Дисертації з теми "Microrna targets"
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Sætrom, Ola. "Predicting MicroRNA targets." Thesis, Norwegian University of Science and Technology, Department of Computer and Information Science, 2005. http://urn.kb.se/resolve?urn=urn:nbn:no:ntnu:diva-9266.
Повний текст джерелаMicroRNAs are a large family of short non-encoding RNAs that regulated protein production by binding to mRNAs. A single miRNA can regulate an mRNA by itself, or several miRNAs can cooperate in regulating the mRNAs. This is all dependent on the degree of complementarity between the miRNA and the target mRNA. Here, we present the program TargetBoost that, using a classifier generated by a combination of hardware accelerated genetic programming and boosting, allows for screening several large dataset against several miRNAs, and computes a likelihood of that genes in the dataset is regulated by the set of miRNAs used in the screening. We also present results from comparison of several different scoring functions for measuring cooperative effects. We found that the classifier used in TargetBoost is best for finding target sites that regulate mRNAs by themselves. A demo of TargetBoost can be found on http://www.interagon.com/demo.
Migliore, Chiara Maria. "RNA-sequencing based identification of microRNA-204 targets." Doctoral thesis, Università degli studi di Trieste, 2011. http://hdl.handle.net/10077/4595.
Повний текст джерелаWith the completion of the sequencing and annotation of hundreds of genomes, and the accumulation of data on the mammalian transcriptome, greater emphasis has been placed on elucidating the function of non-coding DNA and RNA sequences. It is well known that the non-coding portion of the genome can transcribe functional RNAs. Several categories of non-coding RNAs (ncRNAs) have been defined, such as transport RNAs (tRNAs) ribosomal RNAs (rRNAs), small nuclear RNAs (snRNAs) and small nucleolar RNAs (snoRNAs). A larger group of ncRNAs comprises the so-called microRNAs (miRNAs) and long non-coding RNAs serving key regulatory roles. It has been shown that miRNAs directly target a large number of genes, thus affecting significantly major pathways. In my project, I focused on miR-204, a microRNA that is highly conserved from zebrafish to human and located in the sixth intron of the human TRPM3 gene. I sought to identify mir-204 targets by using the Medaka fish (Oryzias latipes), where mir-204 is expressed at very low levels in the nervous system, as a model for perturbation of the mir-204 network. Transient transgenic Medaka fish were produced to knock down and over-express mir-204. Next-generation sequencing was used to sequence the Medaka transcriptome, dissect the putative targets of miR-204, and thus gain further insight about its function. Potential target genes of mir-204 were selected by choosing genes, which presented lower expression in the wild-type (wt) fish than in the knock down, a lower expression in the over-expression than in the wt and, finally, a higher expression in the knock down than in the over-expression. At the same time, I collected a list of putative miR-204 mouse and human targets using the prediction softwares miRanda, PicTar and TargetScan, obtained the Medaka orthologues and verified that the selected genes in Medaka had a statistically significant enrichment in miR-204 targets as compared to the complete set of genes obtained from the RNA-Sequencing approach. The combined RNA-Sequencing and bioinformatics analysis revealed 147 predicted targets of mir-204, which showed a significant enrichment for the axon guidance pathway. In order to confirm this data, real time quantitative PCR has been performed on total RNA from wt and morphant fish. Results showed a higher expression in the knock down fish for 15 out of 25 putative targets (Neo1, Trim71, Ddx3y, Prkar1a, MyoX, Sema3B, Sema3F, Ptprg, Slit2, Epha4, Epha7, Amot, Lpp, Odz4, Jarid2). I further validated these genes by both Q-PCR and luciferase assays. To this aim, I cloned five putative target sequences into the 3’UTR of a luciferase reporter vector (pGL3-TK-luc Promega) to use them in luciferase assays: co-transfection with miR-204 reduced the luciferase activity of Sema3F, belonging to the class of receptors involved upstream of the axon guidance pathway. These results indicate that mir-204 directly targets key genes involved in the axon guidance pathway such as Sema3F in the nervous system. Further validation of the disruption of axon guidance in the transgenic fish has been undertaken in vivo by our collaborators: the experiment demonstrated a clear role of this microRNA in axon path finding during retinal development.
XXII Ciclo
1981
Wang, Qi. "Using Imputed Microrna Regulation Based on Weighted Ranked Expression and Putative Microrna Targets and Analysis of Variance to Select Micrornas for Predicting Prostate Cancer Recurrence." Thesis, North Dakota State University, 2014. https://hdl.handle.net/10365/27341.
Повний текст джерелаDavis, M. P. "Generation of a murine ES cell system deficient in microRNA processing for the identification of microRNA targets." Thesis, University of Cambridge, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.598389.
Повний текст джерелаTorkey, Hanaa A. "Machine Learning Approaches for Identifying microRNA Targets and Conserved Protein Complexes." Diss., Virginia Tech, 2017. http://hdl.handle.net/10919/77536.
Повний текст джерелаPh. D.
Woodcock, M. Ryan. "Network Analysis and Comparative Phylogenomics of MicroRNAs and their Respective Messenger RNA Targets Using Twelve Drosophila species." VCU Scholars Compass, 2010. http://scholarscompass.vcu.edu/etd/155.
Повний текст джерелаBudd, William. "Development and Implementation of a Tissue Specific MicroRNA Prediction Tool for Identifying Targets of the Tumor Suppressor microRNA 17-3p." VCU Scholars Compass, 2010. http://scholarscompass.vcu.edu/etd/2116.
Повний текст джерелаJoo, Lauren Jin Suk. "RET-regulated microRNAs as Recurrence Biomarkers and Therapeutic Targets in Medullary Thyroid Carcinoma." Thesis, The University of Sydney, 2018. http://hdl.handle.net/2123/19945.
Повний текст джерелаRose, Jarod. "An Investigation and Visualization of MicroRNA Targets and Gene Expressions and Their Use in Classifying Cancer Samples." University of Akron / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=akron1302303717.
Повний текст джерелаYoussef, Ninwa. "Analysis of conserved microRNA targets in the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster." Thesis, Södertörns högskola, Institutionen för naturvetenskap, miljö och teknik, 2013. http://urn.kb.se/resolve?urn=urn:nbn:se:sh:diva-19211.
Повний текст джерелаMartinez-Nunez, Rocio Teresa. "Role of microRNA-155 in dendritic cells and macrophages : MiR-155 directly targets PU.1 and IL13Rα1". Thesis, University of Southampton, 2010. https://eprints.soton.ac.uk/196569/.
Повний текст джерелаSamols, Mark Atienza. "Identification and Functional Analysis of Micro-RNAs Encoded by Kaposi’s Sarcoma-Associated Herpesvirus." Case Western Reserve University School of Graduate Studies / OhioLINK, 2007. http://rave.ohiolink.edu/etdc/view?acc_num=case1181143062.
Повний текст джерелаCONTU, RICCARDO. "MIRNA-1 IN THE HEART: ITS TARGETS AND ROLE IN CARDIAC HYPERTROPHY AND DIABETIC CARDIOMYOPATHY." Doctoral thesis, Università degli Studi di Milano, 2012. http://hdl.handle.net/2434/168776.
Повний текст джерелаSavary, Grégoire. "Rôles et mécanismes d’action des microARN dans la fibrogenèse : applications thérapeutiques et diagnostiques dans les fibroses pulmonaires et rénales." Thesis, Université Côte d'Azur (ComUE), 2016. http://www.theses.fr/2016AZUR4151.
