications for the design of anti-HIV drugs.
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Functional analysis on the interactions of the human immunodeficiency virus type 1 integrase with its cofactors that regulate viral replicationZheng, Yingfeng 03 1900 (has links)
Like all viruses, the replication of HIV-1 relies heavily on host proteins due to its limited genome products. HIV-1 integrase (IN) catalyzes the integration of viral DNA into host genome and also impacts other steps of viral replication cycle, all of which are assisted by various cellular proteins. Among them, LEDGF/p75 acts as the IN-to-chromatin tethering factor. However, whether other cellular cofactors also participate in this process still remains elusive. To gain insight into the mechanism of action of HIV-1 IN during viral integration, we used a previously described IN/yeast lethality system and our results revealed that the HIV-1 IN-induced yeast lethality absolutely required its chromatin binding ability. Since there is no yeast homolog of LEDGF/p75, it raises the possibility that IN may recruit other cellular cofactors for its chromatin targeting. Consistently, further analysis in mammalian cells indicated that HIV-1 IN was able to mediate chromatin binding independent of IN-LEDGF/p75 interaction and that HIV-1 fitness relied more on chromatin binding than LEDGF/p75 binding of IN. These data greatly enrich our current knowledge on the dynamic interplay within the ternary complex IN/LEDGF/chromatin.
HIV-1 exploits multiple cellular cofactors not only to facilitate viral replication, but also to evade the host defense system in favor of the virus. IN is known to be an unstable protein, degraded by the host ubiquitin-proteasome pathway. To investigate how IN avoids the host degradation machinery in the context of viral infection, we showed that IN interacted with host protein Ku70 and protected itself from the Lys48-linked polyubiquitination proteasomal pathway. More importantly, Ku70 was shown to be incorporated into the progeny virus in an IN-dependent manner, and both cell- and virus- associated Ku70 were essential for HIV-1 replication. Finally, the data demonstrated that the interactions between HIV-1 IN and host cofactors can be regulated through its SUMO-interacting motifs (SIMs). Three putative SIMs (72VILV75; 200IVDI203 and 257IKII260) in IN were examined and shown to be essential for IN-LEDGF/p75 but not IN-Ku70 interaction.
In summary, this study advances our knowledge of the interaction network between IN and its cofactors, which would have important implications for the design of anti-HIV drugs.
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Rôle de la protéine de réparation de l'ADN Ku dans la régulation traductionnelle de l'ARNm p53 / Role of the DNA repair protein Ku in the translational regulation of p53 mRNALamaa-Mallak, Assala 08 December 2015 (has links)
L'augmentation des taux cellulaires de p53 en réponse aux dommages à l'ADN a été largement attribuée à une augmentation de la demi-vie de la protéine. Il est maintenant bien établi que la régulation traductionnelle de l'ARNm de p53 est également critique à la fois pour la répression de l'accumulation de p53 dans des conditions normales, et l'induction de la protéine en réponse aux dommages de l'ADN. Nos travaux se sont accès sur l'étude du rôle de facteur de réparation de l'ADN Ku dans la régulation traductionnelle de l'ARNm P53. Nous avons montré que Ku réprime la synthèse de la protéine p53 et l'apoptose p53-dépendante via la liaison à une structure en tige-boucle dans la 5'UTR de l'ARNm. Cependant, la répression traductionnelle exercée par Ku est levée après un stress génotoxique. Le mécanisme sous-jacent implique l'acétylation de Ku qui perturbe les interactions Ku-ARNm p53. Ces résultats suggèrent que la répression traductionnelle de p53 par Ku constitue un nouveau mécanisme cytoprotectif liant la réparation de l'ADN et la traduction des ARNm. / Increases in p53 protein levels after DNA damage have largely been attributed to an increase in the half-life of the p53 protein. It is now well accepted that translational regulation of p53 mRNA is also critical for both repression of p53 accumulation in unstressed conditions and induction of the p53 protein in response to DNA damage. Our work focused on studying the role of DNA repair factor Ku in the regulation of P53 mRNA translation. We showed that Ku represses p53 protein synthesis and p53-mediated apoptosis by binding to a stem-loop structure within the p53 5'UTR. However, Ku-mediated translational repression is relieved after genotoxic stress. The underlying mechanism involves Ku acetylation which disrupts Ku-p53 mRNA interactions. These results suggest that Ku-mediated repression of p53 mRNA translation constitutes a novel cytoprotective mechanism linking DNA repair and mRNA translation.
