Bibliographies thématiques / Singole cellule

Littérature scientifique sur le sujet « Singole cellule »

Auteur : Grafiati
Publié le 11 mars 2023

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Articles de revues sur le sujet "Singole cellule"

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Andreula, C. F., A. M. N. Recchia-Luciani, A. Tarantino, V. Pavone, A. P. Garribba, R. De Blasi et A. Carella. « I linfomi secondari del sistema nervoso centrale ». Rivista di Neuroradiologia 7, no 6 (décembre 1994) : 883–93. http://dx.doi.org/10.1177/197140099400700605.

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Esponiamo i risultati della nostra esperienza nello studio dei linfomi secondari del SNC con la risonanza magnetica, in confronto con i dati disponibili in letteratura. In particolare, abbiamo analizzato i dati epidemiologici e l'eventuale ricaduta delle risultanze della RM sul protocollo diagnostico-terapeutico dei linfomi secondari. Inoltre, abbiamo tentato di identificare delle ipotesi di correlazione tra quadro anatomo-patologico e segnale RM. Nei 10 pazienti sono state individuate 20 lesioni, in 7 casi singole, in 3 multiple, queste ultime da un minimo di 2 a un massimo di 6. Complessivamente le lesioni sono risultate così distribuite: — 15 lesioni intrassiali, 5 delle quali singole, 3 multiple; 1 lesione intra-assiale aveva localizzazione midollare; — 5 lesioni extrassiali, di cui 3 meningo-durali e 2 leptomeningee, tutte singole. Nelle lesioni intrassiali in T1 la zona 1 è apparsa sostanzialmente isointensa (80%) e raramente ipero iso-iperintensa (20%). In T2 si è evidenziata una prevalente iperintensità (70%), raramente una isointensità (20%) o una ipointensità (10%). La zona 2 è risultata evidente nel 30% dei casi. L '80% delle lesioni ha mostrato un potenziamento dopo contrasto, in tutti i casi da moderato a marcato e di aspetto omogeneo. In nessun caso è stata evidenziata una diffusione subependimale. Nelle immagini tardive solo nel 10% dei casi si è osservato un aumento del grado di impregnazione e senza estensione alla zona 2. Le lesioni meningodurali, così come le leptomeningee, si presentano isointense in T1, male apprezzabili in T2, ma vengono rivelate dopo mdc dalla netta impregnazione. All'esame istologico, tali forme secondarie si sono rivelate eterogenee: 5 casi a grandi cellule, 1 a piccole cellule, 1 linfoblastico, 1 tipo Burkitt, 2 linfomi di Hodgkin. In un caso a presentazione contemporanea nel SNC ed in sede periferica, il riscontro istologico cerebrale (a grandi cellule), si è mostrato differente da quello bioptico linfonodale (a piccole cellule). È stata valutata infine la risposta al trattamento, in massima parte chemioterapico; in 2 pazienti questo è stato associato a radioterapia. Si è osservata una regressione o una riduzione volumetrica lesionale nel 50% dei casi, una progressione nel 30%, ed un reperto sostanzialmente invariato nel 20%. La durata minima di tali regressioni è stata di circa 2 mesi. Solo un paziente è attualmente in remissione completa dopo circa 12 mesi dalla regressione delle lesioni. L'esame RM ha confermato di possedere una elevatissima sensibilità alle ripetizioni secondarie dei linfomi a livello del SNC: il rilievo di una elevata frequenza di tali localizzazioni, comunque in misura inferiore a quanto segnalato nei lavori anatomopatologici, è da mettere in rapporto alla selezione delle forme linfomatose a più elevata malignità, forme nelle quali il contributo dell'esame di risonanza ci appare irrinunciabile, al punto di caldeggiarne l'introduzione nei protocolli diagnostici standard.
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Andreula, F. C., A. M. N. Recchia-Luciani et L. Garofalo. « Linfomi del sistema nervoso centrale e Aids ». Rivista di Neuroradiologia 10, no 2_suppl (octobre 1997) : 206. http://dx.doi.org/10.1177/19714009970100s292.

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I linfomi del sistema nervoso centrale, a lungo eteroplasie intracraniche rare (1–2%) sono in continuo aumento percentuale in relazione con l'immunodepressione virale dell' AIDS (6% dei pazienti, 3% in età pediatrica), così come con quella iatrogena. Tipiche dei linfomi AIDS l'associazione con l'EB virus, l'elevata malignità, la scarsa risposta alla terapia, la localizzazione (SNC, midollo, intestino, cute, anoretto). Oggi tali tumori sono riscontrati in tutte le età (60 anni è la decade di presentazione tipica negli immunocompetenti). Le forme intracraniche, soprattutto B (80%), sono l'1% dei Non-Hodgkin, e dovrebbero essere considerate in realtà secondarie, dal punto di vista fisiopatologico, anche nei casi in cui l'esordio riguardi il SNC. Dal 20 al 40% dei casi sono forme multiple. Il ruolo giocato dall'Imaging deve essere considerato importante, poiché, nonostante le frequenti recidive a breve termine (la sopravvivenza media dalla diagnosi supera di poco l'anno, ed è minore nell'AIDS), queste forme rispondono, quando correttamente inquadrate, assai bene alle alte dosi di cortisonici (nel 40% dei soggetti trattati, già in 24 ore, per linfolisi e ripristino della b.e.e.) così come alla radioterapia. Nella patogenesi sono invocati differenti meccanismi di interconnessione tra neoplasie e agenti virali. La sede preferenziale è sopratentoriale in regione dei nuclei della base o comunque in strutture in cui la componente prevalente è la sostanza bianca. L'estensione dell'edema è incongrua rispetto all'entità della lesione, in ragione della esigua neoangiogenesi indotta. Queste masse hanno margini relativamente ben definiti solo macroscopicamente, con ben maggiore infiltrazione all'istologia; foci di rammollimento necrotico o emorragico sono rari nei pazienti AIDS. All'istologia la zona centrale di cellularità elevata, più rarefatta in periferia, mostra un caratteristico aspetto a “bulbo di cipolla” della trama reticolare. Queste neoplasie si localizzano a livello degli “involucri” cerebrali: sedi caratteristiche sono infatti le leptomeningi e le aree lungo lo spazio subependimal (40–50%), aree di coinvolgimento rese manifeste dalla impregnazione del m.d.c. L'impregnazione lungo le pareti ventricolari suggerisce la diagnosi specie se le immagini RM rivelano l'ulteriore diffusione delle localizzazione leptomeningee lungo gli spazi perivascolari di Virchow Robin. Questi tumori metastatizzano per via ematica, determinando la comparsa di lesioni parenchimali, leptomeningee e meningo-durali. In sede meningo-durale un notevole infiltrato linfomatoso può assumere aspetto a lente biconvessa. Non esistono significative differenze di imaging tra forme linfomatose primitive e secondarie del S.N.C. La TC dimostra lesioni solide singole o multiple, rotondeggianti, isodense al parenchima, (nel 20% dei casi iperdense) con quasi costante accentuazione dopo m.d.c., raramente solo periferica. La RM dimostra isoiperintensità in T1, modesto incremento in DP e ipointensità rispetto alla grigia in T2, da scarso citoplasma delle cellule componenti. L'impregnazione è unicamente da alterazione della barriera emato-encefalica (scarsa la componente neovascolare).
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OASA, Sho. « Report ; Single Protein Dynamics in Cellulo 2014 ». Seibutsu Butsuri 54, no 5 (2014) : 280–82. http://dx.doi.org/10.2142/biophys.54.280.

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O. H. Abdelwahed, O. H. Abdelwahed, et M. El-Sayed Wahed. « Optimizing Single Layer Cellular Neural Network Simulator using Simulated Annealing Technique with Neural Networks ». Indian Journal of Applied Research 3, no 6 (1 octobre 2011) : 91–94. http://dx.doi.org/10.15373/2249555x/june2013/31.

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Miller, W., N. Abrosimov, I. Rasin et D. Borissova. « Cellular growth of single crystals ». Journal of Crystal Growth 310, no 7-9 (avril 2008) : 1405–9. http://dx.doi.org/10.1016/j.jcrysgro.2007.11.046.

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Aonuma, Yuki, Taiji Adachi, Mototsugu Tanaka, Masaki Hojo, Teruko Takano-Yamamoto et Hiroshi Kamioka. « MECHANOSENSITIVITY OF A SINGLE OSTEOCYTE : DIFFERENCE IN CELL PROCESS AND CELL BODY(3A1 Cellular & ; Tissue Engineering & ; Biomaterials I) ». Proceedings of the Asian Pacific Conference on Biomechanics : emerging science and technology in biomechanics 2007.3 (2007) : S165. http://dx.doi.org/10.1299/jsmeapbio.2007.3.s165.