Повний текст джерелаFibrotic diseases are characterized by the accumulation of extracellular matrix components in response to chronic aggression leading to the destruction of tissue architecture. Myofibroblasts, controlled by TGFβ, are central effectors in this process. MiRNAs, negative regulator of gene expression, are involved in many patho-physiological mechanisms, including tissue fibrosis but their mechanisms of action and the potential synergistic activity between co-regulated miRNAs within the same cluster remain poorly understood. Our work consisted in characterizing the involvement of the miR-199/214 cluster, generated from LncRNA DNM3os, in pulmonary fibrosis. We have shown that these miRNAs are involved in the activation and the differentiation of fibroblasts to myofibroblasts through the regulation of canonical and non-canonical TGF-β pathways. This cluster also acts as an inhibitor of epithelial repair. In vivo, the inhibition of one of this “FibromiR”, significantly decrease fibrosis, in a murine lung fibrosis model. In addition, because of their presence and their stability in biological fluids, miRNAs also represent a new class of non-invasive diagnostic or prognostic biomarker. We showed that serum levels of miR-21-5p were increased in patients with severe kidney fibrosis. These studies highlight the importance of miRNAs in pathogenesis of fibrotic disorders and show that they represent new therapeutic targets or non-invasive biomarkers
Alvarez-Saavedra, Matias Alberto. "MicroRNA-132-Dependent Post-Transcriptional Regulation of Clock Entrainment Physiology Via Modulation of Chromatin Remodeling and Translational Control Gene Targets." Thesis, University of Ottawa (Canada), 2010. http://hdl.handle.net/10393/28722.
Повний текст джерелаBranscheid, Anja. "Phosphate homeostasis and posttranscriptional gene regulation during arbuscular mycorrhizal symbiosis in Medicago truncatula." Phd thesis, Universität Potsdam, 2012. http://opus.kobv.de/ubp/volltexte/2012/6210/.
Повний текст джерелаPhosphat ist ein essentieller Bestandteil der pflanzlichen Ernährung und ein Mangel führt zu schwerwiegenden Folgen für Wachstum, Entwicklung und Reproduktion der Pflanze. Eine der wichtigsten Strategien, um einen Mangel an löslichem Phosphat im Boden auszugleichen, ist die arbuskuläre Mykorrhiza, einer Wurzelsymbiose zwischen Pflanzen und im Boden lebenden Mykorrhizapilzen. Die Symbiose dient dem gegenseitigen Nährstoffaustausch, der über bäumchenartige Strukturen in Wurzelzellen, den Arbuskeln, realisiert wird. Über ein weit reichendes Netzwerk im Boden verbessert der Pilz die Phosphatversorgung der Pflanzen, wohingegen die Pflanze photosynthetisch erzeugte Zucker zur Verfügung stellt. Ein erhöhter Phosphatgehalt in der Pflanze führt zur Unterdrückung der Symbiose. Da weitestgehend unbekannt ist, wie genau Pflanzen diese Einschränkung der Symbiose regulieren, kann die Erforschung dieses Zusammenhangs einen wichtigen Beitrag für Agrarwirtschaft und Umweltschutz leisten. Im Rahmen dieser Arbeit konnte durch die Entdeckung eines neuen, bisher unbekannten Zielgens aufgezeigt werden, dass die für den Ausgleich des pflanzlichen Phosphathaushalts wichtige Mikro-RNA (miR) 399 auch in der Regulation der arbuskulären Mykorrhizasymbiose von besonderer Bedeutung ist. MiRNAs regulieren die Aktivität von Zielgenen indem sie die jeweiligen Transkripte durch Bindung für den Abbau markieren. In kolonisierten Wurzeln, insbesondere in arbuskelhaltigen Wurzelzellen, konnte eine erhöhte Anhäufung der miR399 beobachtet werden. Durch das Verfahren der Hochdurchsatz-Sequenzierung des Wurzeldegradoms, bei dem alle abgebauten Transkripte analysiert werden, konnte das neue Zielgen der miR399 Familie, MtPT8, identifiziert werden. Dieses codiert für einen Phosphat-Transporter, der diesen Studien zufolge ausschließlich in mykorrhizierten Wurzeln vorkommt und dessen Transkription auf arbuskelhaltige Zellen beschränkt ist. Mit der Identifizierung dieses neuen Zielgens konnte erstmals der Beweis für die direkte Verbindung der pflanzlichen Phosphathomöostase durch miR399 und der arbuskulären Mykorrhizasymbiose gezeigt werden. Die Untersuchung der physiologischen Funktion dieses mykorrhizaspezifischen Phosphat-Transporters bietet die Möglichkeit, die Zusammenhänge der phosphatabhängigen Regulation der Symbiose aufzuklären und weit reichende Einblicke in die Regulationsmechanismen während der Pflanze-Pilz-Interaktion zu erhalten.
El, Hajj Petra. "New prognosis markers and new targets for therapy in high risk melanoma: evaluation of TYRP1 as a melanoma prognostic marker and its regulation by miRNA(s)." Doctoral thesis, Universite Libre de Bruxelles, 2015. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/209064.
Повний текст джерелаfiable en se basant sur les analyses d’histopathologies de la lésion primitive et est souvent ajustée
durant la progression de la maladie. Notre étude vise à élargir nos observations initiales au niveau
des métastases cutanées et d’évaluer la valeur pronostique de tyrosinase related protein 1 (TYRP1)
dans les métastases ganglionnaires des patients atteints de mélanome de stades III et IV. TYRP1 est
une enzyme mélanosomale qui partage des similitudes structurelles avec la tyrosinase, l'enzyme clé
de la mélanogenèse.
L’expression de l'ARNm de TYRP1 a été quantifiée dans 104 métastases ganglionnaires par PCR
en temps réel et normalisée par rapport à l’expression de l’ARNm de S100B (marqueur reconnu du
mélanome) pour corriger l’expression de TYRP1 suivant la charge tumorale de l’échantillon. Le
rapport TYRP1/S100B a été calculé et la médiane a été utilisée en tant que valeur seuil. Ensuite
nous avons étudié la relation entre les valeurs de TYRP1/S100B, le suivi clinique et les
caractéristiques histopathologiques de la tumeur primitive.
Un rapport élevé de l’ARNm TYRP1/S100B corrélait significativement avec une survie sans
récidive et une survie globale plus courtes, avec une épaisseur de Breslow plus élevée et avec la
présence d'une ulcération au niveau de la tumeur primitive. En outre, une expression élevée de
TYRP1/S100B était de meilleure valeur pronostique pour la survie globale que l'épaisseur de
Breslow et l'ulcération des primitifs. De plus, cette expression est bien conservée au cours de la
progression de la maladie par rapport aux groupes de TYRP1 bas/élevé.
Nous avons constaté qu’une expression élevée de TYRP1/S100B dans les métastases de patients
atteints de mélanome est associée à un résultat clinique défavorable et une survie courte. Menée sur
des patients atteints d'un mélanome à haut risque de récidive, cette première étude a suggéré que
l'ARNm de TYRP1 dans les métastases pourrait servir de biomarqueur pour affiner le pronostic
initial des patients surtout ceux ayant des lésions primitives de localisation inconnues ou non
évaluables et peut permettre une gestion différente des deux groupes de patients. Son expression
conservée au cours de la progression de la maladie est en faveur de son utilisation comme cible
thérapeutique.