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Cyclin E provides a link between Cell Cycle, DNA Repair and ApoptosisPlesca, Dragos Costin 18 April 2008 (has links)
No description available.
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Structure and Function Studies of Human T-Lymphotrophic Virus Type 1 p30Bowden, Nadine Yvonne 16 December 2011 (has links)
No description available.
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Mise en évidence d'une fonction non-transcriptionnelle du facteur de transcription homéotique Cdx2 / New non-transcriptional function of the homeotic transcription factor Cdx2Soret, Christine 18 September 2014 (has links)
Le cancer colorectal (CCR) représente la 2ème cause de mortalité par cancer dans les pays industrialisés. De nouveaux traitements permettant de bloquer l’évolution de la maladie sont nécessaires. Il est donc important de mieux connaitre les acteurs impliqués dans la cancérogenèse. Lors du développement du cancer, des gènes suppresseurs de tumeur sont inhibés et des oncogènes sont activés, perturbant ainsi l’équilibre des cellules et les transformant. Au cours de ma thèse, je me suis intéressée à deux gènes homéotiques qui possèdent des rôles opposés dans les CCR. Cdx2 exerce un rôle suppresseur de tumeur, alors que HoxB7 agit comme un oncogène. Mon travail de thèse a permis (i) de mettre en évidence une nouvelle fonction non-transcriptionnelle de Cdx2 : inhibiteur de la réparation des cassures de l'ADN spécifiquement dans le côlon, (ii) et de révéler que le niveau d'expression des gènes Cdx2 et Hoxb7 au sein de la tumeur peut avoir une importance pour le choix du traitement des CCR. / Colorectal cancer is the 2nd cause of mortality by cancer in industrialized countries. New treatments allowing to prevent the evolution of the disease are needed. It is important to better understand the actors implicated in carcinogenesis. During cancer development, tumor suppressor genes are inhibited and oncogenes are activated, thus disrupting the homeostasis of the tissue and transforming the cells. During my thesis, I have been interested in two genes having two opposite functions in CCR : Cdx2 is a tumor suppressor while Hoxb7 acts as an oncogene. My work allows to highlight (i) a new non-transcriptional function of Cdx2 : inhibitor of the reparation of DNA breaks specifically in the colon, (ii) and that the expression level of Cdx2 and Hoxb7 genes inside the tumor can have an importance in the choice of the CCR treatment.
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Regulation of Bax Activation and Apoptosis by Src and Acetylated Mutant p53Woods, Nicholas Taylor 25 August 2008 (has links)
Apoptosis is an inherent suicide mechanism that cells invoke for a variety of reasons including embryo cavitation, tissue homeostasis, excessive DNA damage and aberrant oncogene activation. Apoptosis is regulated by a diverse set of proteins including, but not limited to, the Bcl-2 family. This family set is comprised of both pro-death and pro-survival proteins whose relative expression, localization and/or modifications regulate the balance between life and death for each cell. The keystones to this system are the proapoptotic proteins Bax and Bak, which are regulated by their conformation and localization. However, the exact mechanisms by which Bax and Bak become activated remains to be resolved. Similarly, research focusing on the cancer cell's ability to deregulate apoptosis by preventing the activation of Bax or Bak will provide further insight into the development of targeted therapies for cancer that will hopefully contribute to the cure of this formidable disease.
Src, the classic oncogenic kinase, has been found to deregulate Bax activation in response to the detachment of a cell from its substratum support thereby preventing anoikis, the Bax-dependent apoptotic response involved in the impairment of metastatic dissemination of cancer. Our findings indicate that Src deregulates this response by altering the relative expression of Bcl-2 family members Mcl-1 and Bim through the PI3-K/Akt and Erk1/2 pathways. However, Src retains its ability to prevent anoikis even in the setting of Akt and Erk1/2 signaling inhibition. Further evaluation of the role of Src in this process revealed that Bif-1, a protein known to associate with and activate Bax, could be directly phosphorylated by Src which prevented the association of Bax with Bif-1 and impaired the anoikis response.