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O. H. Abdelwahed, O. H. Abdelwahed, et M. El-Sayed Wahed. « Optimizing Single-Layer Raster Cellular Neural Network Simulator Using Simulated Annealing Technique and RK4(2), RK4(3) and RK 6(4) ». International Journal of Scientific Research 2, no 6 (1 juin 2012) : 108–12. http://dx.doi.org/10.15373/22778179/june2013/35.

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Ge, Xiaohu, Meidong Huang, Jiaqi Chen, Hui Xu, Jing Xu, Wuxiong Zhang et Yang Yang. « Wireless Single Cellular Coverage Boundary Models ». IEEE Access 4 (2016) : 3569–77. http://dx.doi.org/10.1109/access.2016.2582960.

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Lee, J., J. Y. Sul et J. H. Eberwine. « Single Cell/Cellular Subregion-Targeted Phototransfection ». Cold Spring Harbor Protocols 2014, no 9 (1 septembre 2014) : pdb.prot072421. http://dx.doi.org/10.1101/pdb.prot072421.

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Leake, M. C. « Analytical tools for single-molecule fluorescence imaging in cellulo ». Phys. Chem. Chem. Phys. 16, no 25 (2014) : 12635–47. http://dx.doi.org/10.1039/c4cp00219a.

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Thèses sur le sujet "Singole cellule"

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Antoniolli, Francesca. « PROGETTAZIONE E CARATTERIZZAZIONE DI UN BIOSENSORE MEMS ». Doctoral thesis, Università degli studi di Trieste, 2008. http://hdl.handle.net/10077/2755.

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2003/2004
Negli ultimi anni, le cellule sono state oggetto di studio approfondito e, in taluni casi, di esperimenti molto sofisticati. Tuttavia, benché si conosca molto circa la loro struttura, poche sono le informazioni sulla meccanica cellulare e sulla risposta cellulare agli stimoli meccanici. Le cellule, infatti, possono sentire forze meccaniche e convertirle in risposte biologiche, oppure, viceversa, è da tempo noto come segnali biologici e biochimici influenzino l’abilità cellulare nel sentire, generare e sopportare forze di tipo meccanico. Negli ultimi anni sono stati ideati e realizzati svariati meccanismi per l’applicazione di forze meccaniche su cellule e la rilevazione delle conseguenti deformazioni. Questi sistemi, però, presentano dei limiti: - la forza esercitata non è adeguata al fenomeno investigato; - lo studio viene effettuato su un’intera popolazione di cellule; - la forza è esercitata localmente e non sull’intera cellula. Il presente lavoro di tesi, avente come obiettivo primo lo sviluppo, la progettazione e la realizzazione di un dispositivo per la sollecitazione meccanica della singola cellula e la rilevazione delle conseguenti deformazioni, si è focalizzato sullo studio di dispositivi che potessero bypassare i suddetti limiti. La scelta è ricaduta nei Sistemi Micro Elettro Meccanici, dal momento che, oltre ad avere dimensioni compatibili con le caratteristiche cellulari ed assicurare modesti costi realizzativi ed operativi, garantiscono - la possibilità di applicare forze in un ampio range (pN-µN); - la possibilità di effettuare studi sulla singola cellula, ed in particolare su cellule aderenti; - la possibilità di stimolare l’intera cellula, e non soltanto una porzione locale di questa. La prima parte del lavoro è stata rivolta alla messa a punto di dispositivi che, concepiti in maniera analoga a quelle che sono le tradizionali macchine universali per test meccanici, potessero consentire l’ancoraggio della singola cellula su di una piattaforma di geometrie differenti a seconda che si volesse applicare una sollecitazione di trazione uniassiale, biassiale, pluriassiale oppure di taglio. Tali dispositivi tuttavia hanno riscontrato diverse problematiche quando operanti in soluzioni saline quali i medium cellulari. Sono stai quindi concepiti e sviluppati dei nuovi dispositivi che potessero bypassare le problematiche riscontrate con i primi: il MEMS è stato quindi sdoppiato su due outline di 2x2 mm, di cui una ospitante il motore per l’attuazione del dispositivo operante in aria l’altra ospitante la piattaforma per la collocazione della cellula in esame. Per completare il funzionamento di tali dispositivi è stata sviluppata e realizzata con successo una tecnica di collegamento di questi mediante una fibra di carbonio ancorata ai MEMS mediante wire bonding. Infine sono state acquisite e messe a punto la strumentazione e le tecniche che potessero consentire di operare con cellule viventi: è stato individuato un materiale tale da consentire un ancoraggio ottimale della cellula e con il quale si potesse funzionalizzate localmente la piattaforma per la cellula; è stato allestito un laboratorio per colture cellulari presso il Dipartimento dei Materiali e delle Risorse Naturali; è stata messa a punto una tecnica per la manipolazione di singole cellule; sono state infine eseguite alcune preliminari prove di trazione sulla singola cellula.
XX Ciclo
1979
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Moussy, Alice. « Caractérisation des premières étapes de différenciation des cellules hématopoïétiques à l'échelle de la cellule unique ». Thesis, Paris Sciences et Lettres (ComUE), 2017. http://www.theses.fr/2017PSLEP029/document.

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Bien que largement étudiés, les mécanismes fondamentaux de prise de décision dans les processus de différenciation cellulaire restent mal compris. Les théories déterministes, souvent basées sur des études populationnelles, atteignent rapidement leur limite lorsqu’il s’agit d’expliquer les différences de choix individuels de cellules, pourtant exposées au même environnement. L’objectif de ma thèse est donc d’étudier les premières étapes de la différenciation des cellules hématopoïétiques à l’échelle de la cellule unique, par des analyses transcriptomiques, protéomiques et morphologiques. Ce travail a été effectué sur deux modèles de différenciation : les lymphocytes T régulateurs et les cellules CD34+ humaines issues de sang de cordon. Nous avons observé le comportement de ces cellules uniques après stimulation. Grâce à la combinaison de la microscopie en time lapse et des analyses moléculaires réalisées à l’échelle de la cellule individuelle, nous avons pu démontrer que le choix du devenir cellulaire n’était pas unique, programmé. La cellule passe d’abord par un état dit « multi-primed », métastable où elle exprime des gènes de plusieurs lignées différentes, puis elle passe par une phase dite « incertaine », instable où elle hésite entre deux phénotypes avant de se stabiliser dans un état fixe. Nos observations sont cohérentes avec une explication stochastique de la prise de décision. La différenciation serait donc un processus spontané, dynamique, fluctuant et non un processus prédéterminé. Les décisions du destin cellulaire sont prises séparément par les cellules individuelles
Despite intensively studies, the fundamental mechanisms of cell fate decision during cellular differentiation still remain unclear. The deterministic mechanisms, often based on studies of large cell populations, cannot explain the difference between individual cell fates choices placed in the same environment. The aim of my thesis work is to study the first steps of hematopoietic cell differentiation at the single cell level thanks to transcriptomic, proteomic and morphological analyses. Two differentiation models have been used: T regulatory lymphocytes and human cord blood-derived CD34+ cells. The behavior of individual cells following stimulation has been analyzed. Using time-lapse microscopy coupled to single cell molecular analyses, we could demonstrate that the cell fate choice is not a unique, programmed event. First, the cell reaches a metastable “multi-primed” state, which is characterized by a mixed lineage gene expression pattern. After transition through an “uncertain”, unstable state, characterized by fluctuations between two phenotypes, the cell reaches a stable state. Our observations are coherent with a stochastic model of cell fate decision. The differentiation is likely to be a spontaneous, dynamic, fluctuating and not a deterministic process. The cell fate decisions are taken by individual cells
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MALLIA, SELENE. « La genomica su singola cellula rivela la gerarchia e l'architettura clonale nelle Neoplasie Mieloproliferative ». Doctoral thesis, Università degli studi di Modena e Reggio Emilia, 2022. http://hdl.handle.net/11380/1278821.