En second lieu, en évaluant l’expression de la protéine TYRP1 par immunohistochimie dans les
métastases cutanées et ganglionnaires, nous avons observé qu’elle n'était pas détectée dans la moitié
7
des tissus exprimant bel et bien l'ARNm correspondant et qu’elle, contrairement à l'ARNm, n’était
pas associée à la survie.
Des données récentes ont indiqué que le 3'-UTR de l’ARNm de TYRP1 contient trois sites de
liaison putatifs de miR-155 dont deux présentant un polymorphisme d'un seul nucléotide (SNPs:
rs683 et rs910) qui favorisent la dégradation en cas d’hybridation miARN-ARNm parfaite de
l’ARNm ou non en cas d’hybridation imparfaite. Nous avons cherché à examiner si miR-155 peut
affecter l’expression de l’ARNm et de la protéine TYRP1 en fonction de ces SNPs. Tout d'abord,
nous avons transfecté deux lignées de mélanome ayant chacune l’une ou l’autre de l’allèle (au
niveau rs683 et rs910) avec différentes concentrations de pré-miR-155 et nous avons évalué
l’expression du miR-155 et l’ARNm TYRP1 par PCR en temps réel ainsi que l’expression de la
protéine TYRP1 par western blot. Nous avons constaté qu’une surexpression de miR-155 a induit
une dégradation importante des ARNm TYRP1 et a perturbé sa traduction en protéine dans la lignée
avec le génotype “hybridation parfaite”. Ensuite, nous avons examiné l'expression des ARNm et
protéines de TYRP1, le niveau de miR-155 et les SNPs rs683 et rs910 dans 192 échantillons de
métastases cutanées et ganglionnaires de mélanome. Nous avons trouvé que le groupe d'échantillons
avec le génotype “hybridation parfaite” était significativement associé à un niveau de protéine de
TYRP1 plus bas alors qu'aucune différence de niveau d’expression n'a été trouvée pour l’ARNm de
TYRP1 ou miR-155 entre les deux groupes de génotype, confirmant que les SNPs au niveau de 3’-
UTR de TYRP1 peuvent spécifiquement affecter l'expression de la protéine TYRP1. En outre, nous
avons montré que l’ARNm de TYRP1 est inversement corrélé avec l’expression miR-155, mais pas
avec la protéine TYRP1 dans le groupe " hybridation parfaite", alors qu'il corrèle positivement avec
la protéine mais pas avec miR-155 dans le groupe "hybridation imparfaite" où la protéine corrélait
inversement à la survie. Cela montre que les SNPs dans le 3'-UTR de l'ARNm TYRP1 affectent la
régulation de l’ARNm par miR-155 et la traduction en protéine. Ces SNPs rendent la régulation de
l’ARNm et la protéine de TYRP1 indépendante de miR-155 et confèrent une valeur pronostique à
la protéine TYRP1
Doctorat en Sciences biomédicales et pharmaceutiques
info:eu-repo/semantics/nonPublished
Webster, Rebecca. "Complementary investigations of the molecular biology of cancer : assessment of the role of Grb7 in the proliferation and migration of breast cancer cells; and prediction and validation of microRNA targets involved in cancer." University of Western Australia. School of Medicine and Pharmacology, 2008. http://theses.library.uwa.edu.au/adt-WU2008.0179.
Повний текст джерелаGao, Cen. "Research in target specificity based on microRNA-target interaction data." Case Western Reserve University School of Graduate Studies / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=case1275685130.
Повний текст джерелаAbdelhadi, Ep Souki Ouala. "MicroRNA target prediction based upon metastable RNA conformations." Thesis, King's College London (University of London), 2017. https://kclpure.kcl.ac.uk/portal/en/theses/microrna-target-prediction-based-upon-metastable-rna-conformations(9d426d58-d529-4fe7-a98a-1fa2ecdc7a2e).html.
Повний текст джерелаRUBOLINO, CARMELA. "NEW FINDINGS INTO TARGET-DIRECTED MICRORNA DEGRADATION MECHANISM." Doctoral thesis, Università degli Studi di Milano, 2022. https://hdl.handle.net/2434/945228.
Повний текст джерелаPiovezani, Amanda Rusiska. "SIMTar: uma ferramenta para predição de SNPs interferindo em sítios alvos de microRNAs." Universidade de São Paulo, 2013. http://www.teses.usp.br/teses/disponiveis/95/95131/tde-09052014-155546/.
Повний текст джерелаSingle nucleotide polymorphisms (SNPs) can be involved in alteration of not only open reading frame but also important genomic positions of gene regulation process such as transcription initiation sites, splicing sites and microRNA target sites. In particular, the identification of SNPs interfering on microRNA target sites is still an open problem, despite its increasing prominence in recent years due to the discoveries about the microRNA abilities as regulatory elements in the genome and association with severals diseases such as cancer and psychiatric disorders. The computational resources currently available for this purpose (four databases and one tool) are restricted to the analysis of SNPs in the 3UTR (UnTranslated Regions) of mRNAs, where the microRNAs typically bind in order to repress their translation. However, this is a simplification of the problem, since it is already known the gene transcription activation by microRNAs bound to its promoter region, increasing of the effectiveness of negative regulation of translation when microRNAs are bound to the coding region of the gene or binding of microRNA into non-coding RNAs. These resources are also limited to the identification of SNPs in the seed region of miRNAs, and therefore they can only identify sites creation or disruption. However, SNPs located outside this region can not only create and disrupt target sites but also interfere on the stability of miRNAs binding and therefore on the regulation effectiveness. Moreover, considering the target site length, more than one SNP can occur inside of a site and thus, the combination of these SNPs can have an even greater influence on the microRNA binding. Also, current resources do not display which alleles of SNPs or what combinations of them are causing which effect. Finally, these features are restricted to the Homo sapiens and Mus musculus species. This work presents the computational tool SIMTar (SNPs Interfering on MicroRNA Targets), developed to identify SNPs that alter miRNA target sites and fills the mentioned gaps. Finally, it is described an application of SIMTar on the analysis of 114 SNPs associated with schizophrenia, all of them being predicted interfering with miRNA target sites.
Bokobza, Cindy. "Neurodevelopmental impact of perinatal inflammation : targets for neuroprotection?" Thesis, Université de Paris (2019-....), 2019. http://www.theses.fr/2019UNIP7082.
Повний текст джерелаA general consensus regarding neurodevelopmental disorders including Autism Spectrum Disorder (ASD) is that they originate from early development defects in brain formation, leading to altered neuronal circuitry responsible for the pathological behavior. Preterm birth is often linked to the occurrence of inflammation and preterm infants have a ten times higher risk of developing ADS-like symptoms than infants born at term. Moreover, some clinical studies reported ongoing neuroinflammation processes in different brain regions in autistic infants including frontal cortex, hippocampus and cerebellum. The major relay of the environmental response in the brain, including inflammatory responses, is microglia cells, the brain resident macrophages that continuously survey their local environment. Moreover, during development microglia play a critical role during the synaptic pruning and myelination to contribute to the formation of a mature brain. In an inflammatory context, MG are activated and participated to the local release of pro-inflammatory cytokines. Our hypothesis is, therefore, that an exposition to perinatal inflammation impacts on neurodevelopmental defects leading to ASD. Using a mouse model of perinatal inflammation induced by IL-1? injection between post-natal day (P)1-5, this project demonstrates that i) there is region specific inflammation and an impact of perinatal inflammation on the onset of ASD-like phenotypes at different developmental stages in mice and (ii) we successfully identify two new potential targets to limit neurodevelopmental defects: microRNAs and the 5HT-7 receptors. This innovative project has as objective to identify potential diagnosis markers to facilitate an early detection of ASD in premature infants based on inflammatory indicators and new therapeutic targets for neuroprotection
Hanina, Sophie Alexandra. "Identifying direct targets of mouse embryonic stem cell-specific microRNAs." Thesis, University of Cambridge, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.608582.