In addition, our studies have also found that Bax activation in response to treatment with type I and II histone deacetylase inhibitors is dependent on the expression of the tumor suppressor p53. Acetylation of p53 at carboxy-terminal lysine residues enhances its transcriptional activity associated with cell cycle arrest and apoptosis. Here, we demonstrate that p53 acetylation at K320/K373/K382 is also required for its transcription-independent functions in Bax activation, ROS production, and apoptosis in response to the histone deacetylase inhibitors (HDACi) SAHA and LAQ824. Knockout of p53 in HCT116 cells markedly reduced HDACi-induced apoptosis. Unexpectedly, expression of transactivation-deficient p53 variants sensitized p53-null cancer cells to HDACi-mediated Bax-dependent apoptosis, whereas knockdown of endogenous mutant p53 inhibited HDACi-induced apoptosis. Evaluation of the mechanisms controlling this response led to the discovery of a novel interaction between p53 and Ku70. The association between these two proteins was acetylation independent, but acetylation of p53 could prevent and disrupt the Ku70/Bax complex and enhance apoptosis. These results suggest a new mechanism of acetylated p53 transcription-independent regulation of apoptosis.
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Predicción de respuesta a tratamiento en pacientes con carcinoma escamoso de cabeza y cuelloPavón Ribas, Miguel Ángel 09 June 2009 (has links)
La cirugía radical como único tratamiento en carcinomas escamosos de cabeza y cuello (CECC) ha sido substituida por protocolos que permiten preservar la función del órgano como la quimioradioterapia (QRT) concomitante y la quimioterapia de inducción (QTI) seguida de radioterapia (RT) / quimioradioterapia o cirugía. El objetivo general de este proyecto de tesis es identificar marcadores moleculares en biopsias pre-tratamiento de CECC localmente avanzado que permitan predecir la evolución clínica del paciente. En este sentido, hemos llevado a cabo dos estudios independientes. En el primero hemos analizado los niveles de expresión de los genes Ku70, K80 y DNA-PKcs del sistema de reparación no homóloga por unión de extremos en biopsias pre-tratamiento de pacientes con CECC tratados con quimioterapia de inducción seguida de RT/QRT o cirugía. En el segundo hemos realizado un estudio de microarrays de expresión para determinar que genes y procesos biológicos están implicados en la respuesta tumoral en pacientes localmente avanzados tratados con QTI seguida de RT/QRT o cirugía, o con quimioradioterapia desde un inicio. Los resultados del primer estudio muestran que los tumores con una respuesta a QTI superior al 50% presentan niveles de expresión de Ku70, tanto de mRNA como de proteína, mayores que los tumores con una respuesta inferior al 50%. Además los pacientes con tumores con niveles de expresión de Ku70 elevados tienen una supervivencia libre de recidiva local (SLRL) y una supervivencia global (SG) mayor que los pacientes con tumores que expresan bajos niveles de dicho marcador. En el segundo estudio hemos identificado tres subtipos de tumores con diferencias en la evolución clínica de los pacientes. Los pacientes con tumores del cluster 1 tienen mayor SLRL y SG que el resto de los pacientes. Su perfil de expresión muestra que presentan una mayor capacidad de migración e invasividad, características de transición epitelio mesénquima, sobre-activación de la vía secretora y un menor grado de diferenciación celular. Los pacientes con tumores del cluster 3 tienen una evolución clínica favorable, presentan una SG y SLRL mayor que el resto de los pacientes. Entre sus características destacan un menor grado de diferenciación celular y una sobre-expresión de genes localizados en las regiones cromosómica 1q21 y 19q13. Los pacientes con tumores del cluster 2 tienen una evolución clínica intermedia ya que presentan una SLRL similar a los del cluster 3 y una SG similar a los del cluster 1. Hemos identificado una serie de marcadores de mal pronóstico (tirosina sulfotransferasa, trombospondina 1, liprina-beta1, fibronectina IIIB y alfa1-prolil-4-hidroxilasa) cuyo mayor expresión aumenta el riego de recidiva local a los 2 años y el riesgo de muerte a