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Le Neoplasie Mieloproliferative (MPN) sono disordini ematologici caratterizzati dalla presenza di mutazioni somatiche che colpiscono le cellule staminali ematopoietiche e comprendono la Policitemia Vera, la Trombocitemia Essenziale e la Mielofibrosi Primaria (PMF). La PMF è una neoplasia eterogenea, contraddistinta dalla presenza di fibrosi midollare, iperplasia megacariocitaria e ematopoiesi extramidollare, e mostra la peggiore prognosi tra tutte le MPN. I pazienti spesso non rispondono ai trattamenti e nel 15-20% dei casi sviluppano una Leucemia Mieloide Acuta (LMA). Le mutazioni ricorrenti, conosciute come “mutazioni drivers”, interessano i geni JAK2, CALR e MPL, ma a complicare il profilo mutazionale intervengono altre alterazioni che sono spesso responsabili del peggioramento del quadro clinico e della trasformazione leucemica. La progressione della malattia e l’evoluzione leucemica nella PMF è accompagnata da un aumento della complessità genomica e dell’eterogeneità clonale. Molti studi hanno confermato come l’ordine di acquisizione delle mutazioni influenzi il decorso clinico. Tuttavia sono ancora poco conosciute le caratteristiche dei cloni che determinano la malattia e che guidano la trasformazione leucemica. Studi recenti hanno dimostrato come la genomica su singola cellula sia una tecnica sensibile per studiare l’eterogeneità clonale e l’evoluzione delle leucemie. Per questa ragione, abbiamo adottato un approccio di genomica su singola cellula per risolvere la complessità clonare della PMF. Dapprima abbiamo sviluppato un metodo di isolamento delle cellule staminali e progenitori ematopoietici CD34+ dal sangue cordonale, di fissazione e marcatura del CD34, al fine di ottenere una popolazione cellulare adatta alla separazione in singole cellule, sfruttando il sistema del DEP-array (Menarini Silicon Biosystem). In seguito, abbiamo confrontato diversi protocolli di amplificazione dell’intero genoma su singola cellula al fine di ottenere un’amplificazione omogenea, minimizzando l’effetto di allele drop out, per proseguire con il sequenziamento Sanger. Usando questa procedura, abbiamo analizzato le cellule CD34+ di un paziente affetto da PMF, positivo per la mutazione JAK2V617F e per altre alterazioni genetiche, caratteristiche delle MPN. Il paziente, nonostante il trattamento con il JAK2-inibitore Ruxolitinib, ha sviluppato una LMA. Al fine di ricostruire la gerarchia e l’architettura clonale, abbiamo analizzato le cellule CD34+ alla diagnosi (T1), durante la fase accelerata (T2) e nella fase di LMA (T3). Grazie alla analisi su singola cellula, abbiamo stabilito che il primo evento mutazionale investa TET2, precedendo la mutazione di JAK2, e probabilmente influenzando negativamente la risposta alla terapia. Abbiamo osservato, inoltre, un aumento dei cloni mutati per TP53 durante la progressione della malattia, suggerendo che siano stati questi cloni a supportare la fase T2. Inaspettatamente, già nella fase T1, abbiamo riscontrato una piccola popolazione cellulare recante una mutazione pro-leucemica su FLT3, alterazione che non era stata evidenziata dall'analisi in NGS ma che verosimilmente ha guidato lo sviluppo della fase T3. Infine, abbiamo evidenziato una mutazione omozigote su SRSF2 non ancora descritta. Tutti i nostri dati, confermano quindi come la genomica su singola cellula sia una tecnologia promettente di analisi della eterogeneità clonale delle MPN e che permetta sia di evidenziare precocemente caratteristiche leucemiche sia di ottenere un quadro chiaro degli eventi mutazioni che interessano i disordini ematologici.
Somatic mutations in Hematopoietic Stem Cells (HSCs) cause Myeloproliferative Neoplasms (MPNs), including Polycythemia Vera, Essential Thrombocythemia and Primary Myelofibrosis (PMF). PMF is a heterogeneous disorder consisting of bone marrow fibrosis, megakaryocyte hyperplasia and extramedullary hematopoiesis and is characterized by the worst prognosis among MPNs. About 15-20% of patients are unresponsive to conventional therapies and develop Acute Myeloid Leukemia (AML). In HSCs the main mutations, identified as “driver mutations” during MPNs pathogenesis, involve JAK2, CALR and MPL genes; in addition, many other genetic alterations contribute to the prognosis worsening and the development of AML. Disease progression and leukemic evolution in PMF results from an increase of the genomic complexity and clonal heterogeneity. Many studies confirmed that the mutational acquisition order affects the clinical outcome. However, the clonal architecture determining disease evolution and the clones guiding leukemic transformation are poorly understood. Recent studies demonstrate that single-cell (sc) genomics is a sensitive technique suitable to study clonal heterogeneity and to detect the evolution of the malignant cells in hematological neoplasms. For this reason, we used the sc-genomics approach to clarify the clonal complexity in PMF. Firstly, we developed a workflow for CD34+ Hematopoietic Stem Progenitor Cells (HSPCs) isolation from cord blood, fixation and immunostaining for CD34, in order to singularly separate the cells by DEP-array system (Menarini Silicon Biosystem) and to obtain a cell population suitable for sc-analysis. Then, we compared different whole genome amplification (WGA) protocols for single cells in order to obtain a uniform DNA amplification for Sanger sequencing and minimize allele drop out effect. Based on this method, we analyzed the CD34+ HSPCs of a PMF patient carrying JAK2V617F and other MPN frequent mutations. This patient was treated with JAK2-inhibitor Ruxolitinib but he was unresponsive to therapy and evolved to AML. In order to reconstruct the clonal hierarchy and architecture, we analyzed CD34+ cells during chronic phase (T1), the accelerated phase (T2) and the AML phase (T3). By means to sc-analysis, we established that TET2 was the first mutated gene, preceding JAK2 mutation, and this probably conferred a lower sensitivity to treatment. Moreover, we identified an increase of the allele burden of the TP53 mutation during disease progression, suggesting that TP53-mutated clones supported the accelerated (T2) phase. Interestingly, we already detected in T1 phase a small cell fraction, undetectable by bulk NGS and carrying the leukemogenic FLT3 mutation, probably driving the T3 phase. Finally, we characterized SRSF2 homozygous mutation that has not been described yet. Altogether our data demonstrate that sc-genomics is a promising method to uncover clonal heterogeneity in MPNs, highlighting the early occurrence of pro-leukemic mutations and to describe the real scenario of mutational events in hematological diseases.
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Caccianini, Laura. « Imagerie de l'architecture dynamique de la chromatine dans la cellule unique ». Thesis, Paris Sciences et Lettres (ComUE), 2019. https://tel.archives-ouvertes.fr/tel-02896692.

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La structure de la chromatine joue un rôle crucial dans la régulation de plusieurs fonctions cellulaires chez les cellules de mammifères. Perturber l’organisation spatiale de la chromatine peut avoir des conséquences dramatiques sur la vie d’une cellule et peut amener`des pathologies graves chez les organismes. Deux facteurs nucléaires, CTCF et Cohesine, sont parmi les principaux acteurs dans la régulation et le maintien de l’architecture de l’ADN. Des avancements importants ont révélé ́la complexité ́des mécanismes qui régulent l’organisation de la chromatine, mais le domaine manque encore d’une description dynamique à l’échelle de la cellule et de la molécule unique. Cette étude est centrée sur la description de la dynamique de CTCF et Cohesin réalisé ́avec de méthodes de suivi de la molécule unique dans des cellules souche embryonnaires vivantes de souris. L’interaction entre ces deux facteurs a été étudiée à travers la caractérisation de la dynamique de Cohesin en absence de CTCF et dans le contexte d’autres altérations biologiques
Chromatin structure and cellular function are tightly linked in the nucleus of mammalian cells. Disruption of chromatin spatial organisation dramatically affects the life of a cell and eventually leads to severe pathologies in entire organisms. Two nuclear factors, CTCF and Cohesin, have been found to play a crucial role in the regulation and maintenance of DNA architecture. Huge advancements have been made in the understanding of the mechanisms behind chromatin arrangement but the field is still lacking a dynamic picture at the single cell and single molecule level. This study provide this study provides insight into the dynamics of CTCF and Cohesin through single particle tracking of CTCF and Cohesin dynamics achieved with single molecule tracking in living mouse embryonic stem cells. The interplay between these two factors was studied by looking at Cohesin’s behaviour in the absence of CTCF and in the context of other biological alterations
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Holt, Brian D. « Cellular Processing of Single Wall Carbon Nanotubes ». Research Showcase @ CMU, 2014. http://repository.cmu.edu/dissertations/397.

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Nanostructured materials are hailed to be the solutions of the future for many research areas, and single wall carbon nanotubes (SWCNTs) are one of the more interesting materials due to their highly desirable electronic, optical, thermal and mechanical properties. For instance, this combination of properties is of wide interest for biological applications, including cellular technologies. However, understanding cellular processing of SWCNTs is limited. In this thesis, quantification of sub-cellular events–including SWCNT uptake rates, altered mitosis, redistribution of sub-cellular components and reduced cellular functionalities–is used to formulate insight into how cells internalize and process SWCNTs. By understanding sub-cellular processing of SWCNTs, new basic science endeavors and SWCNT-based biological applications can be more effectively developed, and the insights can be generalized to other nanostructured materials.
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Geisler, Hubert. « Structuration d'hydrogels thermoactivables pour l'analyse de cellules uniques ». Electronic Thesis or Diss., Université Paris sciences et lettres, 2020. http://www.theses.fr/2020UPSLS001.