Повний текст джерелаAgarwal, Vikram. "MicroRNAs : principles of target recognition and developmental roles." Thesis, Massachusetts Institute of Technology, 2015. http://hdl.handle.net/1721.1/101295.
Повний текст джерелаThis electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections.
Cataloged from student-submitted PDF version of thesis.
Includes bibliographical references.
MicroRNAs (miRNAs) are ~21-24 nt non-coding RNAs that mediate the degradation and translational repression of target mRNAs. The genomes of vertebrate organisms encode hundreds of miRNAs, each of which may regulate hundreds of mRNA targets. Thus, miRNAs are crucial post-transcriptional regulators engaged in vast regulatory networks. To date, the characteristics of these networks remain mysterious due to the difficulty of identifying miRNA targets through either experimental or computational means. To understand the physiological roles of miRNAs in animal species, it is of fundamental importance to elucidate the structure of the targeting networks in which they participate. The recognition of a miRNA target is guided largely by perfect Watson-Crick base pairing interactions between nucleotides 2-7 from the 5' end of the miRNA (i.e., the "seed" region) and complementary motifs embedded in the 3' UTRs of the target mRNAs. The prevalence of these motifs throughout the transcriptome poses a challenge to our understanding of how specificity emerges: since the presence of a motif is not sufficient to mediate target repression, what contextual features discriminate effective target sites from ineffective ones? Further complicating this is the proposition that "noncanonical" sites lacking perfect seed pairing might mediate repression, which would expand the potential number of functional target sites by orders of magnitude. In the second chapter of this work, we define the features that predict effective miRNA target sites, incorporating their relative influence into a quantitative model which can outperform existing computational models and experimental approaches in target identification. Though the molecular roles of miRNAs in gene regulation have long been appreciated, the functions of most miRNAs in living organisms has remained elusive. In the third chapter of this work, we discuss the consequences of genetic ablation of miR-196, a deeply conserved miRNA that is predicted to simultaneously repress many HOX genes, in the mouse. We propose a role for miR-196 in the spatial patterning of the vertebrate axial skeleton. Isolating the cell populations that express the miRNA during early mammalian development, we attempt to characterize the direct in vivo targets of miR-196 and dissect the molecular underpinnings of the phenotypes observed.
by Vikram Agarwal.
Ph. D.
Ebert, Margaret S. (Margaret Sarah). "Molecular titration by MicroRNAs and target mimic inhibitors." Thesis, Massachusetts Institute of Technology, 2010. http://hdl.handle.net/1721.1/61886.
Повний текст джерелаVita. Cataloged from PDF version of thesis.
Includes bibliographical references.
MicroRNAs (miRNAs) are short, highly conserved non-coding RNA molecules that repress gene expression in a sequence-dependent manner. Each miRNA is predicted to target hundreds of genes, and a majority of protein-coding genes are computationally predicted to be miRNA targets. To test miRNA functions experimentally, we introduced the miRNA "sponge" method, which uses miRNA target mimics to sequester mature miRNAs and thereby create continuous miRNA loss of function in cell lines and transgenic organisms. Sponge RNAs contain complementary binding sites to a miRNA of interest, and are produced from transgenes within cells. As with most miRNA target genes, a sponge's binding sites are specific to the miRNA seed region, which allows them to block a whole family of related miRNAs. This transgenic approach has proven to be a powerful tool to generate miRNA phenotypes in a variety of experimental systems. Bulk measurements on populations of cells have indicated that, although pervasive, repression due to miRNAs is on average quite modest. To assay repression in single cells, we performed quantitative fluorescence microscopy and flow cytometry to monitor a target gene's protein expression in the presence and absence of regulation by miRNA. We found that repression among individual cells varies dramatically. miRNAs establish a threshold level of target mRNA below which protein production is highly repressed and above which expression responds ultra sensitively to target mRNA input until reaching high enough mRNA levels to almost escape repression. We constructed a mathematical model describing molecular titration of target mRNAs by miRNAs. The model predicted, and experiments confirmed, that the ultrasensitive regime could be shifted to higher target mRNA levels by increasing the miRNA concentration or the number of miRNA binding sites in the 3' untranslated region (UTR) of the target mRNA. Thus even a single species of miRNA can act both as a switch to effectively silence gene expression and as a fine-tuner of gene expression. This fits the emerging paradigm in which miRNAs help to confer robustness to biological processes by reinforcing transcriptional programs, attenuating leaky transcripts, and perhaps buffering random fluctuations in transcript copy number.
by Margaret S. Ebert.
Ph.D.
Liu, Wai-man Raymond, and 廖偉文. "Identification of microRNA-184 target genes in squamous cell carcinomaof tongue." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2010. http://hub.hku.hk/bib/B45208207.
Повний текст джерелаViaut, Camille. "Investigating microRNA-target interactions during skeletal muscle development in chicken embryos." Thesis, University of East Anglia, 2017. https://ueaeprints.uea.ac.uk/63691/.
Повний текст джерелаZahra, Latib. "Identification of novel microRNAs as potential biomarkers for the early diagnosis of ovarian cancer using an in-silico approach." University of the Western Cape, 2019. http://hdl.handle.net/11394/7415.
Повний текст джерелаOvarian cancer (OC) is the most fatal gynaecologic malignancy that is generally diagnosed in the advanced stages, resulting in a low survival rate of about 40%. This emphasizes the need to identify a biomarker that can allow for accurate diagnosis at stage I. MicroRNAs (miRNAs) are appealing as biomarkers due to their stability, non-invasiveness, and differential expression in tumour tissue compared to healthy tissue. Since they are non-coding, their biological functions can be uncovered by examining their target genes and thus identifying their regulatory pathways and processes. This study aimed to identify miRNAs and genes as candidate biomarkers for early stage OC diagnosis, through two distinct in silico approaches. The first pipeline was based on sequence similarity between miRNAs with a proven mechanism in OC and miRNAs with no known role. This resulted in 9 candidate miRNAs, that have not been previously implicated in OC, that showed 90-99% similarity to a miRNA involved in OC. Following a series of in silico experimentations, it was uncovered that these miRNAs share 12 gene targets that are expressed in the ovary and also have proven implications in the disease. Since the miRNAs target genes contribute to OC onset and progression, it strengthens the notion that the miRNAs may be dysregulated as well. Using TCGA, the second pipeline involved analysing patient clinical data along with implementing statistical measures to isolate miRNAs and genes with high expression in OC. This resulted in 26 miRNAs and 25 genes being shortlisted as the potential candidates for OC management. It was also noted that targeting interactions occur between 15 miRNAs and 16 genes identified through this pipeline. In total, 35 miRNAs and 37 genes were identified from both pipelines.
Park, Jong Kook. "Target Identification, Therapeutic Application and Maturation Mechanism of microRNAs." The Ohio State University, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=osu1331096696.