los 3 años de seguimiento. También hemos identificado una serie de marcadores de buen pronóstico (Oxidasa dual 1, LYPD3 y GTPasa Rab25) cuya mayor expresión disminuye el riesgo de recidiva local y muerte de los pacientes. / Induction chemotherapy (IC), followed by radiotherapy (RT) / chemoradiotherapy (CRT) or surgery, and concomitant CRT are commonly used to treat locally advanced head and neck squamous cell carcinoma (HNSCC). The objective of this thesis-project is to identify molecular markers in pre-treatment HNSCC biopsies associated with clinical outcome in patients with advanced stage. We have conducted two independent studies. In the first study, we have evaluated the relationship between Ku80, Ku70 or DNA PKcs expression in pre-treatment tumor biopsies, and tumor response in patients treated with IC, followed by RT / CRT or surgery. In the second analysis, we have performed a microarray study to identify genes and biological processes associated with tumor response in patients treated with IC, followed by RT / CRT or surgery, or concomitant CRT.Results of the first study show that tumors with a response to IC higher than 50% have significantly higher mRNA and protein levels for Ku70 than tumors with a response to IC lower than 50%. Moreover, high tumor Ku70 expression was associated with significantly longer local recurrence-free survival (LRFS) and overall survival (OS).In the second microarray study we have identified three tumor subtypes with differences in patient outcome. Patients with tumor subtype1 have a shorter LRLS and OS. These tumors have a higher migration and invasiveness capacity, epithelial-mesenchymal transition, activation of the secretory pathway and lower differentiation grade. Patients with subtype 3 have a longer LRFS and OS. These tumors have a higher differentiation grade and overexpress genes located on chromosome 1q21 and 19q13.Moreover, we have identified a minimum set of genes (TPST1, THBS1, PPF1BP1, FNDC3B, P4HA1, DUOX1, LYPD3 y RAB25) associated with local recurrence and patient survival.
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The mechanism of DNA double-strand break (DSB) resection in human cellsYang, Soo-Hyun 05 November 2013 (has links)
Homologous recombination (HR) repair is critical for the maintenance of genomic stability, as it is involved in the precise repair of DNA double-strand breaks (DSBs) using an intact homologous template for repair. The initiation of 5' strand resection of DNA ends is a critical determinant in this process, which commits cells to HR repair and prevents repair by non-homologous end joining (NHEJ). The human single-stranded DNA (ssDNA) binding complexes, RPA and SOSS1, are involved in regulating DSB signaling and HR repair. In this study, I demonstrate a novel function of SOSS1 in HR repair, in which SOSS1 stimulates hExo1-dependent resection. Despite its poor activity in binding duplex DNA, SOSS1 facilitates hExo1 recruitment to duplex DNA ends and promotes its activity in resection independently of MRN in vitro. MRN(X) is a highly conserved complex that is involved in the early steps of HR repair by regulating DSB resection. MRN interacts with CtIP and constitutes resection machinery that can perform limiting processing on DNA ends. In this study, I also examine the biochemical activities of MRN and CtIP in DSB resection through reconstituted in vitro assays. I show that the ATP-dependent DNA unwinding activity of MRN is responsible for overcoming Ku inhibition of hExo1- and Dna2/BLM-dependent resection activity in vitro. I propose that this unwinding step displaces Ku away from the DNA ends and facilitates the recruitment of hExo1 to the DNA ends for efficient resection. In addition, I show that CtIP can promote overcoming the inhibitory effect of Ku in resection together with MRN. Further, I demonstrate that MRN nuclease activity is required for efficient processing of covalent adducts from DNA ends in vitro, suggesting that the nucleolytic removal of covalent adducts by MRN generates free ends for hExo1- and Dna2/BLM binding. Overall, this study provides mechanistic insight into the regulation of DSB resection in human cells. / text
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