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Dans cette thèse est présentée une nouvelle technologie microfluidique de capture de cellules uniques basée sur l’utilisation d’hydrogels thermoactivables. Nous utilisons notamment le PolyNIPAM, un polymère dont le volume est augmenté dans l'eau de 400% lorsque la température est inférieure à 32°C et est dégonflé lorsque la température est supérieure à 34°C. Nous exploitons ce gonflement réversible pour ouvrir et fermer des compartiments intégrés dans une chambre microfluidique.Le greffage et la structuration de ces motifs d’hydrogel repose sur la chimie click thiol-ène, initiée par voie thermique ou par irradiation UV. Nous avons développé des méthodes et procédés de microfabrication dans le but de diversifier les substrats d’accroche (du verre vers le PDMS, COC, PMMA, etc), d’élargir la gamme des épaisseurs des structures réalisables (de quelques microns vers la dizaine de microns d’épaisseur) et de renforcer nos connaissances concernant l'incidence de la fabrication sur le comportement de l’hydrogel. Un protocole de photolithographie robuste est finalement obtenu permettant le design de toute sorte de motif 2D sur différents choix de substrat. Une application possible détaillée par la suite est le développement de ces puces microfluidiques capables de capturer des cellules uniques dans des compartiments en hydrogel. (confidentiel)
We present in this work a new microfluidic technology aiming at isolating single cells by the use of thermoactuable polymers. One of the polymers we use is polyNIPAM, a polymer that can expand its volume by 400% in water when the temperature is set under 32°C and can shrink down when it is set over 34°C. We use this reversible swelling capability to open and close compartments embedded in a microfluidic chip.Grafting and structuring these hydrogel features relies on thiol-en click chemistry, initiated thermally or by UV irradiation. We have developed methods and microfabrication protocols in order to diversify the substrate materials (from glass to PDMS, COC, PMMA, etc), to expand the structures thickness range (from few microns to a tenth of microns) and to strengthen our knowledge regarding the fabrication impact on the hydrogel’s behavior. A robust protocol of photolithography has finally been worked on allowing the design of any type of 2D features on a large choice of substrates.One of the realistic applications detailed here is the development of microfluidic chips aiming at isolating single cells in hydrogel compartments. (confidential)
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Boltyanskiy, Rostislav. « Mechanical Response of Single Cells to Stretch ». Thesis, Yale University, 2016. http://pqdtopen.proquest.com/#viewpdf?dispub=10160860.

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A living cell is a complex soft matter system far from equilibrium. While its components have definite mechanical properties such as stiffness and viscosity, cells consume energy to generate force and exhibit adaptation by modulating their mechanical properties through regulatory pathways. In this dissertation, we explore cell mechanics by stretching single fibroblast cells and simultaneously measuring their traction stresses. Upon stretch, there is a sudden, drastic increase in traction stresses, often followed by a relaxation over a time scale of about 1 minute. Upon release of stretch, traction stresses initially drop and often recover on a similar time scale of about 1 minute. We show that a minimal active linear viscoelastic model captures essential features of cell response to stretch. This model is most successful in describing the response of cells within the first 30 seconds of stretch. While perturbations of myosin and vinculin change quiescent traction stresses, they surprisingly have no significant impact on the stiffness or viscoelastic timescale of the cells. On longer time scales, cells may show an adaptive response to stretch that contradicts the minimal mechanical model. The probability of an adaptive response is significantly reduced by myosin de-activation and vinculin knockout. Therefore, we find that while vinculin and myosin are not important in determining passive mechanical properties of cells, such as stiffness and viscosity, they play a significant role in the adaptive mechanisms of cell response to stretch. To perform this work, we have built a novel micro stretching device compatible with live cell microscopy and developed a computational tool to analyze data from large traction stresses. Therefore, this dissertation's contribution is two-fold: (1) a novel experimental approach to explore the mechanics of living cells, and (2) a new model and framework for understanding the mechanical response of cells to stretch.

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Pagliaro, Sarah Beatriz De Oliveira. « Transcriptional control induced by bcr-abl and its role in leukemic stem cell heterogeneity. Single-Cell Transcriptome in Chronic Myeloid Leukemia : Pseudotime Analysis Reveals Evidence of Embryonic and Transitional Stem Cell States Single Cell Transcriptome in Chronic Myeloid Leukemia (CML) : Pseudotime Analysis Reveals a Rare Population with Embryonic Stem Cell Features and Druggable Intricated Transitional Stem Cell States A novel neuronal organoid model mimicking glioblastoma (GBM) features from induced pluripotent stem cells (iPSC) Experimental and integrative analyses identify an ETS1 network downstream of BCR-ABL in chronic myeloid leukemia (CML) ». Thesis, université Paris-Saclay, 2020. http://www.theses.fr/2020UPASQ032.

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La leucémie myéloïde chronique est une hématopoïèse maligne clonale, caractérisée par l'acquisition de la translocation t (9;22) conduisant au chromosome Ph1 et à son homologue l'oncogène BCR-ABL, dans une cellule souche hématopoïétique très primitive. La LMC est un modèle de thérapies ciblées, car il a été démontré que la preuve de la faisabilité du ciblage de l'activité tyrosine kinase (TK) BCR-ABL à l'aide d'inhibiteurs de TK (TKI) entraîne des réponses et des rémissions majeures. Cependant, les problèmes actuels rencontrés dans ces thérapies sont la résistance des cellules souches leucémiques primitives et leur persistance qui serait liée à l'hétérogénéité des cellules souches au moment du diagnostic, ce qui conduit à la sélection clonale de cellules résistant aux thérapies TKI. J'ai appliqué la technologie de l'analyse du transcriptome des cellule uniques aux cellules de la LMC en utilisant un panel de gènes impliqués dans différentes voies, combinée à l'analyse d'inférence de trajectoire au modèle d'expression des gènes. Les résultats ont montré un état transitoire des cellules souches comprenant des gènes embryonnaires identifiés dans les cellules de la LMC au moment du diagnostic, ce qui pourrait contribuer à la résistance et à la persistance de la LSC. En outre, l'oncoprotéine Bcr-Abl est la tyrosine kinase constitutivement active produite par le gène chimérique BCR-ABL dans la leucémie myéloïde chronique (LMC). Les cibles transcriptionnelles de Bcr-Abl dans les cellules leucémiques n'ont pas été étudiées de manière approfondie. Une expérience de transcriptome utilisant la lignée cellulaire UT7 hématopoïétique exprimant BCR-ABL, a identifié la surexpression du facteur d'élongation eucaryote kinase 2 (eEF2K) qui joue un rôle majeur dans la survie des cellules en cas de privation de nutriments. Dans l'ensemble, les données suggèrent que la surexpression de eEF2K dans la LMC est associée à une sensibilité accrue à la privation de nutriments
Chronic myeloid leukemia is a clonal hematopoietic malignancy, characterized by the acquisition of the t (9;22) translocation leading to Ph1 chromosome and its counterpart BCR-ABL oncogene, in a very primitive hematopoietic stem cell. CML is a model of targeted therapies as the proof of concept of the feasibility of targeting the tyrosine kinase (TK) activity BCR-ABL using TK inhibitors (TKI) has been shown to lead to major responses and remissions. However, the current problems encountered in these therapies are primitive leukemic stem cells resistance and their persistence which is thought to be related to the heterogeneity of the stem cells at diagnosis leading to clonal selection of cells resisting to TKI therapies. I have applied the technology of single cell transcriptome analysis to CML cells using a panel of genes involved in different pathways combined with trajectory inference analysis to the gene expression pattern. The results showed a transitional stem cell states including embryonic genes identified in CML cells at diagnosis which could contribute to LSC resistance and persistence. Furthermore, the oncoprotein Bcr-Abl is the constitutively active tyrosine kinase produced by the chimeric BCR-ABL gene in chronic myeloid leukemia (CML). The transcriptional targets of Bcr-Abl in leukemic cells have not been extensively studied. A transcriptome experiment using the hematopoietic UT7 cell line expressing BCR-ABL, has identified the overexpression of eukaryotic elongation factor kinase 2 (eEF2K) which plays a major role in the survival of cells upon nutrient deprivation. Overall, the data suggest that overexpression of eEF2K in CML is associated with an increased sensitivity to nutrient-deprivation
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Simon-Desbois, Linda. « Development of a microfluidic device for single cell transcriptome analysis ». Thesis, Lille 2, 2013. http://www.theses.fr/2013LIL2S007.