Повний текст джерелаSchulz, Nikola. "microRNA profiling and target identification in a mouse model for allergic asthma." Diss., lmu, 2012. http://nbn-resolving.de/urn:nbn:de:bvb:19-143012.
Повний текст джерелаSualp, Merter. "Machine Learning Methods For Using Network Based Information In Microrna Target Prediction." Phd thesis, METU, 2013. http://etd.lib.metu.edu.tr/upload/12615645/index.pdf.
Повний текст джерелаSamenuk, Thomas. "Incorporation of Organ-Specific MicroRNA Target Sequences to Improve Gene Therapy Specificity:." Thesis, Boston College, 2021. http://hdl.handle.net/2345/bc-ir:109174.
Повний текст джерелаThe aim of this study was to utilize a massively parallel reporter assay (MPRA) to identify organ-specific microRNA (miRNA) target sequences to refine the timing and expression of transgene expression for gene therapy. We previously had developed a cardiac gene therapy for Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT) using a systemically delivered adeno-associated virus (AAV9) vector. We hypothesized that incorporation of organ specific miRNA target sites into our vector construct could improve our therapy’s tissue specificity due to the ability of miRNAs to silence transgene expression. Initially, we attempted to incorporate mir-124 target sequences into our vector to detarget the brain. Although these initial attempts were unsuccessful, the study allowed us to develop a protocol to test the effectiveness of miRNA target sequences. Thereafter, we developed a method to screen thousands of putative miRNA target sequences simultaneously. In this study, target sequences of miRNAs specific to the heart, brain and liver were incorporated into a plasmid library. This plasmid library was subsequently made into AAV and injected into mice from a CPVT transgenic line. Total DNA and RNA was later extracted from the target organs, converted into genomic DNA (gDNA) and complementary DNA (cDNA) libraries respectively, and sent for amplicon sequencing. We analyzed the results using Comparative Microbiome Analysis 2.0 software (CoMA) and a custom python script to count the occurrence of each specified barcode per sample. In doing so, we showed that the miRNA suppression mechanism is not only effective but also organ specific. Furthermore, we developed a second script to create a combinatorial library from a set list of miRNA target sequences enabling us to efficiently test thousands of target sequence combinations at once. In doing so, we will be able to identify effective miRNA target sequence combinations to further improve gene therapy specificity
Thesis (BS) — Boston College, 2021
Submitted to: Boston College. College of Arts and Sciences
Discipline: Departmental Honors
Discipline: Biology
Winata, Patrick. "The Development of Artificial microRNAs (amiRs) to Target Multiple Oncogenes in Malignant Pleural Mesothelioma (MPM)." Thesis, The University of Sydney, 2018. https://hdl.handle.net/2123/21883.
Повний текст джерелаChow, Hiu Tung. "Arabidopsis miR163 and its target are involved in defense against Pseudomonas syringae." HKBU Institutional Repository, 2016. https://repository.hkbu.edu.hk/etd_oa/312.
Повний текст джерелаAriede, Jovita Ramos. "MicroRNoma dos carcinomas de fígado." Botucatu, 2017. http://hdl.handle.net/11449/148986.
Повний текст джерелаResumo: Introdução: O carcinoma hepatocelular (CHC) está entre as neoplasias de alta complexidade no diagnóstico, determinação do prognóstico e tratamento, sendo o sexto tipo de câncer mais comum no mundo e a segunda causa mais comum de morte por câncer. Uma variedade de genes alterados foram relatados, revelando heterogeneidade genética entre tumores de diferentes pacientes. Portanto, o insucesso terapêutico da terapia convencional pode ser parcialmente atribuído a essa heterogeneidade associada ao comportamento biológico tumoral. Sendo assim, justifica-se a necessidade de identificação de vias moleculares as quais podem conter biomarcadores clinicamente aplicáveis para a melhoria do diagnóstico e tratamento dos pacientes com CHC. Os resultados podem ter futuras aplicações clínicas utilizando miRNAs e os genes-alvo regulados por miRNAs como biomarcadores com valor diagnóstico, prognóstico e terapêutico. Objetivos: Identificar o perfil global de expressão de miRNAs (microRNoma) e os mRNAs-alvo potencialmente regulados por miRNAs em CHC. Pacientes e Métodos: Foram incluídas 18 amostras de tecido, fixadas em formalina e emblocadas em parafina (FFPE) de carcinoma hepatocelular, sendo 18 amostras tumorais e 18 amostras de tecido hepático histologicamente normal, adjacente ao tumor, dos mesmos pacientes. O perfil de expressão de miRNAs das amostras tumorais foi determinado utilizando o ensaio TaqMan Array Human MicroRAN Cards (TLDA) (card A) (Life Technologies). A análise dos dados util... (Resumo completo, clicar acesso eletrônico abaixo)
Doutor
Tan, Yi. "Functional analysis of microRNA-181a : identification of target proteins and application in HCC therapy." Thesis, Imperial College London, 2014. http://hdl.handle.net/10044/1/24698.
Повний текст джерелаParveen, Alisha [Verfasser], and Norbert [Akademischer Betreuer] Gretz. "Advanced hierarchical learning approach for microRNA and target prediction / Alisha Parveen ; Betreuer: Norbert Gretz." Heidelberg : Universitätsbibliothek Heidelberg, 2020. http://d-nb.info/1205807489/34.
Повний текст джерелаFleischmann, Katrin Kristina. "Identification of MLL-A9 related target genes and microRNAs involved in leukemogenesis." Diss., Ludwig-Maximilians-Universität München, 2012. http://nbn-resolving.de/urn:nbn:de:bvb:19-161310.
Повний текст джерелаMalik, Juliane [Verfasser]. "Evaluierung der diagnostischen Rolle von microRNAs und Validierung ihrer Targets in hepatozellulärem Karzinom / Juliane Malik." Ulm : Universität Ulm, 2021. http://d-nb.info/1238690440/34.
Повний текст джерелаRieger, Megan Elizabeth. "Transcription Cofactor LBH is a Direct Target of the Oncogenic WNT Pathway with an Important Role in Breast Cancer." Scholarly Repository, 2010. http://scholarlyrepository.miami.edu/oa_dissertations/659.
Повний текст джерелаTorossian, Avédis. "Contrôle de l'expression de Bcl-2 dans les lymphomes anaplasiques à grandes cellules par la protéine HuR en réponse au crizotinib : impact sur l'apoptose et l'autophagie." Thesis, Toulouse 3, 2017. http://www.theses.fr/2017TOU30190/document.
Повний текст джерелаAnaplastic large cell lymphoma (ALCL) are T/-null non-hodgkin lymphoma representing most of childhood T-cell lymphoma (up to 30%). More than 80% of cases bear reciprocal chromosomic translocation responsible for abnormal expression and constitutive activation of X-ALK type (Anaplastic Lymphoma Kinase) chimeric proteins (ALK+ ALCL). A striking characteristic of this lymphoma is that B-Cell Lymphoma-2 (BCL-2) remains undetectable in ALK+ cases compared to ALK- cases. This is all the more surprising as the BCL-2 oncogene, which is firmly established as a prototypic anti-apoptotic factor as well as a key autophagy regulator, has been shown to be overexpressed in a majority of lymphomas. On the other hand, the RNA-binding protein HuR (Human Antigen R) is overexpressed in ALCL (as in most cancers). It has been demonstrated that this protein was involved in the sustainability of the tumoral phenotype, and that its subcellular localization and functions were closely related to its phosphorylation status, which in turn heavily depends on ALK activity in ALK+ ALCL. In the cytoplasm, HuR has the ability to bind adenine and uridine-rich elements (ARE) located on the 3'-UTR of target mRNAs, and both protect them from degradation and increase their translation. From a general point of view, HuR is able to establish an interplay with microRNAs (miRNAs), either blocking them through competition, or actually cooperating with them and thus promote their function of negative regulators of gene expression on common target transcripts. The BCL-2 transcript, which expression seems to be silenced in ALK-expressing ALCL, has been described as a potential target of HuR. During my PhD work, I dedicated myself to understand the molecular mechanism at work in the silencing of BCL-2 expression with a focus on HuR and collaborating miRNA. The data I obtained point at a cooperation between HuR and miR-34a leading to the silencing of the BCL-2 transcript. However, when the ALK tyrosine kinase activity is inhibited, it appears the interaction between the BCL-2 mRNA diminishes, which limitates the miR-34a 's access to this transcript and ultimately results in a re-expression of the BCL-2 oncogene in these lymphoma cells. In the current context of clinical trials for ALK-targeting inhibitors, such as the Crizotinib, this BCL-2 re-expression observed upon ALK inhibition shed light on potential reasons behind some therapeutic failures that have recently been reported. Indeed, during my PhD work, I also studied the consequences of the BCL-2 re-expression observed in Crizotinib-treated cells. The data I obtained in vitro and in vivo show that, by blocking this re-expression using RNA interference, the Crizotinib anti-tumoral efficiency can be greatly potentiated. This potentiation took the form of an increase of apoptotic cell death induction and, interestingly, also affected the autophagic response triggered by the drug, making it switch from a cytoprotective- type, protumoral autophagic flux to an enhanced, deletary-type and tumor suppressive flux, adding to the therapeutic effect of the drug. This work in general provides insights for new therapeutic combinations that could potentially benefit to ALK+ ALCL patients, and illustrates the complex cross-regulations between apoptotic and autophagic pathway
Limbu, Sarita. "Identifying Differentially Expressed Human Lung MicroRNAs and Their Molecular Functions." University of Akron / OhioLINK, 2009. http://rave.ohiolink.edu/etdc/view?acc_num=akron1259543051.
Повний текст джерелаArif, Km Taufiqul. "Functional association of Micrornas with molecular subtypes of breast cancer." Thesis, Queensland University of Technology, 2021. https://eprints.qut.edu.au/213110/1/Km%20Taufiqul_Arif_Thesis.pdf.
Повний текст джерела"Identification of microRNA targets." NEW YORK UNIVERSITY, 2010. http://pqdtopen.proquest.com/#viewpdf?dispub=3365704.
Повний текст джерелаLin, Li_Zen, and 林立人. "Computational prediction of host microRNA targets against virus genome and identification of virus microRNAs." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/73394184058148358652.
Повний текст джерела國立交通大學
生物資訊研究所
94
Recent work has indicated that microRNAs (miRNAs) play important roles on development, oncogenesis and apoptosis by binding to mRNAs to regulate the post-transcriptional level of gene expression in mammals, plants and insects. Notably, miRNAs also can be produced from virus, and the expression of virus genes are potentially regulated by host miRNAs after infecting host. In this thesis, two databases, VirMiR and ViTa, are developed to compile the virus miRNAs and the regulatory relationships between host miRNAs and virus genes, respectively. Known virus microRNAs in four species, such as humans, mouse, rat and chicken, were obtained from miRBase and miRNAMap. Virus information was collected from NCBI, ICTVdB, VBRC and VirGen. Experimentally validated host miRNA targets on viruses are identified via a literature survey. Then, miRanda and TargetScan are applied to identify the miRNA targets within virus genomes. The virus miRNAs were detected based on comparative sequence analysis. Finally, virus annotations, virus infected tissues from literatures and tissue-specificity of miRNAs from MIT are integrated, and the possible relationship between miRNAs and viruses are identified. This work also provides comparisons between subtypes in some common viruses, such as influenza viruses, liver viruses, and conserved regions between different strains of the same virus species for users. Both textual and graphical web interfaces are provided to facilitate data retrieval from the two databases. The two databases can be available on the Internet at http://vita.mbc.nctu.edu.tw/ and http://virmir.mbc.nctu.edu.tw/.
"The Functional Evolution of Human microRNA Families." Doctoral diss., 2016. http://hdl.handle.net/2286/R.I.41220.
Повний текст джерелаDissertation/Thesis
Doctoral Dissertation Molecular and Cellular Biology 2016
Chou, Yi-Hsuan, and 周宜璇. "Genome-Wide RNAi Screen to Identify Genetic Interactions with microRNA-26a, and Its Potential Targets." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/43335235472204846860.
Повний текст джерела國立臺灣大學
生物化學暨分子生物學研究所
99
In post genomic era, a major biological challenge is to observe how genes function as networks to carry out and regulate cellular processes. Genetic interaction analysis is a powerful tool for establishing functional linkages between genes. Hepatocellular carcinoma (HCC) is the third-leading cause of death from cancer and the fifth most common malignancy worldwide. Despite great advances in the diagnosis and treatment of this cancer, relapse and metastasis is largely unavoidable and the 5-year survival rate remains unsatisfactory. MicroRNAs (miRNAs) are a class of highly conserved small RNA molecules that function as critical regulators of gene expression. Mature miRNAs bind to the target mRNAs, resulting in mRNA degradation or translation repression dependent on the sequence complementarily. Importantly, the ability of individual miRNAs to regulate hundreds of mRNAs allows these RNAs to coordinate complex programs of gene expression. Dysregulated miRNA expression has been linked to many human diseases. Previous studies have shown that HCC cells exhibit reduced expression of miR-26a compared to the normal tissue. Activation of nuclear factor κB and interleukin-6 was observed in tumors with reduced miR-26a expression. Moreover, ectopically expressed miR-26a in mouse HCC results in inhibition of cell proliferation and restrains disease progression. These findings indicate that miR-26a may function as a tumor suppressor gene and provide a new approach for HCC treatment. Although the biochemical mechanisms of miRNA have been specified and some candidate miRNA target genes may be predicted by bioinformatics approaches, identification of physiologically relevant targets of individual miRNA remains challenging. In this research, we carried out high-throughput RNA interference screen in human hepatocellular carcinoma cells to identify genes interacting with the query miR-26a gene, which is able to reveal the upstream regulators and downstream effectors. We generated query stable HepG2 cell lines harboring overexpression constructs. We delivered miR26a expression construct into HepG2 cells using lentiviruses followed by TaqMan miRNA assay to confirm the overexpression level. Furthermore, we found that forced expression of miR-26a2 in these stable cell lines render them less capable of growing without anchorage, which is consistent with the proposed tumor suppressor phenotype of this miRNA. Finally, we performed genome-wide genetic interaction screens to identify some interesting genetic interacting partner genes of miR-26a2. These may provide us direct informations on networks, pathways and dynamics of miRNA in hepatocarcinogenesis.
Sims, Emily K. "Microrna 21 targets B Cell Lymphoma 2 (Bcl2) Mrna to increase beta cell apoptosis and exosomal Microrna 21 could serve as a biomarker of developing Type 1 Diabetes Mellitus." 2018. https://doi.org/10.7912/C2T366.