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L’hétérogénéité cellulaire au sein d’une même population cellulaire a été observée chez des organismes procaryotes, chez des organismes plus complexes tels que les mammifères et chez les cellules cancéreuses. L’expression des gènes est un phénomène stochastique ou « bruité ». La connaissance, à l’échelle d’une cellule isolée, des variations stochastiques de l’expression génique, ses oscillations dans le temps et en fonction des fluctuations extérieures constitue un enjeu majeur. Elle permettrait de connaitre les mécanismes de croissance, de différenciation et de migration cellulaire et par voie de conséquence de développer de nouveaux outils thérapeutiques. Par ailleurs, les techniques actuelles d’analyses sur cellule unique nécessitent des protocoles longs et laborieux. Cependant, l’avènement des microcircuits dans les années 80, puis celle de la microfluidique dans les années qui ont suivi, ont ouvert les portes des analyses (chimiques, biologiques et biochimiques) à haut débit, sur de très petits volumes (pl à fl). Nous avons développé un microsystème fluidique, permettant la génération de micro gouttelettes ultrastables et facilement manipulables ; et ce, en utilisant les qualités de changement de phase de l’agarose. Par ailleurs, nous avons conçu un system innovant de génération de microgouttelettes le système « push-pull » qui permet de réduire les volumes morts. Le très petit volume des microgouttelettes font d’elles des microréacteurs dans lesquels nous avons pu encapsuler des suspensions d’ADN, puis des cellules isolées, réaliser des réactions biochimiques, et analyser leur transcriptome ; et améliorer encore l’efficacité et l’intérêt des microémulsions, une technique à très haut débit, et en plein essor.L’hétérogénéité cellulaire au sein d’une même population cellulaire a été observée chez des organismes procaryotes, chez des organismes plus complexes tels que les mammifères et chez les cellules cancéreuses. L’expression des gènes est un phénomène stochastique ou « bruité ». La connaissance, à l’échelle d’une cellule isolée, des variations stochastiques de l’expression génique, ses oscillations dans le temps et en fonction des fluctuations extérieures constitue un enjeu majeur. Elle permettrait de connaitre les mécanismes de croissance, de différenciation et de migration cellulaire et par voie de conséquence de développer de nouveaux outils thérapeutiques. Par ailleurs, les techniques actuelles d’analyses sur cellule unique nécessitent des protocoles longs et laborieux. Cependant, l’avènement des microcircuits dans les années 80, puis celle de la microfluidique dans les années qui ont suivi, ont ouvert les portes des analyses (chimiques, biologiques et biochimiques) à haut débit, sur de très petits volumes (pl à fl). Nous avons développé un microsystème fluidique, permettant la génération de micro gouttelettes ultrastables et facilement manipulables ; et ce, en utilisant les qualités de changement de phase de l’agarose. Par ailleurs, nous avons conçu un system innovant de génération de microgouttelettes le système « push-pull » qui permet de réduire les volumes morts. Le très petit volume des microgouttelettes font d’elles des microréacteurs dans lesquels nous avons pu encapsuler des suspensions d’ADN, puis des cellules isolées, réaliser des réactions biochimiques, et analyser leur transcriptome ; et améliorer encore l’efficacité et l’intérêt des microémulsions, une technique à très haut débit, et en plein essor
In the post-genomic era, it is now critical to characterize living organisms at the singlecell level. CAGE (Cap Analysis of Gene Expression) is a technology developed by agroup of RIKEN instituteto get genome-wide profile of gene expression. It can beused for profiling of gene expression and identifying the TSS (transcription start site)to analyze promoters architecture. By using the CAGE technology, it could be foundthat different tissues and families of genes differentially use distinct types ofpromoters. Applying CAGE technology against single cells is an ideal way tounderstand life phenomenon based on genome and will have a major impact inbiology. To address this, a novel platform to manipulate single cell and analyze itsown transcriptome with higher precision and efficiency is required.This project aims to develop a microfluidic platform to realize the protocol of CAGEtechnology against single cells with higher-throughput and sensitivity overconventional microtube-based way. For this, we encapsulated single cells inmicrodroplets, lysed them, and performed RT reaction in order to sequence andanalyze their transcriptome
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Chen, Peng. « Single cell assays of exocytosis / ». free to MU campus, to others for purchase, 2002. http://wwwlib.umi.com/cr/mo/fullcit?p3074384.

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Livres sur le sujet "Singole cellule"

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Single-molecule cellular biophysics. Cambridge : Cambridge University Press, 2012.

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Chemical cytometry : Ultrasensitive analysis of single cells. Weinheim : Wiley-VCH, 2010.

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Sako, Yasushi. Cell Signaling Reactions : Single-Molecular Kinetic Analysis. Dordrecht : Springer Science+Business Media B.V., 2011.

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Sardin, Marie-Neige. Celle qui dit non. Paris : Œuvre, 2011.

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1961-, Duijn Bert van, et Wiltink Anneke 1961-, dir. Signal transduction--single cell techniques. Berlin : Springer, 1998.

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The diversity of life : From single cells to multicellular organizations. Oxford : Heinemann Library, 2003.

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Snedden, Robert. The diversity of life : From single cells to multicellular organisms. Chicago, Ill : Heinemann Library, 2008.

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Arnaud, Chauvière, Preziosi Luigi et Verdier Claude 1962-, dir. Cell mechanics : From single scale-based models to multiscale modeling. Boca Raton : Chapman & Hall/CRC, 2009.

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Arnaud, Chauvière, Preziosi Luigi et Verdier Claude, dir. Cell mechanics : From single scale-based models to multiscale modeling. Boca Raton : Chapman & Hall/CRC, 2009.

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Luigi, Preziosi, et Verdier Claude, dir. Cell mechanics : From single scale-based models to multiscale modeling. Boca Raton : Chapman & Hall/CRC, 2009.

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Chapitres de livres sur le sujet "Singole cellule"

1

De Keijzer, Sandra, B. Ewa Snaar-Jagalska, Herman P. Spaink et Thomas Schmidt. « Single-Molecule Imaging of Cellular Signaling ». Dans Single Molecules and Nanotechnology, 107–29. Berlin, Heidelberg : Springer Berlin Heidelberg, 2008. http://dx.doi.org/10.1007/978-3-540-73924-1_5.

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Huang, Howard, Constantinos B. Papadias et Sivarama Venkatesan. « Single-user MIMO ». Dans MIMO Communication for Cellular Networks, 35–78. Boston, MA : Springer US, 2011. http://dx.doi.org/10.1007/978-0-387-77523-4_2.

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Shinde, Ashwini, Srabani Kar, Moeto Nagai, Fan-Gang Tseng et Tuhin Subhra Santra. « Light-Induced Cellular Delivery and Analysis ». Dans Handbook of Single Cell Technologies, 1–29. Singapore : Springer Singapore, 2021. http://dx.doi.org/10.1007/978-981-10-4857-9_4-1.

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Shinde, Ashwini, Srabani Kar, Moeto Nagai, Fan-Gang Tseng et Tuhin Subhra Santra. « Light-Induced Cellular Delivery and Analysis ». Dans Handbook of Single-Cell Technologies, 3–30. Singapore : Springer Singapore, 2021. http://dx.doi.org/10.1007/978-981-10-8953-4_4.

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Patsch, Katherin, Shannon M. Mumenthaler et Daniel Ruderman. « Image-Based Tracking of Heterogeneous Single-Cell Phenotypes ». Dans Cellular Heterogeneity, 47–63. New York, NY : Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7680-5_3.

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Navas-Moreno, Maria, et James W. Chan. « Laser Tweezers Raman Microspectroscopy of Single Cells and Biological Particles ». Dans Cellular Heterogeneity, 219–57. New York, NY : Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7680-5_13.

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Kwon, Jungeun Sarah, Xia Wang et Guang Yao. « Study Quiescence Heterogeneity by Coupling Single-Cell Measurements and Computer Modeling ». Dans Cellular Quiescence, 287–99. New York, NY : Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7371-2_20.

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Sanchez-Adams, Johannah, et Kyriacos A. Athanasiou. « Biomechanical Characterization of Single Chondrocytes ». Dans Cellular and Biomolecular Mechanics and Mechanobiology, 247–66. Berlin, Heidelberg : Springer Berlin Heidelberg, 2010. http://dx.doi.org/10.1007/8415_2010_20.

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Brun, Caroline E., Yu Xin Wang et Michael A. Rudnicki. « Single EDL Myofiber Isolation for Analyses of Quiescent and Activated Muscle Stem Cells ». Dans Cellular Quiescence, 149–59. New York, NY : Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7371-2_11.

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Barteneva, Natasha S., et Ivan A. Vorobjev. « Heterogeneity of Metazoan Cells and Beyond : To Integrative Analysis of Cellular Populations at Single-Cell Level ». Dans Cellular Heterogeneity, 3–23. New York, NY : Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7680-5_1.

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Actes de conférences sur le sujet "Singole cellule"

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Liu, Yu-San, Yonghua Luo, John O. Baker, Yining Zeng, Michael E. Himmel, Steve Smith et Shi-You Ding. « A single molecule study of cellulase hydrolysis of crystalline cellulose ». Dans BiOS, sous la direction de Jörg Enderlein, Zygmunt K. Gryczynski et Rainer Erdmann. SPIE, 2010. http://dx.doi.org/10.1117/12.840975.