Повний текст джерелаThe role of beta cell miR-21 in Type 1 Diabetes (T1D) pathophysiology has been controversial. Here, we sought to define the context of beta cell miR-21 upregulation in T1D and the phenotype of beta cell miR-21 overexpression through target identification. Furthermore, we sought to identify whether circulating extracellular vesicle (EV) beta cell-derived miR-21 may reflect inflammatory stress within the islet during T1D development.. Results suggest that beta cell miR-21 is increased in in-vivo models of T1D and cytokine-treated cells/islets. miR-21 overexpression decreased cell count and viability, and increased cleaved caspase-3 levels, suggesting increased cell death. In silico prediction tools identified the anti-apoptotic mRNA B Cell Lymphoma 2 (BCL2) as a conserved miR-21 target. Consistent with this, miR-21 overexpression decreased BCL2 transcript and protein expression, while miR-21 inhibition increased BCL2 protein levels and reduced cleaved caspase-3 levels following cytokine-treatment. miR-21-mediated cell death was abrogated in 828/33 cells, which constitutively overexpress BCL-2. Luciferase assays suggested a direct interaction between miR-21 and the BCL2 3’untranslated region. With miR-21 overexpression, PRP revealed a shift of BCL-2 message toward monosome-associated fractions, indicating inhibition of BCL2 translation. Finally, overexpression in dispersed human islets confirmed a reduction in BCL2 transcripts and increased cleaved caspase 3 production. Analysis of EVs from human beta cells and islets exposed to cytokines revealed a 3-5-fold increase in miR-21. Nanoparticle tracking analysis showed no changes in EV quantity in response to cytokines, implicating specific changes within EV cargo as responsible for the miR-21 increase. Circulating EVs from diabetic non-obese diabetic (NOD) mice displayed progressive increases in miR-21 that preceded diabetes onset. To validate relevance to human T1D, we assayed serum samples collected from 19 pediatric T1D subjects at the time of diagnosis and 16 healthy controls. Consistent with our NOD data, EV miR-21 was increased 5-fold in T1D samples. In conclusion, in contrast to the pro-survival role reported in other systems, our results demonstrate that miR-21 increases beta cell death via BCL2 transcript degradation and inhibition of BCL2 translation. Furthermore, we propose that EV miR-21 may be a promising marker of developing T1D.
"Identification of molecular targets for Brucein D and metastasis suppressor genes in cancer through microRNA and RNAi screening." 2012. http://library.cuhk.edu.hk/record=b5549124.
Повний текст джерела胰腺癌是一种预后很差的恶性肿瘤,常常在确诊时已发生转移。为了找出在胰腺癌转移过程中发挥决定性作用的基因,我们进行了全基因组范围的RNA干扰筛选。一个包含针对全部人类基因的shRNA文库被导入胰腺癌细胞系capan-2.然后将这些细胞移植到裸鼠的胰腺中来建立一个原位胰腺癌小鼠模型。我们的假设是下调某个基因会促使低转移潜力的capan-2细胞转移到肝脏。通过从肝转移结节中回收shRNA模板,我们找到了几个推定的转移抑制基因。其中之一,SOX9,通过体内实验验证,证明下调SOX9基因的表达可促进胰腺癌转移。
化疗适用于进展期胰腺癌病人。然而他们对一线化疗药吉西他滨的反应并不乐观,这进一步使胰腺癌的预后变差。我们展开了一个全基因组范围的RNA干扰筛选来确定一些在化疗耐药过程中起关键作用的基因。携带上述shRNA文库的capan-2细胞被用于吉西他滨药物处理之下的筛选。通过微阵列分析,一些基因被筛选成为可影响癌细胞对药物敏感性的潜在的靶基因。通过进一步验证,LLGL1基因被确定为在调节癌细胞对化疗敏感性过程中起重要作用的基因。
MicroRNAs (miRNAs) are endogenous small non-coding RNAs that have been shown to play important roles in tumorigenesis. Brucein D (BD), a chemical compound isolated from Brucea javanica fruit, has previously been reported to have anti-cancer effect in pancreatic cancer. In this study, we showed that BD also inhibited the growth of liver cancer cells both in vitro and in vivo. To investigate whether BD exerts its anti-cancer effect through regulation of miRNAs, we performed a cancer miRNA qPCR array profiling. From the profiling, miR-95 was found to be significantly down-regulated after BD treatment. Subsequently, a pro-apoptotic gene CUGBP2 was identified as a direct downstream target of miR-95. These findings suggested BD suppressed liver cancer cell growth through down-regulation of miR-95 and reinforcing CUGBP2.
Pancreatic cancer is an aggressive malignancy with extremely poor prognosis. It is usually diagnosed when metastases are already present. To identify genes that play critical roles in the processes of pancreatic cancer metastasis, a whole genome RNAi screening was performed. An shRNA library targeting all human genes was introduced into a human pancreatic cancer cell line capan-2. The infected cells were then transplanted into the pancreas of nude mice. Because capan-2 is of low metastatic potential, we hypothesized that knocking down of metastasis suppressor genes would facilitate capan-2 cells to spread to the liver. By retrieving shRNA templates from the liver metastatic nodules, several candidate genes were found. One of them, SOX9, has been validated as metastasis suppressor gene in vivo, implying that loss of expression of SOX9 promotes pancreatic cancer metastasis.
Chemotherapy is recommended for patients of pancreatic cancer in advanced stage. However, their response to the first-line chemotherapy drug gemcitabine is not satisfactory. A genome-wide RNAi screening was conducted to identify genes that were critical in chemotherapy resistance. Capan-2 cells containing the above shRNA library were applied for the screening under gemcitabine treatment. Through microarray analysis, a number of genes were screened as potential gemcitabine sensitivity genes. Validation experiments implied that the gene LLGL1 may play an important role in modulating pancreatic cancer cells’ sensitivity to gemcitabine.
Detailed summary in vernacular field only.
Detailed summary in vernacular field only.
Detailed summary in vernacular field only.
Xia, Tian.
Thesis (M.Phil.)--Chinese University of Hong Kong, 2012.
Includes bibliographical references (leaves 125-134).
Abstracts also in Chinese.
Chapter Abstract --- p.I
Chapter Acknowledgements --- p.V
Chapter Abbreviations --- p.VI
Chapter List of Figures --- p.XV
Chapter List of Tables --- p.XVI
Chapter Part I: --- Brucein D-modulated microRNA-95 expression inhibits hepatocellular carcinoma cell growth --- p.1
Chapter 1 --- Introduction --- p.1
Chapter 1.1 --- Hepatocellular carcinoma --- p.1
Chapter 1.1.1 --- Definition and classification --- p.1
Chapter 1.1.2 --- Epidemiology --- p.1
Chapter 1.1.3 --- Etiology --- p.3
Chapter 1.1.4 --- Molecular pathogenesis of HCC --- p.4
Chapter 1.1.4.1 --- Genomic instability --- p.4
Chapter 1.1.4.2 --- Deregulation of key signaling pathways --- p.5
Chapter 1.1.4.3 --- Epigenetic changes of HCC --- p.6
Chapter 1.1.4.4 --- Two models of HCC pathogenesis --- p.7
Chapter 1.1.5 --- Therapeutic methods and prognosis of HCC --- p.8
Chapter 1.2 --- Apoptosis --- p.9
Chapter 1.2.1 --- Types of cell death --- p.9
Chapter 1.2.2 --- Apoptosis --- p.10
Chapter 1.2.3 --- Morphological features of apoptosis --- p.10
Chapter 1.2.4 --- Molecular mechanisms of apoptosis --- p.11
Chapter 1.2.5 --- Apoptosis and cancer --- p.13
Chapter 1.2.5.1 --- Imbalance of pro-apoptotic proteins and anti-apoptotic proteins --- p.13
Chapter 1.2.5.2 --- Impaired caspases activity --- p.14
Chapter 1.2.5.3 --- Deregulated death receptor signaling --- p.15
Chapter 1.2.6 --- Cancer therapy targeting apoptotic defects --- p.15
Chapter 1.3 --- microRNA --- p.16
Chapter 1.3.1 --- Overview --- p.16
Chapter 1.3.2 --- Biogenesis and maturation of microRNA --- p.17
Chapter 1.3.3 --- Gene silencing by microRNA --- p.18
Chapter 1.3.4 --- MicroRNA and cancers --- p.19
Chapter 1.3.5 --- MicroRNA’s involvement in HCC --- p.21
Chapter 1.3.6 --- Involvement of miR-95 in cancer --- p.22
Chapter 1.4 --- Brucein D --- p.22
Chapter 1.5 --- Aims of study --- p.23
Chapter 2 --- Materials and Methods --- p.25
Chapter 2.1 --- Cell Culture --- p.25
Chapter 2.1.1 --- Mammalian Cell Culture --- p.25
Chapter 2.1.2 --- Preparation of cell stock --- p.25
Chapter 2.1.3 --- Cell recovery from liquid nitrogen stock --- p.26
Chapter 2.1.4 --- Preparation of drugs for treatments --- p.26
Chapter 2.1.5 --- Drug treatment --- p.26
Chapter 2.1.6 --- Transfection of siRNA --- p.27
Chapter 2.1.7 --- MTT Assay --- p.28
Chapter 2.1.8 --- Luciferase reporter assays --- p.28
Chapter 2.1.9 --- Annexin V Assay --- p.29
Chapter 2.2 --- In vivo mouse model --- p.29
Chapter 2.3 --- Tunel Assay (Terminal deoxynucleotide transferase dUTP Nick End Labeling Assay) --- p.30
Chapter 2.4 --- RNA manipulation --- p.31
Chapter 2.4.1 --- RNA Isolation --- p.31
Chapter 2.4.2 --- Synthesis of cDNA from miRNA --- p.32
Chapter 2.4.3 --- Synthesis of cDNA from RNA and quantitative PCR --- p.33
Chapter 2.4.4 --- miRNA qPCR array --- p.34
Chapter 2.5 --- DNA manipulation --- p.34
Chapter 2.5.1 --- Agarose gel electrophoresis and purification of DNA --- p.34
Chapter 2.5.2 --- Restriction enzymes digestion --- p.35
Chapter 2.5.3 --- Ligation of DNA fragments --- p.36
Chapter 2.5.4 --- Polymerase chain reaction --- p.36
Chapter 2.5.5 --- Preparation of competent E. coli cells --- p.37
Chapter 2.5.6 --- Transformation of E. coli cells --- p.37
Chapter 2.5.7 --- Small scale plasmid isolation from E. coli (mini-prep) --- p.38
Chapter 3 --- Results --- p.39
Chapter 3.1 --- Brucein D inhibited the growth of HCC cells both in vitro and in vivo --- p.39
Chapter 3.2 --- BD induced apoptosis in HCC cells --- p.43
Chapter 3.3 --- miR-95 is an target of BD to modulate cell growth --- p.46
Chapter 3.4 --- Identification of CUGBP2 as a downstream target of miR-95 --- p.55
Chapter 4 --- Discussion --- p.60
Chapter Part II: --- Genome-wide RNAi screening identifies tumor metastasis suppressor genes and drug sensitivity genes in pancreatic cancer --- p.65
Chapter 1 --- Introduction --- p.65
Chapter 1.1 --- Pancreatic cancer --- p.65
Chapter 1.1.1 --- Overview --- p.65
Chapter 1.1.2 --- Pancreatic ductal adenocarcinoma (PDAC) --- p.67
Chapter 1.1.3 --- Molecular basis of PDAC --- p.67
Chapter 1.1.3.1 --- KRAS --- p.67
Chapter 1.1.3.2 --- TP53 --- p.68
Chapter 1.1.3.3 --- CDKN2A --- p.69
Chapter 1.1.4 --- Gemcitabine treatment in PDAC --- p.69
Chapter 1.2 --- Metastasis --- p.71
Chapter 1.2.1 --- Overview --- p.71
Chapter 1.2.2 --- The stepwise process of metastasis --- p.72
Chapter 1.2.3 --- Metastasis of pancreatic cancer --- p.74
Chapter 1.3 --- SOX9 --- p.75
Chapter 1.4 --- Aims of study --- p.77
Chapter 2 --- Materials and Method --- p.78
Chapter 2.1 --- Cell culture --- p.78
Chapter 2.1.1 --- Mammalian Cell Culture --- p.78
Chapter 2.1.2 --- MTT Assay --- p.78
Chapter 2.1.3 --- Colony formation assay --- p.79
Chapter 2.1.4 --- Wound healing assay --- p.79
Chapter 2.1.5 --- Transwell migration chamber assay --- p.80
Chapter 2.1.6 --- Immunocytochemistry --- p.80
Chapter 2.1.7 --- Transient transfection of siRNA --- p.81
Chapter 2.2 --- Establishment of in-vivo and in-vitro models --- p.82
Chapter 2.2.1 --- shRNA library introduction --- p.82
Chapter 2.2.2 --- Establishment of the orthotopic pancreatic cancer mouse model --- p.82
Chapter 2.2.3 --- Package of lentivirus expressing shRNA --- p.83
Chapter 2.2.4 --- Generation of stable cell line expressing shRNA --- p.84
Chapter 2.3 --- DNA manipulation --- p.84
Chapter 2.3.1 --- Large scale plasmid isolation from E. coli (maxi-prep) --- p.84
Chapter 2.4 --- Analysis of Protein --- p.85
Chapter 2.4.1 --- Preparation of protein cell lysates --- p.85
Chapter 2.4.2 --- Protein concentration determination --- p.86
Chapter 2.4.3 --- SDS-PAGE --- p.86
Chapter 2.4.4 --- Immunoblotting (Western blotting) --- p.87
Chapter 2.5 --- RNA manipulations --- p.88
Chapter 2.5.1 --- RNA Isolation --- p.88
Chapter 2.5.2 --- Synthesis of cDNA from RNA and quantitative PCR --- p.89
Chapter 2.6 --- Analysis of Clinical Samples --- p.90
Chapter 2.6.1 --- Clinical specimens --- p.90
Chapter 2.6.2 --- Immunohistochemistry --- p.90
Chapter 3 --- Results --- p.92
Chapter 3.1 --- Genome-wide RNAi screening identifies genes as metastasis suppressors in an orthotopic pancreatic cancer mouse model --- p.92
Chapter 3.2 --- SOX9 is a metastasis suppressor gene in pancreatic cancer --- p.97
Chapter 3.3 --- Investigation into cellular functions of SOX9 --- p.102
Chapter 3.3.1 --- SOX9’s effect on cell growth --- p.102
Chapter 3.3.2 --- SOX9’s effect on cell migration --- p.105
Chapter 3.4 --- Implication of SOX9 in human pancreatic cancer samples --- p.109
Chapter 3.5 --- Genome-wide RNAi screening for the identification of gemcitabine sensitivity genes --- p.113
Chapter 4 --- Discussion --- p.120
Chapter General conclusions --- p.125