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Coskun, Ulas C., Matthew L. Ferguson, Alexander Vallmitjana, Huynh Anh, Julianna Goelzer, Yuansheng Sun, Shih-Chu J. Liao, Sunil Shah, Enrico Gratton et Beniamino Barbieri. « Nano-resolution in vivo 3D orbital tracking system to study cellular dynamics and bio-molecular processes ». Dans Single Molecule Spectroscopy and Superresolution Imaging XIII, sous la direction de Ingo Gregor, Rainer Erdmann et Felix Koberling. SPIE, 2020. http://dx.doi.org/10.1117/12.2546690.

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Mandyam, Giridhar D. « Single-Sideband OFDM for Cellular Systems ». Dans 2006 Fortieth Asilomar Conference on Signals, Systems and Computers. IEEE, 2006. http://dx.doi.org/10.1109/acssc.2006.355064.

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Chen, Fei. « Abstract IA02 : Slide-seq : A platform for understanding cellular circuits in tissue ». Dans Abstracts : AACR Virtual Special Conference on Tumor Heterogeneity : From Single Cells to Clinical Impact ; September 17-18, 2020. American Association for Cancer Research, 2020. http://dx.doi.org/10.1158/1538-7445.tumhet2020-ia02.

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Wu, Catherine J. « Abstract IA11 : Detecting and targeting cellular heterogeneity of the lymphoid blood malignancies ». Dans Abstracts : AACR Virtual Special Conference on Tumor Heterogeneity : From Single Cells to Clinical Impact ; September 17-18, 2020. American Association for Cancer Research, 2020. http://dx.doi.org/10.1158/1538-7445.tumhet2020-ia11.

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Ashlock, Daniel, et Sharon McNicholas. « Single parent generalization of cellular automata rules ». Dans 2012 IEEE Congress on Evolutionary Computation (CEC). IEEE, 2012. http://dx.doi.org/10.1109/cec.2012.6256126.

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Kaufmann, Rainer, Paul Lemmer, Manuel Gunkel, Yanina Weiland, Patrick Müller, Michael Hausmann, David Baddeley, Roman Amberger et Christoph Cremer. « SPDM : single molecule superresolution of cellular nanostructures ». Dans SPIE BiOS : Biomedical Optics, sous la direction de Jörg Enderlein, Zygmunt K. Gryczynski et Rainer Erdmann. SPIE, 2009. http://dx.doi.org/10.1117/12.809109.

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Lin, C. M., C. C. Tseng, C. L. Chen et Andrew M. Wo. « Hydrodynamic single-cell trapping for cellular assays ». Dans TRANSDUCERS 2009 - 2009 International Solid-State Sensors, Actuators and Microsystems Conference. IEEE, 2009. http://dx.doi.org/10.1109/sensor.2009.5285399.

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Babaee, Sahab, Babak Haghpanah Jahromi, Amin Ajdari, Hamid Nayeb-Hashemi et Ashkan Vaziri. « Mechanical Properties of Open-Cell Cellular Structures With Rhombic Dodecahedron Cells ». Dans ASME 2010 International Mechanical Engineering Congress and Exposition. ASMEDC, 2010. http://dx.doi.org/10.1115/imece2010-39924.

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We present a series of analytical models and finite element results (FE) for special 3-D open cellular foam to determine the effective material properties of a 3D rhombic dedecahedron open-cell cellular structure. The analytical approach is based on minimizing the total energy associated with small deformation of a single unit cell of the cellular structure. The finite element models were developed for both a single unit cell and three dimensional foam structure and used to obtain the mechanical properties in all three principal directions.
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Dahan, Maxime. « Probing Cellular Events with Single Quantum Dot Imaging ». Dans Laser Science. Washington, D.C. : OSA, 2009. http://dx.doi.org/10.1364/ls.2009.lsthb4.

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Rapports d'organisations sur le sujet "Singole cellule"

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Harmon, Jennifer. Cotton Versus Bacterial Cellulose : A Comparison of Single Ply Yarns. Ames (Iowa) : Iowa State University. Library, janvier 2019. http://dx.doi.org/10.31274/itaa.8789.

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Hiddessen, A. SINGLE-CELL LEVEL INVESTIGATION OF CYTOSKELETAL/CELLULAR RESPONSE TO EXTERNAL STIMULI. Office of Scientific and Technical Information (OSTI), février 2007. http://dx.doi.org/10.2172/902301.

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Sun, Lina, Yanan Han, Hua Wang, Huanyu Liu, Shan Liu, Hongbin Yang, Xiaoxia Ren et Ying Fang. MicroRNAs as Potential Biomarkers for the Diagnosis of Inflammatory Bowel Disease : A Systematic Review and Meta-analysis. INPLASY - International Platform of Registered Systematic Review and Meta-analysis Protocols, février 2022. http://dx.doi.org/10.37766/inplasy2022.2.0027.

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Review question / Objective: The purpose of this systematic review was to systematically review the clinical studies regarding miRNAs as diagnostic biomarkers for inflammatory bowel disease and assess the overall diagnostic accuracy of miRNAs. Condition being studied: The symptoms of inflammatory bowel disease (IBD) are highly variable. The diagnosis of IBD must be made through medical history, physical, laboratory, radiologic, endoscopic, and histological examinations. However, these diagnostic techniques are not specific and sometimes even equivocal. Therefore, reliable biomarkers are urgently needed in the diagnosis of IBD. Several clinical and preclinical researches have shown that dysregulated microRNAs (miRNAs) play a crucial role in IBD development. miRNAs, as single-stranded noncoding RNAs that contain 22-24 nucleotides, can post-transcriptionally regulate gene expression by blocking mRNA translation or degrading target mRNAs. miRNAs are widely involved in physiological and pathological cellular processes, such as differentiation, proliferation and apoptosis. Besides, they are stable, noninvasive, and resistant to degradation by ribonucleases, making them valuable targets in the diagnosis, monitoring, prognosis, and treatment of diseases. To date, inconsistent results have been found about miRNA expression profiling in the patients with IBD. Moreover, the diagnostic accuracy of miRNAs for IBD has not been reported in any meta-analysis.
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Barg, Rivka, Erich Grotewold et Yechiam Salts. Regulation of Tomato Fruit Development by Interacting MYB Proteins. United States Department of Agriculture, janvier 2012. http://dx.doi.org/10.32747/2012.7592647.bard.

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Background to the topic: Early tomato fruit development is executed via extensive cell divisions followed by cell expansion concomitantly with endoreduplication. The signals involved in activating the different modes of growth during fruit development are still inadequately understood. Addressing this developmental process, we identified SlFSM1 as a gene expressed specifically during the cell-division dependent stages of fruit development. SlFSM1 is the founder of a class of small plant specific proteins containing a divergent SANT/MYB domain (Barg et al 2005). Before initiating this project, we found that low ectopic over-expression (OEX) of SlFSM1 leads to a significant decrease in the final size of the cells in mature leaves and fruits, and the outer pericarp is substantially narrower, suggesting a role in determining cell size and shape. We also found the interacting partners of the Arabidopsis homologs of FSM1 (two, belonging to the same family), and cloned their tomato single homolog, which we named SlFSB1 (Fruit SANT/MYB–Binding1). SlFSB1 is a novel plant specific single MYB-like protein, which function was unknown. The present project aimed at elucidating the function and mode of action of these two single MYB proteins in regulating tomato fruit development. The specific objectives were: 1. Functional analysis of SlFSM1 and its interacting protein SlFSB1 in relation to fruit development. 2. Identification of the SlFSM1 and/or SlFSB1 cellular targets. The plan of work included: 1) Detailed phenotypic, histological and cellular analyses of plants ectopically expressing FSM1, and plants either ectopically over-expressing or silenced for FSB1. 2) Extensive SELEX analysis, which did not reveal any specific DNA target of SlFSM1 binding, hence the originally offered ChIP analysis was omitted. 3) Genome-wide transcriptional impact of gain- and loss- of SlFSM1 and SlFSB1 function by Affymetrix microarray analyses. This part is still in progress and therefore results are not reported, 4) Search for additional candidate partners of SlFSB1 revealed SlMYBI to be an alternative partner of FSB1, and 5) Study of the physical basis of the interaction between SlFSM1 and SlFSB1 and between FSB1 and MYBI. Major conclusions, solutions, achievements: We established that FSM1 negatively affects cell expansion, particularly of those cells with the highest potential to expand, such as the ones residing inner to the vascular bundles in the fruit pericarp. On the other hand, FSB1 which is expressed throughout fruit development acts as a positive regulator of cell expansion. It was also established that besides interacting with FSM1, FSB1 interacts also with the transcription factor MYBI, and that the formation of the FSB1-MYBI complex is competed by FSM1, which recognizes in FSB1 the same region as MYBI does. Based on these findings a model was developed explaining the role of this novel network of the three different MYB containing proteins FSM1/FSB1/MYBI in the control of tomato cell expansion, particularly during fruit development. In short, during early stages of fruit development (Phase II), the formation of the FSM1-FSB1 complex serves to restrict the expansion of the cells with the greatest expansion potential, those non-dividing cells residing in the inner mesocarp layers of the pericarp. Alternatively, during growth phase III, after transcription of FSM1 sharply declines, FSB1, possibly through complexing with the transcription factor MYBI serves as a positive regulator of the differential cell expansion which drives fruit enlargement during this phase. Additionally, a novel mechanism was revealed by which competing MYB-MYB interactions could participate in the control of gene expression. Implications, both scientific and agricultural: The demonstrated role of the FSM1/FSB1/MYBI complex in controlling differential cell growth in the developing tomato fruit highlights potential exploitations of these genes for improving fruit quality characteristics. Modulation of expression of these genes or their paralogs in other organs could serve to modify leaf and canopy architecture in various crops.
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Lapidot, Moshe, et Vitaly Citovsky. molecular mechanism for the Tomato yellow leaf curl virus resistance at the ty-5 locus. United States Department of Agriculture, janvier 2016. http://dx.doi.org/10.32747/2016.7604274.bard.

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Tomato yellow leaf curl virus (TYLCV) is a major pathogen of tomato that causes extensive crop loss worldwide, including the US and Israel. Genetic resistance in the host plant is considered highly effective in the defense against viral infection in the field. Thus, the best way to reduce yield losses due to TYLCV is by breeding tomatoes resistant or tolerant to the virus. To date, only six major TYLCV-resistance loci, termed Ty-1 to Ty-6, have been characterized and mapped to the tomato genome. Among tomato TYLCV-resistant lines containing these loci, we have identified a major recessive quantitative trait locus (QTL) that was mapped to chromosome 4 and designated ty-5. Recently, we identified the gene responsible for the TYLCV resistance at the ty-5 locus as the tomato homolog of the gene encoding messenger RNA surveillance factor Pelota (Pelo). A single amino acid change in the protein is responsible for the resistant phenotype. Pelo is known to participate in the ribosome-recycling phase of protein biosynthesis. Our hypothesis was that the resistant allele of Pelo is a “loss-of-function” mutant, and inhibits or slows-down ribosome recycling. This will negatively affect viral (as well as host-plant) protein synthesis, which may result in slower infection progression. Hence we have proposed the following research objectives: Aim 1: The effect of Pelota on translation of TYLCV proteins: The goal of this objective is to test the effect Pelota may or may not have upon translation of TYLCV proteins following infection of a resistant host. Aim 2: Identify and characterize Pelota cellular localization and interaction with TYLCV proteins: The goal of this objective is to characterize the cellular localization of both Pelota alleles, the TYLCV-resistant and the susceptible allele, to see whether this localization changes following TYLCV infection, and to find out which TYLCV protein interacts with Pelota. Our results demonstrate that upon TYLCV-infection the resistant allele of pelota has a negative effect on viral replication and RNA transcription. It is also shown that pelota interacts with the viral C1 protein, which is the only viral protein essential for TYLCV replication. Following subcellular localization of C1 and Pelota it was found that both protein localize to the same subcellular compartments. This research is innovative and potentially transformative because the role of Peloin plant virus resistance is novel, and understanding its mechanism will lay the foundation for designing new antiviral protection strategies that target translation of viral proteins. BARD Report - Project 4953 Page 2
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Gafni, Yedidya, et Vitaly Citovsky. Molecular interactions of TYLCV capsid protein during assembly of viral particles. United States Department of Agriculture, avril 2007. http://dx.doi.org/10.32747/2007.7587233.bard.

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Tomato yellow leaf curl geminivirus (TYLCV) is a major pathogen of cultivated tomato, causing up to 100% crop loss in many parts of the world. The present proposal, a continuation of a BARD-funded project, expanded our understanding of the molecular mechanisms by which CP molecules, as well as its pre-coat partner V2, interact with each other (CP), with the viral genome, and with cellular proteins during assembly and movement of the infectious virions. Specifically, two major objectives were proposed: I. To study in detail the molecular interactions between CP molecules and between CP and ssDNA leading to assembly of infectious TYLCV virions. II. To study the roles of host cell factors in TYLCV assembly. Our research toward these goals has produced the following major achievements: • Characterization of the CP nuclear shuttling interactor, karyopherin alpha 1, its pattern of expression and the putative involvement of auxin in regulation of its expression. (#1 in our list of publication, Mizrachy, Dabush et al. 2004). • Identify a single amino acid in the capsid protein’s sequence that is critical for normal virus life-cycle. (#2 in our list of publications, Yaakov, Levy et al. in preparation). • Development of monoclonal antibodies with high specificity to the capsid protein of TYLCV. (#3 in our list of publications, Solmensky, Zrachya et al. in press). • Generation of Tomato plants resistant to TYLCV by expressing transgene coding for siRNA targeted at the TYLCV CP. (#4 in our list of publications, Zrachya, Kumar et al. in press). •These research findings provided significant insights into (i) the molecular interactions of TYLCV capsid protein with the host cell nuclear shuttling receptor, and (ii) the mechanism by which TYLCV V2 is involved in the silencing of PTGS and contributes to the virus pathogenicity effect. Furthermore, the obtained knowledge helped us to develop specific strategies to attenuate TYLCV infection, for example, by blocking viral entry into and/or exit out of the host cell nucleus via siRNA as we showed in our publication recently (# 4 in our list of publications). Finally, in addition to the study of TYLCV nuclear import and export, our research contributed to our understanding of general mechanisms for nucleocytoplasmic shuttling of proteins and nucleic acids in plant cells. Also integration for stable transformation of ssDNA mediated by our model pathogen Agrobacterium tumefaciens led to identification of plant specific proteins involved.
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Chejanovsky, Nor, et Bruce A. Webb. Potentiation of Pest Control by Insect Immunosuppression. United States Department of Agriculture, janvier 2010. http://dx.doi.org/10.32747/2010.7592113.bard.

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The restricted host range of many baculoviruses, highly pathogenic to Lepidoptera and non-pathogenic to mammals, limits their use to single or few closely related Lepidopteran species and is an obstacle to extending their implementation for pest control. The insect immune response is a major determinant of the ability of an insect pathogen to efficiently multiply and propagate. We have developed an original model system to study the Lepidopteran antiviral immune response based on Spodoptera littoralis resistance to AcMNPV (Autographa californica multiple nucleopolyhedrovirus) infection and the fascinating immunosuppressive activity of polydnaviruses .Our aim is to elucidate the mechanisms through which the immunosuppressive insect polydnaviruses promote replication of pathogenic baculoviruses in lepidopteran hosts that are mildly or non-permissive to virus- replication. In this study we : 1- Assessed the extent to which and the mechanisms whereby the immunosuppressive Campoletis sonorensis polydnavirus (CsV) or its genes enhanced replication of a well-characterized pathogenic baculovirus AcMNPV, in polydnavirus-immunosuppressedH. zea and S. littoralis insects and S. littoralis cells, hosts that are mildly or non-permissive to AcMNPV. 2- Identified CsV genes involved in the above immunosuppression (e.g. inhibiting cellular encapsulation and disrupting humoral immunity). We showed that: 1. S. littoralis larvae mount an immune response against a baculovirus infection. 2. Immunosuppression of an insect pest improves the ability of a viral pathogen, the baculovirus AcMNPV, to infect the pest. 3. For the first time two PDV-specific genes of the vankyrin and cystein rich-motif families involved in immunosuppression of the host, namely Pvank1 and Hv1.1 respectively, enhanced the efficacy of an insect pathogen toward a semipermissive pest. 4. Pvank1 inhibits apoptosis of Spodopteran cells elucidating one functional aspect of PDVvankyrins. 5. That Pvank-1 and Hv1.1 do not show cooperative effect in S. littoralis when co-expressed during AcMNPV infection. Our results pave the way to developing novel means for pest control, including baculoviruses, that rely upon suppressing host immune systems by strategically weakening insect defenses to improve pathogen (i.e. biocontrol agent) infection and virulence. Also, we expect that the above result will help to develop systems for enhanced insect control that may ultimately help to reduce transmission of insect vectored diseases of humans, animals and plants as well as provide mechanisms for suppression of insect populations that damage crop plants by direct feeding.
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Sharon, Amir, et Maor Bar-Peled. Identification of new glycan metabolic pathways in the fungal pathogen Botrytis cinerea and their role in fungus-plant interactions. United States Department of Agriculture, 2012. http://dx.doi.org/10.32747/2012.7597916.bard.

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The involvement of glycans in microbial adherence, recognition and signaling is often a critical determinant of pathogenesis. Although the major glycan components of fungal cell walls have been identified there is limited information available on its ‘minor sugar components’ and how these change during different stages of fungal development. Our aim was to define the role of Rhacontaining-glycans in the gray mold disease caused by the necrotrophic fungus B. cinerea. The research was built on the discovery of two genes, Bcdhand bcer, that are involved in formation of UDP-KDG and UDP-Rha, two UDP- sugars that may serve as donors for the synthesis of cell surface glycans. Objectives of the proposed research included: 1) To determine the function of B. cinereaBcDh and BcEr in glycan biosynthesis and in pathogenesis, 2) To determine the expression pattern of BcDH and BcERand cellular localization of their encoded proteins, 3) Characterize the structure and distribution of Rha- containing glycans, 4) Characterization of the UDP-sugar enzymes and potential of GTs involved in glycanrhamnosylation. To address these objectives we generated a series of B. cinereamutants with modifications in the bchdhand bcergenes and the phenotype and sugar metabolism in the resulting strains were characterized. Analysis of sugar metabolites showed that changes in the genes caused changes in primary and secondary sugars, including abolishment of rhamnose, however abolishment of rhamnose synthesis did not cause changes in the fungal phenotype. In contrast, we found that deletion of the second gene, bcer, leads to accumulation of the intermediate sugar – UDP- KDG, and that such mutants suffer from a range of defects including reduced virulence. Further analyses confirmed that UDP-KDG is toxic to the fungus. Studies on mode of action suggested that UDP-KDG might affect integrity of the fungal cell wall, possibly by inhibiting UDP-sugars metabolic enzymes. Our results confirm that bcdhand bcerrepresent a single pathway of rhamnose synthesis in B. cinerea, that rhamnose does not affect in vitro development or virulence of the fungus. We also concluded that UDP-KDG is toxic to B. cinereaand hence UDP-KDG or compounds that inhibit Er enzymes and lead to accumulation of UDP-KDG might have antifungal activity. This toxicity is likely the case with other fungi, this became apparent in a collaborative work with Prof. Bart Thomma of Wageningen University, NETHERLANDS . We have shown the deletion of ER mutant in Verticillium dahlia gave plants resistance to the fungal infection.
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Hansen, Peter J., Zvi Roth et Jeremy J. Block. Improving oocyte competence in dairy cows exposed to heat stress. United States Department of Agriculture, janvier 2014. http://dx.doi.org/10.32747/2014.7598163.bard.

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Original Objectives. The overall goal is to develop methods to increase pregnancy rate in lactating dairy cows exposed to heat stress through methods that minimize damage to the oocyte and embryo caused by heat stress. Objectives were as follows: (1) examine the protective effects of melatonin on developmental competence of oocytes exposed to elevated temperature in vitro; (2) test whether melatonin feeding can improve developmental competence of oocytes in vivo and, if so, whether effects are limited to the summer or also occur in the absence of heat stress; and (3) evaluate the effectiveness of improving fertility by facilitating follicular turnover in the summer and winter. Revised Objectives. (1) Examine protective effects of melatonin and follicular fluid on developmental competence of oocytes exposed to elevated temperature in vitro; (2) examine the protective effects of melatonin on developmental competence of embryos exposed to elevated temperature in vitro; (3) evaluate effectiveness of improving fertility by administering human chorionicgonadotropin (hCG) to increase circulating concentrations of progesterone and evaluate whether response to hCG depends upon genotype for four mutations reported to be related to cow fertility; and (4) identify genes with allelic variants that increase resistance of embryos to heat shock. Background. The overall hypothesis is that pregnancy success is reduced by heat stress because of damage to the oocyte and cleavage-stage embryo mediated by reactive oxygen species (ROS), and that fertility can be improved by provision of antioxidants or by removing follicles containing oocytes damaged by heat stress. During the study, additional evidence from the literature indicated the potential importance of treatment with chorionicgonadotropin to increase fertility of heat- stressed cows and results from other studies in our laboratories implicated genotype as an important determinant of cow fertility. Thus, the project was expanded to evaluate hCG treatment and to identify whether fertility response to hCG depended upon single nucleotide polymorphisms (SNP) in genes implicated as important for cow fertility. We also evaluated whether a SNP in a gene important for cellular resistance to heat stress (HSPA1L, a member of the heat shock protein 70 family) is important for embryonic resistance to elevated temperature. Major conclusions, solutions & achievements. Results confirmed that elevated temperature increases ROS production by the oocyte and embryo and that melatonin decreases ROS. Melatonin reduced, but did not completely block, damaging effects of heat shock on the oocyte and had no effect on development of the embryo. Melatonin was protective to the oocyte at 0.1-1 μM, a concentration too high to be achieved in cows. It was concluded that melatonin is unlikely to be a useful molecule for increasing fertility of heat-stressed cows. Treatment with hCG at day 5 after breeding increased first-service pregnancy rate for primiparous cows but not for multiparous cows. Thus, hCG could be useful for increasing fertility in first-parity cows. The effectiveness of hCG depended upon genotype for a SNP in COQ9, a gene encoding for a mitochondrial-function protein. This result points the way to future efforts to use genetic information to identify populations of cows for which hormone treatments will be effective or ineffective. The SNP in HSPA1L was related to embryonic survival after heat shock. Perhaps, genetic selection for mutations that increase cellular resistance to heat shock could be employed to reduce effects of heat stress on fertility. Implications, both scientific and agricultural. This project has resulted in abandonment of one possible approach to improve fertility of the heat-stressed cow (melatonin therapy) while also leading to a method for improving fertility of primiparous cows exposed to heat stress (hCG treatment) that can be implemented on farms today. Genetic studies have pointed the way to using genetic information to 1) tailor hormonal treatments to cow populations likely to respond favorably and 2) select animals whose embryos have superior resistance to elevated body temperatures.
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Tzfira, Tzvi, Michael Elbaum et Sharon Wolf. DNA transfer by Agrobacterium : a cooperative interaction of ssDNA, virulence proteins, and plant host factors. United States Department of Agriculture, décembre 2005. http://dx.doi.org/10.32747/2005.7695881.bard.

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Agrobacteriumtumefaciensmediates genetic transformation of plants. The possibility of exchanging the natural genes for other DNA has led to Agrobacterium’s emergence as the primary vector for genetic modification of plants. The similarity among eukaryotic mechanisms of nuclear import also suggests use of its active elements as media for non-viral genetic therapy in animals. These considerations motivate the present study of the process that carries DNA of bacterial origin into the host nucleus. The infective pathway of Agrobacterium involves excision of a single-stranded DNA molecule (T-strand) from the bacterial tumor-inducing plasmid. This transferred DNA (T-DNA) travels to the host cell cytoplasm along with two virulence proteins, VirD2 and VirE2, through a specific bacteriumplant channel(s). Little is known about the precise structure and composition of the resulting complex within the host cell and even less is known about the mechanism of its nuclear import and integration into the host cell genome. In the present proposal we combined the expertise of the US and Israeli labs and revealed many of the biophysical and biological properties of the genetic transformation process, thus enhancing our understanding of the processes leading to nuclear import and integration of the Agrobacterium T-DNA. Specifically, we sought to: I. Elucidate the interaction of the T-strand with its chaperones. II. Analyzing the three-dimensional structure of the T-complex and its chaperones in vitro. III. Analyze kinetics of T-complex formation and T-complex nuclear import. During the past three years we accomplished our goals and made the following major discoveries: (1) Resolved the VirE2-ssDNA three-dimensional structure. (2) Characterized VirE2-ssDNA assembly and aggregation, along with regulation by VirE1. (3) Studied VirE2-ssDNA nuclear import by electron tomography. (4) Showed that T-DNA integrates via double-stranded (ds) intermediates. (5) Identified that Arabidopsis Ku80 interacts with dsT-DNA intermediates and is essential for T-DNA integration. (6) Found a role of targeted proteolysis in T-DNA uncoating. Our research provide significant physical, molecular, and structural insights into the Tcomplex structure and composition, the effect of host receptors on its nuclear import, the mechanism of T-DNA nuclear import, proteolysis and integration in host cells. Understanding the mechanical and molecular basis for T-DNA nuclear import and integration is an essential key for the development of new strategies for genetic transformation of recalcitrant plant species. Thus, the knowledge gained in this study can potentially be applied to enhance the transformation process by interfering with key steps of the transformation process (i.e. nuclear import, proteolysis and integration). Finally, in addition to the study of Agrobacterium-host interaction, our research also revealed some fundamental insights into basic cellular mechanisms of nuclear import, targeted proteolysis, protein-DNA interactions and DNA repair.
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